COVID-19

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Canadian Journal of Respiratory Therapy: CJRT = Revue Canadienne de la Thérapie Respiratoire : RCTR
. 2020; 56: 25–31.
Published online 2020 Jul 23. doi: 10.29390/cjrt-2020-015
PMCID: PMC7428000
PMID: 32844112

Low level laser therapy as a modality to attenuate cytokine storm at multiple levels, enhance recovery, and reduce the use of ventilators in COVID-19

Soheila Mokmeli, MD Anesthesiologistcorresponding author1 and Mariana Vetrici, MD, PhDcorresponding author2
1Canadian Optic and Laser Center (Training Institute), Victoria, BC, Canada
2Department of Biological Sciences, University of Lethbridge, Lethbridge, AB T1K 3M4, Canada
corresponding authorCorresponding author.
Correspondence: Soheila Mokmeli, Canadian Optic and Laser Center (Training Institute), 744A Lindsay Street, Victoria, BC V8Z 3E1, Canada. Tel.: +1 (250) 480-7868, E-mail: moc.oohay@ilemkom.rd Mariana A. Vetrici, Department of Biological Sciences, University of Lethbridge, 4401 University Drive, Lethbridge, AB T1K 3M4, Canada, Tel.: +1 (865) 888-3095, E-mail: moc.liamg@icirtevanairam
This open-access article is distributed under the terms of the Creative Commons Attribution Non-Commercial License (CC BY-NC) (http://creativecommons.org/licenses/by-nc/4.0/), which permits reuse, distribution and reproduction of the article, provided that the original work is properly cited and the reuse is restricted to noncommercial purposes. For commercial reuse, contact moc.trsc@rotide

Abstract

The global pandemic COVID-19 is a contagious disease and its mortality rates ranging from 1% to 5% are likely due to acute respiratory distress syndrome (ARDS), and cytokine storm. A significant proportion of patients who require intubation succumb to the disease, despite the availability of ventilators and the best treatment practices. Researchers worldwide are in search of anti-inflammatory medicines in the hope of finding a cure for COVID-19. Low-level laser therapy (LLLT) has strong, anti-inflammatory effects confirmed by meta-analyses, and it may be therapeutic to ARDS. LLLT has been used for pain management, wound healing, and other health conditions by physicians, physiotherapists, and nurses worldwide for decades. In addition, it has been used in veterinary medicine for respiratory tract disease such as pneumonia. Laser light with low-power intensity is applied to the surface of the skin to produce local and systemic effects. Based on the clinical experience, peer-reviewed studies, and solid laboratory data in experimental animal models, LLLT attenuates cytokine storm at multiple levels and reduces the major inflammatory metabolites. LLLT is a safe, effective, low-cost modality without any side-effects that may be combined with conventional treatment of ARDS. We summarize the effects of LLLT on pulmonary inflammation and we provide a protocol for augmenting medical treatment in COVID-19 patients. LLLT combined with conventional medical therapy has the potential to prevent the progression of COVID-19, minimize the length of time needed on a ventilator, enhance the healing process, and shorten recovery time.

Keywords: COVID-19, ARDS, cytokine storm, low level laser therapy, anti-inflammatory, ventilator, photobiomodulation

INTRODUCTION

What is low level laser therapy?

Low level laser therapy (LLLT) is also known as cold laser therapy or photobiomodulation therapy. LLLT utilizes visible light and infrared laser beams in the range of 450–1000 nm. Single wavelength or monochromatic light is emitted from a low-intensity laser diode (<500 mW). The light source is placed in contact with the skin, allowing the photon energy to penetrate tissue, where it interacts with various intracellular biomolecules to restore normal cell function and enhance the body’s healing processes []. This contrasts with the thermal effects produced by the high-power lasers that are used in cosmetic and surgical procedures to destroy tissue [], as mentioned in the PubMed Medical Subject Heading (MeSH) subheading for LLLT.

LLLT effects are not due to heat but rather to a photochemical reaction that occurs when a photoacceptor molecule within the cell absorbs a photon of light, becomes activated, and changes the cell’s membrane permeability and metabolism. Presently, cytochrome c oxidase, opsins and their associated calcium channels, and water molecules have been identified as the main mediators of the photochemical mechanisms []. This leads to increased mRNA synthesis and cell proliferation. LLLT produces reactive oxygen species (ROS) in normal cells, but ROS levels are lowered when it is used in oxidatively stressed cells, like in animal models of disease. LLLT up-regulates antioxidant defenses and decreases oxidative stress [].

Low-level lasers are a safe, noninvasive technology approved by both the US Food and Drug Administration and Health Canada for several chronic and degenerative conditions, temporary pain relief, cellulite treatment, body contouring, lymphedema reduction, hair growth, and chronic musculoskeletal injuries. LLLT increases microcirculation, lymphatic drainage, and cellular metabolism, thereby relieving many acute and chronic conditions.

The MeSH database in PubMed contains more than 7000 articles on LLLT. The effects of LLLT have been confirmed through several meta–analysis studies and include anti-inflammatory [] and analgesic effects [], tissue healing [], treating tendinopathy [], and improving lymphedema []. Recent lab and animal studies suggest LLLT is ready for clinical trials over myocardial infarction []. In 2010, a meta-analysis concluded that there was strong evidence of an anti-inflammatory effect of LLLT [].

To date, published reports indicate that LLLT up-regulates antioxidant defenses and decreases ROS in oxidatively stressed cells and animal models of disease. The anti-inflammatory effect of LLLT directly addresses the main pathology of disorders such as musculoskeletal, lungs, wounds, brain, trauma, etc. LLLT reduces NF-kB, a protein complex that controls transcription of DNA, in pathological conditions. Reports have shown reductions in reactive nitrogen species and prostaglandins in various animal models [].

LLLT has diverse effects []:

  • reduces pain related to inflammation via dose-dependent reduction of prostaglandin E2, prostaglandin-endoperoxide synthase-2, IL-1, IL-6, TNFa, as well as the cellular influx of neutrophils, oxidative stress, edema, and bleeding;

  • decreases edema and swelling by increasing lymphatic drainage;

  • increases collagen and protein production, and cell proliferation;

  • accelerates wound healing and scar formation;

  • improves quality and tensile strength of tissue;

  • stimulates nerve function and regeneration;

  • accelerates bone regeneration and remineralization;

  • reduces the pain threshold and enhances endorphins;

  • washes inflammatory debris away from the injured site; and

  • augments blood flow.

LLLT has been used in respiratory tract diseases since 1978. Empirical practice on over 1000 patients produced data pertaining to chronic pneumonia, acute pneumonia, asthma, and chronic bronchitis in children, adults, and elderly. Common findings include reduced chest pain and heaviness; normalization of respiratory function; improved blood, immunological, and radiological parameters; and shortened recovery times. In community-acquired pneumonia, intravenous LLLT of blood added to conventional treatment significantly promoted the bactericidal activity of neutrophils. In asthma, the addition of LLLT was more effective than medical treatment alone and it shortened the duration of treatment and recovered bronchial sensitivity to sympathomimetics []. In newborns with pneumonia, LLLT combined with conventional medical regimens optimized the treatment infectious and inflammatory diseases, reduced the incidence of complications, and shortened recovery periods [].

LLLT is a well-known treatment modality in veterinary medicine. Upper and lower respiratory conditions in dogs and cats are common, and viral and bacterial infections are often highly contagious. Regardless of etiology, inflammation is the major pathology of these conditions. The addition of LLLT to conventional treatment alleviates symptoms and stimulates the healing process in tissues. General guidelines for the use of laser therapy in animals and protocols for specific conditions are published [].

The pathogenesis of COVID-19 in respiratory tract

Coronaviruses are a large group of viruses that affect animals. In humans, they produce diseases such as the common cold, severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome. The disease caused by the novel coronavirus, SARS-CoV-2, has been named COVID-19 and the clinical manifestations range from asymptomatic to severe acute respiratory distress syndrome (ARDS) to death [].

Respiratory viruses infect either the upper or lower airways. Typical upper-respiratory infections are milder, more contagious, and spread easily, whereas lower-respiratory infections spread much less frequently but are more severe and dangerous. SARS-CoV-2 appears to infect both upper and lower airways. It spreads while still limited to the upper airways, before traveling into the deeper respiratory tract and leading to severe symptoms [].

SARS-CoV-2 attaches to a protein called angiotensin converting enzyme (ACE2), on the surface of cells in the respiratory tract. As SARS-CoV-2 attacks the cells, dead cells flow down and block the airways with debris while the virus moves deeper into the lungs. Breathing becomes difficult because the lungs become clogged with dead cells and fluid. The immune system attacks the virus causing inflammation and fever. In severe cases, the immune system goes wild, causing more damage to the lungs than the actual virus. Blood vessels dilate to increase blood flow and become more permeable to maximize transport of chemical and cellular mediators the infection site. Inevitably, the lungs get filled with fluid. This exaggerated immune response is called cytokine storm and it leads to ARDS, fever, multiorgan failure, and death [].

During cytokine storm, the immune system attacks indiscriminately without clearing the specific targets. Cytokine storm also affects other organs, especially if people already have chronic diseases []. The severity of cytokine storm determines who is hospitalized and who will be treated in the intensive care unit (ICU). The classification of COVID-19 is summarized in Table 1 [].

Table 1

The staging and classification of COVID-19 []
Class Symptoms Imaging Respiratory criteria
Mild infection Mild Negative signs of pneumonia Normal
Moderate infection Fever and upper respiratory tract symptoms Positive signs of pneumonia Normal
Severe infection Fever, upper and lower respiratory tract symptoms >50% lesion progression within 24–48 hours Respiratory rate ? 30 /min
O2 saturation ? 93% at rest
Arterial partial pressure of O2 (PaO2)/oxygen concentration (FiO2) ? 300 mm Hg
Critical infection Respiratory failure requiring mechanical ventilator and (or) presence of shock and (or) other organ failure that requires monitoring and (or) treatment in the ICU > 50% lesion progression within 24–48 hours Early stage:

  • Oxygenation index 100.1–149.9 mmHg.

  • Respiratory system compliance (RSC) ? 30 ml/cmH2O.

  • No organ failure other than the lungs.

Middle stage:

  • 60 mmHg < O2 index ? l00 mmHg.

  • 30 mL/cmH2O > RSC ? 15 mL/cmH2O.

  • Maybe complicated by mild or moderate dysfunction of other organs.

Late stage:

  • O2 index ? 60 mmHg.

  • RSC < 15 mL/cmH2O.

  • Diffuse consolidation of both lungs that requires the use of extracorporeal membrane oxygenation or failure of other vital organs.

Note: A confirmed case is based on the epidemiological history (including cluster transmission), clinical symptoms (fever and respiratory symptoms), lung imaging, and results of SARS-CoV-2 nucleic acid detection and serum-specific antibodies [].

The morbidity and mortality of COVID-19 are due to excessive inflammatory cytokine production and immune hyperactivity. Alveolar macrophage activation and cytokine storm are the main pathogenesis of severe COVID-19. The pathological features include exudation and hemorrhage, epithelial injuries, infiltration of macrophages into the lungs, and fibrosis of lung tissue. The mucous plug with fibrinous exudate in the alveoli and the activation of alveolar macrophage are characteristic abnormalities []. Chemical and genetic studies have shown that the pulmonary endothelium is a key component of the cytokine storm. Therefore, modulation of the involved cellular signaling pathways may have therapeutic effects [].

COVID-19 begins when SARS-CoV-2 uses ACE2 as the entry receptor for infection []. This induces ACE2 downregulation and shedding. Loss of ACE2 from the endothelium causes dysfunction of the renin-angiotensin system, and it enhances inflammation and vascular permeability. Shedding of ACE2 from the endothelium releases enzymatically active soluble ACE2 (sACE2), which is tightly linked to tumor necrosis factor alpha (TNF-?) production in cell culture [].

Multiple signaling pathways are activated during an immune response and cytokine storm. The P2X purinoceptor 7 (P2X7r) is major factor involved in activation of the cytokine storm and lung pathology in response to viruses [], infection, inflammation, hypoxia, or trauma []. P2X7r is an adenosine triphosphate (ATP) gated, nonselective cation channel, allowing Ca2+ and Na+ influx and K+ efflux. Extracellular ATP plays a central role in apoptotic cell death [], the induction of inflammation [], and mitochondrial failure in monocytes []. P2X7r mediates ATP-induced cell death in different cells and it promotes assembly and release of proinflammatory interleukins (IL-1? and IL-18) from immune cells after exposure to lipopolysaccharide and ATP []. P2X7r is constitutively expressed in many cells, including respiratory epithelial cells and most immune cells like neutrophils, monocytes, macrophages, dendritic, natural killer, B and T lymphocytes [].

Studies stratified COVID-19 patients as: (i) severe symptoms and ICU admission and (ii) mild and moderate symptoms requiring hospitalization but not ICU []. The severe patients have significantly higher levels of plasma pro-inflammatory factors (IL-2, IL-7, IL-10, GSCF, IP-10, MCP-1, MIP1A, TNF-?) [] and (IL-2, IL-6, IL-10, TNF-?) [] than non-ICU patients, and they were likely in cytokine storm []. These findings justify the use of IL-6 receptor antagonists []; however, a therapy to reduce inflammation at multiple levels, such as LLLT, could be more successful in controlling the unbalanced immune response (Figure 1).

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The effects of SARS-CoV-2 on alveolar cell and cytokine storm.

The effects of LLLT on pulmonary inflammation

LLLT is effective against cytokine storm and ARDS while promoting healing and tissue regeneration. Experimental and animal models of pulmonary disease and infection have revealed multiple cellular and molecular effects, which are both local and systemic. LLLT reduces inflammation without impairing lung function in acute lung injuries and is a promising therapeutic approach for lung inflammatory diseases such as Chronic obstructive pulmonary disease [].

In murine models of acute inflammation of the airways and lungs, transcutaneous LLLT delivered over the trachea decreases pulmonary microvascular leakage [], IL-1b levels [], IL-6 [], MIP-2 mRNA expression [], and intracellular ROS production []. LLLT produces anti-inflammatory effects on tracheal hyperactivity, and reduces neutrophil influx [] by inhibiting COX-2-derived metabolites []. In ARDS, LLLT elevates cyclic adenosine monophosphate [], a signaling molecule that stimulates IL-10 and G-CSF expression and blocks TNF-a and MIP-1. LLLT also reduces TNF-a levels in bronchoalveolar lavage fluid and alveolar macrophages []. In hemorrhagic lesions of the lungs, LLLT significantly reduces the hemorrhagic index and myeloperoxidase activity, to levels comparable to Celecoxib [].

LLLT contributes to the resolution of inflammation by upregulating IL-10 and downregulating P2X7r. LLLT changes the profile of inflammatory cytokines and elevates IL-10 [], known as human cytokine synthesis inhibitory factor, in the lung and abolishes lung inflammation via a reduction of inflammatory cytokines and mast cell degranulation []. LLLT decreases collagen deposition as well as the expression of the P2X7r [].

LLLT contributes to healing by promoting apoptosis of inflammatory cells while suppressing apoptotic pathways in lung tissue. In a model of acute lung injury, LLLT reduced DNA fragmentation and apoptotic pathways via increased B-cell-lymphoma-2 (Bcl-2), the key regulator of the intrinsic or mitochondrial pathway for apoptosis, in alveolar epithelial cells while promoting DNA fragmentation in inflammatory cells []. In pulmonary idiopathic fibrosis, LLLT inhibits pro-inflammatory cytokines and increases expression of proliferating cell nuclear antigen [], attenuates airway remodeling by balancing pro- and anti-inflammatory cytokines in lung tissue, and inhibiting fibroblast secretion of the pro-fibrotic cytokines [].

LLLT provides synergy in combination with medical treatment. It has a synergic anti-inflammatory action over alveolar macrophages pretreated with N-acetyl cysteine, an effective oral medicine for coughs and some lung conditions []. The synergic effects of LLLT combined with conventional treatments were reported on over 1000 patients in Russian studies [].

Extended time on ventilators causes lung injury but LLLT minimizes this side effect. In experimental models of ventilator-induced lung injury (VILI), LLLT following VILI resulted in lower injury scores, decreased total cell count and neutrophil count in bronchoalveolar lavage, and reduced alveolar neutrophil infiltration. LLLT in an experimental model of VILI in rats demonstrated the anti-inflammatory effect via decreased lung injury scores and lower counts of neutrophils in alveolar, interstitial, and bronchial lavage [] (Figure 2).

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The effects of SARS-CoV-2 versus LLLT on cytokine storm and lung tissue.

Evidence from the literature supports the use of LLLT for the treatment of COVID-19.

  • It has significant anti-inflammatory effects confirmed by meta–analyses. Eleven cell studies, 27 animal studies, and another six animal studies for drug comparisons and LLLT interactions verified that there is strong evidence of an anti-inflammatory effect of LLLT. The scale of the anti-inflammatory effect is not significantly different than non-steroidal anti-inflammatory drugs, but it is slightly less than glucocorticoid steroids [].

  • It has diverse applications and effects confirmed through several meta-analysis studies include analgesia [], tissue healing [], treating tendinopathy [], and improved lymphedema [].

  • LLLT is approved by the US FDA and Health Canada for several chronic and degenerative conditions, temporary pain relief, cellulite treatment, body contouring, lymphedema reduction, and hair growth. It has been used in veterinary medicine for upper and lower respiratory conditions in dogs and cats [].

  • It has been used for human respiratory tract disease. Empirical use on over 1000 patients produced data pertaining to chronic pneumonia, acute pneumonia, asthma, and chronic bronchitis in children, adults, and the elderly []. Light therapy and LLLT has been mentioned as a potential treatment for pandemic coronavirus infections [].

  • The anti-inflammatory effect of LLLT in lung inflammation is confirmed in at least 14 experimental animal studies. LLLT attenuates cytokine storm at multiple levels and reduces the major inflammatory metabolites such as IL-6 and TNF-?. IL-6 antagonists are being investigated for treating COVID-19 but LLLT reduces the production of IL-6, as well as other chemokines and metabolites [].

  • There are US FDA and Health Canada approved laser machines for pain management, lymphedema after breast cancer surgery, and cellulite treatments that can be used and set to treat lung inflammation.

  • LLLT is an affordable modality compared with other treatments and medicines like IL-6 antagonists. LLLT is a safe, effective, low-cost modality without any reported side-effects compared with other approaches. A laser machine costs Can$35,000.00–200,000.00, and each machine can fully treat 20,000 patients for COVID-19. In comparison, an IL-6 antagonist costs US$1000.00 per injection, and each patient would need 3–6 injections for complete COVID-19 treatment. Treating 20,000 patients would cost US$ 60,000,000.00–US$ 120,000,000.00.

Based on this information, LLLT will accelerate recovery from COVID-19 and will get patients off ventilator support and out of the ICU more rapidly. This could significantly decompress our severely overburdened health care systems.

Therapeutic technique and dosage of LLLT

Laser dose is the amount of energy delivered per second per cm2. The effect of laser therapy is related to the amount of laser energy per cm2. The Arndt-Schultz Law is considered the standard to describe the dose dependent effects of LLLT []. The minimum therapeutic dose for a bio-stimulatory effect for red and infrared laser is 0.01 J/cm2 while for ultraviolet, blue, green laser it is 0.001 J/cm2. LLLT has a noticeable biphasic dose response. The effective stimulation dose is 1 J/cm2 on the target tissue. Doses greater than 10 J/cm2 produces inhibitory effects. The inhibitory effects are used in conditions requiring inhibition and suppression [].

Therapeutic protocol: early phase of COVID-19: (Figure 3Table 2)

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LLLT for COVID-19.

Table 2

Therapeutic protocol: Early phase of COVID-19
Laser system parameters
Wavelengths Infrared laser (780–900 nm), or red laser (630–660 nm)
Average power 50–100 mW
Dose 4–6 J/cm2
Area 10 cm2
Sessions 3–8 once-daily sessions
Laser probe positions
Intranasal:
1 minute/cm2 (100 mW)
2 minutes/cm2 (50 mW)
Noncontact technique
Over right and left tonsils
1 minute/cm2 (100 mW)
2 minutes/cm2 (50 mW)
Transcutaneous (place laser over the skin)
Over the trachea
1 minute/cm2 (100 mW)
2 minutes/cm2 (50 mW)
Transcutaneous
Over the veins in the cubital areas
8 minute/cm2 (100 mW)
15 minutes/cm2 (50 mW)
Transcutaneous blood laser therapy

Laser parameters:

  • Laser type: infrared laser (780–900 nm), or red laser (630–660 nm)

  • Average power: 50–100 mW

  • Dose: 4–6 J/cm2

  • Area: 10 cm2

  • Time: 1–2 minutes/cm2

  • Sessions: 3–8 once-daily sessions

Laser probe positions:

  • Intranasal: 2 minutes, noncontact technique

  • Over right and left tonsils: transcutaneous (place laser over the skin)

  • Over the trachea: transcutaneous

  • Over the veins in the cubital areas: transcutaneous blood laser therapy, 10–15 minutes

Therapeutic protocol: medium–severe phase of COVID-19: (Figure 3Table 3)

Table 3

Therapeutic protocol: medium–severe phase of COVID-19
Laser system parameters
Wavelengths Infrared laser (780–900 nm), or red laser (630–660 nm)
Average power 50–100 mW
Dose 6–10 J/cm2
Area 10 cm2
Sessions 3–10 once-daily sessions
Laser probes positions
Intranasal:
1 minute/cm2 (100 mW)
2 minutes/cm2 (50 mW)
Noncontact technique
Over right and left tonsils
1 minute/cm2 (100 mW)
2 minutes/cm2 (50 mW)
Transcutaneous (place laser over the skin)
Over the trachea
1 minute/cm2 (100 mW)
2 minutes/cm2 (50 mW)
Transcutaneous
Over the veins in the cubital areas
8 minute/cm2 (100 mW)
15 minutes/cm2 (50 mW)
Transcutaneous blood laser therapy
Over the lungs
1:30–2 minute/cm2 (100 mW)
2–3 minutes/cm2 (50 mW)
Bilaterally over apical, middle, and lower lobes, front and back of thorax; transcutaneous over the intercostal spaces
Over the bronchus
1:30–2 minute/cm2 (100 mW)
2–3 minutes/cm2 (50 mW)
Upper mediastinal area: transcutaneous

Laser parameters:

  • Laser type: infrared laser (780–900 nm) or red laser (630–660 nm)

  • Average power: 50–100 mW

  • Dose: 6–10 J/cm2

  • Area: 10 cm2

  • Time: 2–3 minutes/cm2

  • Sessions: 3–10 once-daily sessions

Laser Probe Positions:

  • Over the lungs: bilaterally over apical, middle, and lower lobes and front and back of thorax, transcutaneous over the intercostal spaces

  • Over the trachea: transcutaneous

  • Over the bronchus: upper mediastinal area, transcutaneous

  • Over right and left tonsils: transcutaneous

  • Over the veins in the cubital areas: transcutaneous blood laser therapy; 10–15 minutes

Contraindications and side effects of LLLT []

Although LLLT is safe and noninvasive and there are no reports of mutagenicity, genotoxicity, or carcinogenicity of LLLT after 60 years of its use. However, there are some contraindications:

  • work over the site of tumors and cancer;

  • benign tumors with possibility of converting to malignant tumors;

  • the first 3 months of pregnancy (in the second and third trimesters, avoid work on abdominal and spine area); and

  • light sensitivity conditions.

Precautions []

  • epiphyseal line in children;

  • glands: avoid ovaries, testes;

  • in patients with severe end organ damage: heart, kidney, liver, and lung;

  • epilepsy: the possibility of nerve discharge is increased in LLLT, especially with low-frequency protocols, 5–10 HZ.

Side effects of LLLT

Optical side effects

Because of the high intensity of lasers and the absorption of its wavelengths by different parts of ocular system, there is a possibility of damage to the eyes. It is important to use protective glasses that can absorb the specific wavelength. Protective glasses for each wavelength are different; therefore, choose the protective goggles specified for each wavelength. Both therapists and clients should wear protective goggles [].

Early sense of healing

The analgesic effect of laser manifests earlier than its healing effect, and the patients feel better because of this, but the actual tissue damage has not yet healed. Patients feel relaxed and more energetic because the pain is gone. However, they must allow enough time for recovery [].

Fatigue and tiredness

Fatigue is the most common symptom following LLLT. This is due to hormonal and metabolite changes after laser therapy that increase expression natural pain killers like endorphins and enkephalins. These metabolites induce relaxation and sleepiness [].

Low blood pressure and dizziness

Very rarely, when the treated area is close to large blood vessels, a patient may experience a temporary drop in the blood pressure and orthostasis. This is due to vasodilatation and increased circulation to the limbs. To avoid dizziness, it is recommended that patients drink fluids before LLLT, and then wait for a few minutes before getting up from the supine position [].

CONCLUSION

COVID-19 is potentially lethal because of cytokine storm and ARDS. Although most patients who contract COVID-19 may recover at home, a significant proportion require hospitalization and (or) ICU treatment. Many of the patients that are placed on ventilators succumb to the disease despite the best treatment practices. Often, patients are maintained on ventilators for longer than expected, and this may contribute to ventilator induced lung injury while depleting the patient’s convalescent resources. Modulation of inflammatory factors and a boost to healing are necessary to help patients come off the ventilators. LLLT is a safe and noninvasive modality that has been used for decades in pain management, wound healing, and health conditions including diseases of the respiratory tract. LLLT was combined successfully with standard medical care to optimize response to treatments, reduce inflammation, promote healing, and accelerate recovery times. Scientific evidence shows that LLLT attenuates the inflammatory cytokines and chemokines in cytokine storm at multiple levels. In addition, LLLT promotes apoptosis of inflammatory cells and protects alveolar cells from damage. These findings suggest that LLLT is a feasible modality in the treatment of ARDS. LLLT can be added to the conventional treatment in COVID-19 at different stages of the disease. Because of its anti-inflammatory effect, and ability to shorten recovery times, LLLT can reduce the need of ventilators in the healing process. Clinical trials are necessary to objectively evaluate the effect of LLLT on COVID-19 treatment and recovery.

Contributors

Soheila Mokmeli and Mariana Vetrici contributed to the conception and design of the work.

Competing interests

All authors have completed the ICMJE uniform disclosure form at www.icmje.org/coi_disclosure.pdf and declare: no financial relationships with any organizations that might have an interest in the submitted work in the previous 3 years; no other relationships or activities that could appear to have influenced the submitted work.

Ethical approval

Informed consent was obtained from all participants.

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Articles from Canadian Journal of Respiratory Therapy: CJRT = Revue Canadienne de la Thérapie Respiratoire : RCTR are provided here courtesy of Canadian Society of Respiratory Therapy

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. 2020 Aug 19 : 111999.
doi: 10.1016/j.jphotobiol.2020.111999 [Epub ahead of print]
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Light-based technologies for management of COVID-19 pandemic crisis

aDepartment of Clinical Analysis, Faculty of Pharmaceutical Sciences, University of São Paulo, SP, Brazil
bBioLambda, Scientific and Commercial LTD, São Paulo, SP, Brazil
cGAMA Therapeutics LLC, Massachusetts Biomedical Initiatives, Worcester, USA
dDepartment of Biochemistry, Institute of Chemistry, University of São Paulo, São Paulo, SP, Brazil.
eWellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA
fVaccine and Immunotherapy Center, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA
gLaser Research Centre, Faculty of Health Science, University of Johannesburg, Doornfontein, South Africa
hCenter for Lasers and Applications, Nuclear, and Energy Research Institute, National Commission for Nuclear Energy, São Paulo, SP, Brazil
iDepartment of Chemistry Rice University, Houston, TX, USA
jIdISBA – Fundación de Investigación Sanitaria de las Islas Baleares, Palma, Spain
kDepartment of Internal Medicine, School of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo, SP, Brazil
lSchool of Veterinary Medicine, Metropolitan University of Santos, Santos, Brazil.
mMicromoria LLC, Marlborough, USA
nSchool of Pharmacy and Biomolecular Sciences, Liverpool John Moores University, Liverpool, UK
Caetano P. Sabino: moc.adbmaloib@onateacMauricio S. Baptista: rb.psu.qi@atsitpab
?Corresponding author. rb.psu.qi@atsitpab
??Corresponding author at: Department of Clinical Analysis, Faculty of Pharmaceutical Sciences, University of São Paulo, SP, Brazil. moc.adbmaloib@onateac
1All authors contributed equally to the manuscript
Since January 2020 Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company’s public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre – including this research content – immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active.

Abstract

The global dissemination of the novel coronavirus disease (COVID-19) has accelerated the need for the implementation of effective antimicrobial strategies to target the causative agent SARS-CoV-2. Light-based technologies have a demonstrable broad range of activity over standard chemotherapeutic antimicrobials and conventional disinfectants, negligible emergence of resistance, and the capability to modulate the host immune response. This perspective article identifies the benefits, challenges, and pitfalls of repurposing light-based strategies to combat the emergence of COVID-19 pandemic.

Keywords: photoinactivation, ultraviolet, photodynamic, photobiomodulation, germicidal, virucidal, photobiology

1. Introduction

The pandemic spread of the novel coronavirus disease (COVID-19), caused by the SARS-CoV-2 virus, is a red-alert global health threat [,]. In December 2019, COVID-19 expanded from Wuhan throughout China and was then exported throughout the world []. So far, more than 10 million people have been diagnosed with COVID-19 infection, and many more are expected to be diagnosed within the coming months [,]. As the epidemic evolves, national and global organizations are facing an urgent need to coordinate and combat this unprecedented large-scale public health crisis [].

The epidemiological features of COVID-19 (i.e., severity, full spectrum of disease, transmissibility) have not been fully dissected []. The consensus is that the risk for severe acute disease symptoms and death is higher among the elderly and the immunocompromised []. In severe cases, infected patients need to be transferred to intensive care units for tracheal intubation []. This phenomenon is particularly worrisome because it can overwhelm healthcare facilities during the epidemic peak [].

The spread and persistence of SARS-CoV-2 in diverse environments, such as healthcare, community, and residential areas, underlines the urgency for developing effective decontamination approaches as the pandemic crisis evolves []. A successful disinfection strategy coupled with additional infection-prevention countermeasures may substantially reduce transmissibility from asymptomatic carriers, a feature that is considered pivotal in the rapid dissemination of SARS-CoV-2. New light-mediated disinfection protocols are currently validated in hospitals and healthcare facilities for surface, air, and water as well as personal protective equipment (PPE), including eyewear, N95 respirators, and masks. Additionally, photobiomodulation, a light-based anti-inflammatory therapy, may have some palliative effects on patients suffering from severe COVID-19. This review examines the potential of light-based technologies to prevent COVID-19 infection and control its dissemination by direct viral inactivation and to treat COVID-19 by modulating the host immune system. The direct antimicrobial actions of solar and UV radiation, photodynamic therapy, antimicrobial blue light, and ultrafast pulsed lasers for disinfection or in vivo use are considered, and the application of photobiomodulation to stimulate the host to mount an anti-viral response is discussed.

2. SARS-CoV-2 Stability Outside The Human Body

SARS-CoV-2 is highly infectious [] and transmission occurs through contaminated air, water, and surfaces, which plays a pivotal role in its unbridled dissemination. A recent study by van Doremalen and colleagues investigated the stability of SARS-CoV-2 in aerosols and on inanimate surfaces (e.g., glass, metal, plastic, or cardboard) that can act as important transmission vectors []. Their findings suggest that aerosol and fomite transmission of SARS-CoV-2 is likely, indicating that the virus can remain viable and infectious for hours in aerosols and up to days on surfaces. This is in agreement with a recent comparative analysis of 22 studies looking at the persistence of a broader panel of human coronaviruses on inanimate surfaces [] This study included prominent pathogenic coronavirus species such as Severe Acute Respiratory Syndrome (SARS), Middle East Respiratory Syndrome (MERS) and endemic human coronaviruses (HCoV) and concluded that: 1) viruses can remain infectious from 2 h to 9 days; 2) incubation temperature is critical, as some viruses can remain viable at 4 °C for up to 28 days whereas at 30–40 °C viral viability is reduced.

3. Historical Milestones of Antimicrobial Light

The microbicidal effects of light have been widely known for more than a century. In 1885, Duclaux experimented with several microbial species and concluded that “sunlight is the best, cheapest, and most universally applicable microbicidal agent that we have” []. As early as 1877, Downes and Blunt observed that light could effectively kill a series of microorganisms and reported that this effect was dependent on light parameters such as intensity, duration (i.e., light dose) and that the shortest wavelengths (e.g., blue to ultraviolet light) were the most effective [] The first report on the virucidal effects of UV radiation dates back to 1928 when Rivers and Gates used UV light to inactivate viral particles in suspension and proved the efficacy of the method through subsequent subcutaneous inoculation of rabbits [].

In 1903, Niels Finsen was awarded the Nobel Prize in Physiology or Medicine for his contribution to the treatment of infectious diseases, especially cutaneous tuberculosis, using visible light [,]. Virtually at the same time, Herman Von Tappeiner and Oscar Raab discovered by accident that the use of fluorescent dyes could enhance the microbial killing effect of visible light via photodynamic reactions []. By the 1930s, germicidal low-pressure Mercury (Hg) discharge lamps emitting quasi-monochromatic UV-C light (peak emission at 254 nm) had been introduced into the market as highly efficient disinfection equipment []. Thus, since the pre-antibiotic era, light-based strategies have been extensively studied and used to treat and prevent infections []. However, each photoinactivation strategy has its pros and cons that must be carefully considered when designing a new microbial control strategy.

4. Natural Ultraviolet Light

Ultraviolet (UV) radiation is naturally and ubiquitously emitted by the sun, representing 10% of its total light output. Only a small portion of the sunlight spectra has direct antimicrobial properties (UV-C). However, since most UV-C light is filtered by the atmospheric ozone layer, in practical terms, the antimicrobial activity associated with sunlight is mostly caused by photochemical reactions induced by UV-A and UV-B photons which are absorbed by endogenous chromophores such as amino acid residues, flavins, and porphyrin derivatives []. While UV-A alone does not seem to exert any significant virucidal effects, natural and artificial sunlight, as well as radiation in the UV-B spectrum, have been shown to inactivate bacteriophages and human viruses []. A model for the potential of solar UV radiation to inactivate viruses aerosolized in the atmosphere concluded that a full day of sun exposure would on average decrease the infectivity of UV-sensitive viruses by 3 log10 [].

Besides its virucidal potential, solar UV radiation can also play a protective role against infectious diseases via its modulating effect on vitamin D production []. Vitamin D is known to upregulate the production of human cathelicidin, LL-37. This antimicrobial peptide has both antimicrobial and antiendotoxin activities, and also attenuates the production of proinflammatory cytokines which typically accompany respiratory tract infections. Accordingly, it was recently suggested that vitamin D could reduce the incidence, severity, and risk of death due to respiratory tract infections, notably those caused by COVID-19 []. However, conclusive evidence for an association between vitamin D supplementation and decreased risk of respiratory tract infections is still lacking.

UV-C is directly absorbed by pyrimidine bases causing their dimerization, which leads to viral inactivation via DNA or RNA damage []. Thymine is the main chromophore in DNA while uracil is its counterpart in RNA. Upon UV-C exposure, thymine and uracil form cyclobutane-dimers and pyrimidine-protein cross-links []. It must be stressed that UV-C usage must be limited to inanimate objects since it is highly dangerous to human skin. The viral protein coat has been shown to protect nucleic acids from UV-C radiation, by shielding the RNA, quenching the excited states of RNA, and/or by surrounding the bases with a hydrophobic environment and limiting the mobility of the individual bases. This results in a reduction of the overall rate of photoreactions, which allows the formation of non-cyclobutane-type dipyrimidines and uridine hydrates. Viral coating proteins themselves may suffer UV photodamage and become cross-linked to RNA.

The International Ultraviolet Association (IUVA) recently released a fact sheet detailing the efficacy of UV on SARS-CoV-2 [] in which they reviewed all the appropriate requirements for the safety of UV-C disinfection devices and discussed the corresponding performance standards and validation protocols. Coronaviruses display a wide range of UV-C LD90 (UV-C dose necessary to inactivate 90% of a microbial population) values, from 7 to 241 J/m2 so one might assume that the UV-C susceptibility of the novel SARS-CoV-2 (COVID-19) virus probably lies within this range []. Therefore, based on previous studies with SARS-CoV-1 and other RNA-based coronaviruses, UV-C light can be used to effectively inactivate such pathogens present in the air, liquids and over several surfaces [,].

5. Ultraviolet Germicidal Irradiation (UVGI)

UV-C lamps have long been used in hospital and industrial settings for decontamination purposes. In the context of a mitigation approach to infection spreading, UV-C can be particularly helpful in the inactivation of virus-containing aerosols and surfaces.

Air disinfection via upper-room germicidal UV-C light fixtures may be able to reduce viral transmission via the airborne route. Accordingly, an observational study during the 1957 influenza pandemic reported that patients housed in hospital wards with upper-room UV-C had an infection rate of 1.9%, compared to an infection rate of 18.9% among patients housed in wards without UV-C []. However, it is important to note that the germicidal effect of UV-C seems to be strongly dependent on the relative humidity of the air, with UV-C effectiveness against influenza virus decreasing with increasing relative humidity [].

The potential of viral spreading via contaminated surfaces depends on the ability of the virus to maintain infectivity in the environment, which in turn is influenced by several biological, physical, and chemical factors, including the type of virus, temperature, relative humidity, and type of surface []. Importantly, single-stranded nucleic acid (ssRNA and ssDNA) viruses were more susceptible to UV inactivation than viruses with double-stranded nucleic acid (dsRNA and dsDNA). Also, the UV dose necessary to achieve the same level of virus inactivation at 85% relative humidity (RH) was higher than that at 55% RH [].

In a recent study, Fischer et al. showed that UV-C light can inactivate more than 99.9% of SARS-CoV-2 viral particles deposited over the filtering material of N95 masks and stainless steel surface []. As expected, inactivation kinetics over stainless steel was much faster (i.e., more than 99.9% for (0.33 J/cm2). However, after sufficient exposure (1.98 J/cm2) UV-C could promote germicidal efficacy levels that were similar to those promoted by ethanol, dry heat or vaporized hydrogen peroxide. Older studies have hypothesized that the necessary dose to inactivate 90% of viruses present in N95 filtering facepiece respirator (FFR) material would be about 30 times higher than over the surface of non-porous materials []. This was an interesting estimation, but we should keep in mind that UV-C emission spectrum and irradiance of different UV-C equipment as well as material composition are widely variable []. Therefore, such estimatives cannot be used as a robust procedure and experimental demonstrations must always be presented. Indeed, a recent in silico study demonstrated that for effective and fast decontamination one should consider the FFR shape besides the optical properties of the FFR model, which has to be determined at the UV-C specific wavelength []. Even though UV does not seem to affect the filtrating capacity of FFRs, it is important to note that high UV-C doses can lead to reduced tensile strength of its materials [,].

The combination of multiple light wavelengths has been explored for cosmetic, environmental (water disinfection) and clinical (microbial catheter disinfection) applications. However, the precise photobiological mechanism of action and the experimental workflow to develop a marketable application is still missing [,].

It must be remarked that UV-C light at 254 nm is harmful to the eyes and skin and, therefore, it is recommended to use it in setups that avoid direct human exposure. Although, far-UV-C (207–222 nm) has been proposed as a disinfection technology that seems to be safer to human exposure []. This has been claimed because far-UV-C range is strongly absorbed by amino acid residues and, therefore, is further blocked by the acellular stratum corneum of the skin and the cornea of the eye, leading to lower levels of UV-C light reaching the cellular layers of eyes and skin. However, as far as our knowledge goes, robust studies showing the actual safety of far-UV-C towards animal tissues in short and long terms have not been strongly established and degradation of proteins can also lead to serious eye and skin damages. Thus, we can only recommend UV-C application to inanimate objects. Additionally, far-UV-C technology is not broadly available in the market yet and the cost is far higher than common LP-Hg lamps. On the other hand, UV-C LED technology is limited to very compact applications. The shortest wavelengths available are around 255 nm, with the price per Watt being up to 1000 times higher than that of LP-Hg lamps, while displaying an energy efficiency (< 1%) far lower than that of LP-Hg lamps (25–40%) at 254 nm.

6. Photoantimicrobials and Photodynamic Therapy

Visible light can exert antiviral effects via photodynamic mechanisms that are initiated upon absorption of light by exogenous photosensitizer compounds, such as phenothiazinium salts, porphyrins, nanoparticles, and others []. The inactivation of microorganisms and viruses by visible light is based on the generation of lethal oxidant species via photosensitized oxidation reactions, which usually require three components: the chromophore, termed the photosensitizer (PS), light, and oxygen, even though some PS may also work through alternative reactions in the absence of oxygen []. After light absorption, excited oxygen states are quickly formed, initially in the singlet, and subsequently in the triplet states (i.e., considering the photocycle of organic molecules). These species can release the excitation energy in the form of light emission (e.g., fluorescence and phosphorescence) or heat release (non-radiative decay). Since excited states are intrinsically more reactive than ground states, energy and electron transfer reactions can occur. There are two main mechanisms of photosensitized oxidation: Type I reactions depend on the encounter of the excited species with biological substrates. These reactions usually occur through electron or hydrogen abstraction, leading to radical chain reactions; Type II reactions rely on energy transfer reaction from the PS triplet state to molecular oxygen, generating singlet oxygen (1O2) (Fig. 1 ) []. Spacially, type I reactions require the PS to be within a subnanometer distance to the virus, whereas type II reactions allow singlet oxygen diffusion to more than 100 nm [].

Fig. 1

Mechanisms of photosensitized oxidation reactions. The photosensitizer (PS) is a molecule capable of absorbing light depending on its specific absorption spectra. Once excited, the PS is converted from the ground state 1PS to its singlet excited 1PS? and triplet excited 3PS? states. Via Type I (contact-dependent) reactions both 1PS? and 3PS? can react directly with O2 or biomolecules, like carbohydrates, lipids, proteins, or nucleic acids, resulting in the formation of radicals capable of initiating redox chain reactions. Otherwise, 3PS? can react with molecular oxygen (3O2), via the Type II (energy transfer) reaction, generating the reactive state of singlet oxygen (1O2).

Light energy is thus converted into oxidation potential that can damage biomolecules. Antimicrobial photodynamic therapy (aPDT) is based on this process and it has been used to treat localized microbial infections caused by viruses, bacteria, fungi, and parasites []. Among the many pathogens that can be targeted by aPDT, viruses are perhaps the most vulnerable, as they depend on entering a host cell for survival and replication and can be inactivated by damaging the capsid or envelope molecules (lipids, carbohydrates, proteins) or internal molecules (nucleic acids) (Fig. 1). Thus, many viruses can be treated via aPDT, including papillomavirus (HPV), hepatitis A virus (HAV), and herpes simplex virus (HSV) []. Additionally, the disinfection of biological fluids (plasma and blood products) by photoantimicrobials has been performed for decades and is a well-regarded technological application of these compounds. For instance, extracorporeal photoinactivation of coronaviruses and other clinically relevant pathogens using methylene blue (MB)-mediated aPDT has been reported [].

It is possible that photosensitized oxidation can neutralize SARS-CoV-2 and, consequently, play a role in mitigating the ongoing pandemic; however, there is no data available on the photodynamic inactivation of this virus. Thus, here we sought to find and discuss scientific literature that could help predict whether COVID-19 is more or less susceptible to oxidant species generated during aPDT.

While all types of viruses can be neutralized by aPDT, the inactivation efficiency depends on both the PS and the virus [,]. As a rule of the thumb, RNA-type phages are more easily photoinactivated than their DNA-type counterparts, suggesting that SARS-CoV-2, which is an enveloped RNA-type virus, can be easily neutralized by aPDT [,]. Guanine bases are the major targets for oxidation by photosensitizing agents in both RNA and DNA []. The formation of RNA-protein crosslinks may also be an important lesion involved in virus inactivation via aPDT [].

Enveloped viruses are more prone to aPDT neutralization than those without an envelope, due to the role of PS in damaging envelope components [,]. Initial studies on viral inactivation by aPDT demonstrated the importance of the PS reaching specific reaction sites, so-called “marked targets”, for efficient viral inactivation []. Other reports have confirmed the importance of PS binding on the efficiency of virus inactivation via aPDT, and the PS membrane partition coefficients can be used as a predictor of its virus inactivation efficacy [,]. Transmission electron microscopy data has revealed that low PS concentrations degrade envelope surface glycoproteins blocking virus internalization, while higher PS concentrations can destroy lipid membranes []. These results can be interpreted in terms of the current mechanistic understanding of photosensitized oxidation, specifically the important role of direct-contact reactions. Irreversible membrane damage occurs with the abstraction of a hydrogen atom from an unsaturated fatty acid by direct reaction with the triplet excited state of the PS. Subsequent formation of peroxyl and alkoxyl radicals leads to the build-up of truncated lipid aldehydes, which are ultimately responsible for opening membrane pores []. The fact that irreversible damage occurs due to contact-dependent reactions, indicates that the damage can be confined within the nanometer location site of the PS [].

In terms of the application of aPDT to treat COVID-19 patients, it is encouraging to note that this technique is already used to treat several respiratory diseases []. PDT has been used for decades to treat lung cancers and its successful application in the treatment of laryngeal papillomas has also been reported []. Geralde et al. recently demonstrated that acute pneumonia caused by Streptococcus pneumoniae could be treated via inhalation of indocyanine green combined with extracorporeal administration of infrared light []. A prophylactic approach proposed by Soares et al. showed that aPDT can also be used to eliminate bacterial biofilms frequently associated with endotracheal tubes and that can lead to more severe stages of acute respiratory syndromes []. More recently, Schikora and colleagues reported succesfull use of aPDT to disinfect oral and nasal cavity of patients in early stages of COVID-19 infection This approach can potentially lead to a temporary and moderate reduction of disease progression but cannot be regarded as a potential therapeutic procedure since aPDT is limited to local effects and COVID-19 is a systemic disease [].

Considering that: 1) SARS-CoV2 is an enveloped RNA virus, 2) aPDT is efficient at neutralizing such viruses, and 3) light is already used to treat lung and airway-related infections, we propose that aPDT is a good candidate for treating COVID-19 or as an adjunct to disinfect biological fluids. Alternatively, photosensitizers could also be used to decontaminate liquids and surfaces or be incorporated into polymeric matrices such as plastics, fabrics, paper, and paints to produce photoantimicrobial materials [,,]. Allotropes of carbon such as fullerenes, carbon nanotubes, and graphene can also show light-activated antimicrobial effects, including the inactivation of viruses [,,].

7. Antimicrobial Blue Light

Visible blue light exhibits microbicidal effects in the wavelength range of 405–470 nm [,]. High-intensity narrow-spectrum light at 405 nm has been used for continuous decontamination of inpatient and outpatient burn units and patient-occupied intensive care isolation rooms, as well as the treatment of surgical site infections in an orthopedic operating room []. Compared to UV-C, in general terms antimicrobial blue light (aBL) requires a much higher radiant exposure (or longer exposure times) to reach similar levels of microbial inactivation if irradiance of the light sources is similar. Even though aBL displays decreased deleterious effects on mammalian cells, one should avoid direct eye exposure because eye lens focuses visible light and overexposure can promote either flash blindness or retinal lesions.

The exact mechanisms underlying the antimicrobial effects of blue light are not yet completely understood but appear to involve the formation of short-lived reactive oxygen species (ROS) []. The most widely accepted view of the process posits that the photochemical mechanisms of aBL are based on light energy excitation of endogenous microbial intracellular light receptors (chromophores), such as porphyrins and flavins. Once excited, these receptors undergo energy transfer processes that lead to the generation of cytotoxic ROS which react with intracellular components resulting in photodamage and cell death by oxidative stress []. Since endogenous photoreceptors appear to be absent in viruses, the mechanisms by which aBL affects these pathogens remains unclear. However, it is currently known that: 1) the use of exogenous photosensitizers improves the efficiency of inactivation by blue light, and 2) the inactivation is more pronounced when viral particles are present in body fluids, e.g., saliva, feces, and blood plasma, which contain photosensitive substances [,].

Accordingly, antimicrobial blue light has been explored in the treatment of infectious diseases and as a disinfection adjuvant in healthcare settings. Clinical trials have revealed the efficiency of aBL in the treatment of acne, Helicobacter pylori gastrointestinal infections, and dental infections [,]. aBL was recently shown to rescue mice from methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa wound infections [,]. Oral anaerobic periodontopathogenic bacteria (Porphyromonas gingivalisPrevotella intermedia, and P. nigrescens) were also inhibited or completely eradicated under blue light irradiation [,].

In a recent bioinformatics study, SARS-CoV-2 infection was reported to be dependent on porphyrin, which it captures from human hemoglobin, resulting in altered heme metabolism []. However, the in silico methods used to obtain such results have been questioned by a commentary publication, putting into doubt wheter SARS-CoV-2 actually interacts with heme metabolism and accumulates porphyrins []. If this thesis is experimentally proven to be correct, aBL might be able to kill SARS-CoV-2 by photoexcitation of its acquired porphyrins. Thus, experimental studies are required to verify the potential of aBL to prevent and control COVID-19.

8. Photobiomodulation Therapy

Photobiomodulation (PBM) employs low levels of red or near-infrared (NIR) light to treat and heal wounds and injuries, reduce pain and inflammation, regenerate damaged tissue, and protect tissue at risk of dying []. Instead of directly targeting viruses, PBM mainly acts on the host cells, which absorb light in the red and near-infrared spectral region []. Literature indicates that photons are absorbed by multiple cellular chromophores, including mitochondrial enzymes, to trigger the biological effects of PBM []. Cytochrome c oxidase (i.e., unit IV in the mitochondrial respiratory chain) appears to play a main role in this process []. Other molecular chromophores include light and heat-sensitive ion channels (transient receptor potential) that, upon light activation, lead to changes in calcium concentrations. Nanostructured water (interfacial water) is also likely to act as a chromophore. Upon irradiation, the mitochondrial membrane potential is raised and oxygen consumption and ATP generation are increased. Subsequent activation of signaling pathways and transcription factors leads to fairly long-lasting effects even after relatively brief exposure of the tissue to light [].

In the early 1900s, Finsen reported that patients exposed to red light exhibited significantly better recovery from smallpox infections than unexposed counterparts []. Since then, PBM has been used in the treatment of acute lung injury, pulmonary inflammation, and models of acute respiratory distress syndrome (ARDS), due to its ability to substantially reduce systemic inflammation while preserving lung function. []. There are currently 90 published papers on PBM concerning “acute lung injury” [] OR “pulmonary inflammation” [] OR “lung inflammation” [] OR “ARDS” [] OR “lung oxidative stress” [] OR “asthma” [] many involving small animal models where it can be argued that light penetrates more easily than in humans. Because COVID-19 involves a “cytokine storm”, PBM delivered to the torso (chest and back) might not only allow some light to reach the lungs but might also reduce the systemic inflammation responsible for COVID-19 sepsis-like syndrome [] and disseminated intravascular coagulation [] that can be deadly []. Moreover, PBM is more effective on hypoxic cells [], suggesting it could be effective for COVID-19 infection, which seems to be characterized by severe hypoxia []. Nevertheless, so far there are no experimental data supporting the influence of PBM on COVID-19. Therefore, clinical studies have to be performed to understand whether PBM therapy may actually reduce the cytokine storm impacts for COVID-19 patients.

Hospitalized patients receiving mechanical ventilation or under high-oxygen continuous positive airway pressure (CPAP) treatment could be placed on an LED pad. These do not generate unacceptable levels of heat, so the high fever experienced by these patients should not be a problem. LED-based PBM devices similar to these have been approved by the FDA for general health and wellness applications, and there are no reported adverse effects []. However, PBM is not recommended to be used over cancerous lesions since the effects on tumor cells are not fully understood yet [].

9. Ultrafast Laser Irradiation

Ultrashort pulse lasers (USPLs) emitting visible to near-infrared light have been used to inactivate a broad spectrum of viruses (human immunodeficiency virus, human papillomavirus, encephalomyocarditis virus, M13 bacteriophage, tobacco mosaic virus, and murine cytomegalovirus) with no damage to human or murine cells []. Regardless of wavelength, ultrafast laser irradiation at low mean irradiance levels (? 1 W/cm2) does not promote ionization effects that could impair host cells. This irradiation does not appear to destroy either bovine serum albumin or single-stranded DNA, nor cause adverse effects like those produced by toxic or carcinogenic chemicals. Previous works suggest that the antimicrobial effect of USPLs at low mean irradiance is exerted via impulsive stimulated Raman scattering, whereby high-frequency resonance vibrations provoke vibrations of sufficient strength to disintegrate the capsid into subunits through the breaking of weak links (e.g., hydrogen bonds and hydrophobic contacts) in non-enveloped viruses []. For enveloped virus, USPLs promote vibrations on the proteins of the capsid. These excitations break the hydrogen bonds and hydrophobic contacts causing partial unfolding of the proteins. Since the concentration of confined proteins is very high within the capsid of a virus, they can assemble with other neighboring proteins, leading to the aggregation of proteins []. In contrast, an intense laser pulse could generate shock wave-like vibrations upon impact with the virus to promote viral inactivation [].

However, laser pulsing may not be necessary for its antimicrobial action. Recently, Kingsley et al. applied a tunable mode-locked Ti-Sapphire laser emitting femtosecond pulses at wavelengths of 400, 408, 425, 450, 465, and 510 nm to verify inactivation of murine norovirus (MNV) []. Using an average power of 150 mW, authors observed that femtosecond-pulsed light emitting at 408, 425 and 450 nm promoted more than 99.9% of virus inactivation after 3 h of illumination, indicating that the inactivation mechanism is not wavelength-specific. In addition, they reported that a continuous wave 408 nm laser at similar power also promoted reduction of plaque-forming units, although the addition of exogenous photosensitizers has increased MNV inactivation. These data suggest that virus inactivation does not require pulsing and can be improved in the presence of singlet oxygen enhancers, as previously reported for aBL (see section 7).

Potential use of USPLs encompasses the inactivation of pathogens in pharmaceuticals, blood products and uncooked foods as well as chemical-free whole inactivated virus vaccine preparation [,]. Laser treatment resulted in 1-log, 2-log, and 3-log reductions in hepatitis A, human immunodeficiency, and murine cytomegalovirus, respectively, in human plasma with no changes in the structure of fibrinogen []. Further, in mice USPL-induced inactivation of H1N1 influenza virus was more effective than formalin and did not cause damage to viral surface proteins or resulted in the production of carbonyl groups in proteins [].

Concluding remarks

As we presented in this review, light-based technologies have unique features that could be useful to face the COVID-19 pandemic, but could also present pitfalls that deserve to be highlighted. Thus, we compiled at Table 1 their advantages and disadvantages.

Table 1

Light-based strategies available to combat the emergence of COVID-19 pandemic. FFR: filtering facepiece respirator.

Light-based Platform Potential Applications Advantages Disavantages
Natural Ultraviolet Light Synthesis of vitamin D Sunburn following overexposure
Microbicidal activity Long-term aging and cancer risk
Ultraviolet Germicidal Irradiation Surface, FFR reuse, air and water disinfection Low exposure time to reach high levels of pathogen inactivation (< 1 min) depending on irradiance of light source Risk of tissue damage and cancer
Potential long-term degradation of materials
Photoantimicrobials and Photodynamic Therapy Environmental and surface disinfection, therapeutics, virus inactivation in biological products Efficient and selective pathogen inactivation following short period of illumination if photosensitizer is resonant to light source wavelength Photosensitizer could promote material and/or tissue staining
Systemic PS administration may cause photosensitivity
Succesfull results depend on light parameters, type of microorganism, PS concentration and pre-irradiation time
Non-invasive approach
Succesfull results in humans with artificial light sources
Antimicrobial Blue Light Environmental and surface disinfection, therapeutics, virus inactivation in biological materials Can be used in inhabited places and to treat infections in humans Long exposure time (above 30 min)
No notable detrimental effect in materials following long periods of illumination
Effect is more pronounced in the presence of exogenous photoabsorbers
Photobiomodulation Therapy Therapeutics Non-invasive technique Succesfull results depend on light parameters, patient characteristics and disease aetiology
Succesfull results in humans with artificial light sources
Adjuvant to conventional therapies
Ultrafast Laser Irradiation at low irradiance Selective virus inactivation in blood products, pharmaceuticals, food and vaccine development Selective pathogen inactivation Long exposure time (~3 h)
Chemical-free vaccine preparation Expensive light sources

In summary, we have described how light-based strategies can be used to reduce SARS-CoV-2 transmission through air, water, and surfaces as well as potential therapeutic applications that can reduce COVID-19 morbidity and mortality. From our perspective, light provides several practical answers to the new logistical and therapeutic challenges brought by COVID-19. Therefore, we suggest that the death toll and quarantine extent can be significantly mitigated if at least part of these strategies are encouraged and implemented by health systems. Given the urgent demand raised by the current uncontrolled pandemic we must be ready to use all the available armamentarium to fight COVID-19.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

We attest no conflict of interest in the manuscript we are submitting entitled “Antimicrobial light-based technologies for management of COVID-19 pandemic crisis”.

Acknowledgements

CPS was supported by the São Paulo Research Foundation (FAPESP, grant 2017/22406-0) and by the Brazilian National Council for Scientific and Technological Development (CNPq, scholarship 141901/2016-0). FPS is supported by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES Finance code 001). ALS is supported by a Marie-Curie Global Fellowship mentored by GPT (EU project 843116 – REBELLION). TD is supported by USA National Institutes of Health (NIH, grant R01AI123312) and by the Department of Defense (DoD, grant FA9550-17-1-0277). MRH is supported by USA National Institutes of Health (NIH, grants R01AI050875 and R21AI121700). MSR thanks Photonics Institute (INCT/CNPq, grant 465763/2014-6) for financial support. MSB acknowledges FAPESP for the CEPID Redoxoma grant 2013/07937-8.

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Ankylosing Spondylitis

Lasers Med Sci. 2016 Apr;31(3):459-69. doi: 10.1007/s10103-016-1874-2. Epub 2016 Jan 21.

LLLT for the management of patients with ankylosing spondylitis.

Stasinopoulos D1, Papadopoulos K2, Lamnisos D1, Stergioulas A3.

Author information

1
Physiotherapy Program, Department of Health Sciences, School of Sciences, European University Cyprus, Laureate International Universities, 6 Diogenes Street, 2044, Engomi, Nicosia, Cyprus.
2
Physiotherapy Program, Department of Health Sciences, School of Sciences, European University Cyprus, Laureate International Universities, 6 Diogenes Street, 2044, Engomi, Nicosia, Cyprus. kostpap@yahoo.com.
3
Lab of Health, Fitness and Disability Management, Faculty of Human Movement and Quality of Life, University of Peloponnese, Efstathiou & Stamatikis Balioti & Plateon, 231 00, Sparta, Laconia, Greece.

Abstract

This study aimed to compare the effectiveness of the combined lowlevel laser therapy (LLLT) and passive stretching with combined placebo LLLT laser and the same passive stretching exercises in patients suffering from ankylosing spondylitis. Forty-eight patients suffering from ankylosing spondylitis participated in the study and were randomized into two groups. Group A (n=24) was treated with a wavelength = 820 Ga-Al-As laser CW, with power intensity = 60 mW/cm(2), energy per point in each session = 4.5 J, total energy per session =27.0 J, in contact with specific points technique, plus passive stretching exercises. Group B (n = 24), received placebo laser plus the same passive stretching exercises. Both groups received 12 sessions of laser or placebo within 8 weeks; two sessions per week (weeks 1-4) and one session per week (weeks 5-8). Pain and function scales were completed before the treatment, at the end of the fourth and eighth week of treatment, and 8 weeks after the end of treatment (follow-up). Group A revealed a significant improvement after 8 weeks of treatment in all pain and function scales. At 8-week follow-up, the improvement remained only for the pain, while for all other function outcomes the differences were not statistically significant. The results suggested that after an 8-week treatment and after a follow-up, the combination of LLLT and passive stretching exercises decreased pain more effectively than placebo LLLT along with the same passive stretching exercises in patients with ankylosing spondylitis. Future studies are needed to establish the relative and absolute effectiveness of the above protocol.

KEYWORDS:

Ankylosing spondylitis; LLLT; Passive stretching

Prolotherapy and LLLT

Anesth Pain Med. 2017 Oct 15;7(5):e14470. doi: 10.5812/aapm.14470. eCollection 2017 Oct.

Prolotherapy and Low Level Laser Therapy: A Synergistic Approach to Pain Management in Chronic Osteoarthritis.

Tieppo Francio V1,2,3, Dima RS4, Towery C1, Davani S1.

Author information

1
University of Science, Arts and Technology – USAT College of Medicine, Olveston, Montserrat, BWI.
2
Essential Integrative Health – Spine, Orthopaedics and Pain Management, Oklahoma City, OK, USA.
3
Variety Care – Community Health Center, Oklahoma City, OK, USA.
4
School of Interdisciplinary Sciences – McMaster University, Hamilton, ON, Canada.

Abstract

Regenerative injection therapy and low level laser therapy are alternative remedies known for their success in the treatment and symptomatic management of chronic musculoskeletal conditions. In response to the growing demand for alternative therapies in the face of the opioid epidemic, the authors conduct a literature review to investigate the potential for prolotherapy and LLLT to be used adjunctively to manage chronic osteoarthritis (OA). OA is a degenerative chronic musculoskeletal condition on the rise in North America, and is frequently treated with opioid medications. The regenerative action of prolotherapy and pain-modulating effects of LLLT may make these two therapies well-suited to synergistically provide improved outcomes for osteoarthritis patients without the side effects associated with opioid use. A narrative descriptive review through multiple medical databases (Google Scholar, PubMed, and MedLine) is conducted, restricted by the use of medical subject headings. 71 articles were selected for reading in full, and 40 articles were selected for use in the study after reading in full. A review of the literature revealed good clinical results in the use of prolotherapy and LLLT separately to manage chronic musculoskeletal pain due to osteoarthritis and other chronic conditions. It is also recognized in the literature that prolotherapy works most effectively when used adjunctively with other treatments. Downsides to the use of prolotherapy include mild side effects of pain, stiffness and bruising and potential adverse events as a result of injection. This study is limited by the lack of clinical trials available involving both LLLT and prolotherapy injections used adjunctively, and by the low number of high impact literature concerning the treatment of (specifically) osteoarthritis by alternative methods. The authors suggest that practicing health care providers consider utilizing LLLT and prolotherapy together as a supplementary method in the management of chronic pain due to osteoarthritis, to minimize the long-term prescription of opioids and emphasize a less invasive treatment for this debilitating condition.

Hand (N Y). 2014 Dec;9(4):419-46. doi: 10.1007/s11552-014-9642-x.

Non-surgical treatment of lateral epicondylitis: a systematic review of randomized controlled trials.

Sims SE1, Miller K2, Elfar JC1, Hammert WC1.

Author information

1
Department of Orthopaedics and Rehabilitation, University of Rochester Medical Center, 601 Elmwood Ave, Box 665, Rochester, NY 14642 USA.
2
University of Rochester School of Medicine and Dentistry, 601 Elmwood Ave, Box 601, Rochester, NY 14642 USA.

Abstract

BACKGROUND:

Non-surgical approaches to treatment of lateral epicondylitis are numerous. The aim of this systematic review is to examine randomized, controlled trials of these treatments.

METHODS:

Numerous databases were systematically searched from earliest records to February 2013. Search terms included “lateral epicondylitis,” “lateral elbow pain,” “tennis elbow,” “lateral epicondylalgia,” and “elbow tendinopathy” combined with “randomized controlled trial.” Two reviewers examined the literature for eligibility via article abstract and full text.

RESULTS:

Fifty-eight articles met eligibility criteria: (1) a target population of patients with symptoms of lateral epicondylitis; (2) evaluation of treatment of lateral epicondylitis with the following non-surgical techniques: corticosteroid injection, injection technique, iontophoresis, botulinum toxin A injection, prolotherapy, platelet-rich plasma or autologous blood injection, bracing, physical therapy, shockwave therapy, or laser therapy; and (3) a randomized controlled trial design. Lateral epicondylitis is a condition that is usually self-limited. There may be a short-term pain relief advantage found with the application of corticosteroids, but no demonstrable long-term pain relief. Injection of botulinum toxin A and prolotherapy are superior to placebo but not to corticosteroids, and botulinum toxin A is likely to produce concomitant extensor weakness. Platelet-rich plasma or autologous blood injections have been found to be both more and less effective than corticosteroid injections. Non-invasive treatment methods such as bracing, physical therapy, and extracorporeal shockwave therapy do not appear to provide definitive benefit regarding pain relief. Some studies of lowlevel laser therapy show superiority to placebo whereas others do not.

CONCLUSIONS:

There are multiple randomized controlled trials for non-surgical management of lateral epicondylitis, but the existing literature does not provide conclusive evidence that there is one preferred method of non-surgical treatment for this condition. Lateral epicondylitis is a condition that is usually self-limited, resolving over a 12- to 18-month period without treatment.

LEVEL OF EVIDENCE:

Therapeutic Level II. See Instructions to Authors for a complete description of level of evidence.

KEYWORDS:

Elbow tendinopathy; Extensor tendinopathy; Lateral elbow pain; Lateral epicondylalgia; Lateral epicondylitis; Tennis elbow

Am Fam Physician. 2013 Apr 1;87(7):486-90.

Management of chronic tendon injuries.

Childress MA1, Beutler A.

Author information

1
Fort Belvoir Community Hospital, Fort Belvoir, VA 22060, USA.

Abstract

Chronic tendon injuries present unique management challenges. The assumption that these injuries result from ongoing inflammation has caused physicians to rely on treatments demonstrated to be ineffective in the long term. Nonsteroidal anti-inflammatory drugs should be limited in the treatment of these injuries. Corticosteroid injections should be considered for temporizing pain relief only for rotator cuff tendinopathy. For chronic Achilles tendinopathy (symptoms lasting longer than six weeks), an intense eccentric strengthening program of the gastrocnemius/ soleus complex improved pain and function between 60 and 90 percent in randomized trials. Evidence also supports eccentric exercise as a first-line option for chronic patellar tendon injuries. Other modalities such as prolotherapy, topical nitroglycerin, iontophoresis, phonophoresis, therapeutic ultrasound, extracorporeal shock wave therapy, and lowlevel laser therapy have less evidence of effectiveness but are reasonable second-line alternatives to surgery for patients who have persistent pain despite appropriate rehabilitative exercise.

Hemolasertherapy

Photomed Laser Surg. 2018 Apr;36(4):221-226. doi: 10.1089/pho.2017.4349.

Hemolasertherapy: A Novel Procedure for Gingival Papilla Regeneration-Case Report.

Zanin F1, Moreira MS2, Pedroni ACF3, Windlin M1, Brugnera AP4, Brugnera Júnior A5, Marques MM3.

Author information

1
1 Biophotonics Center at Institute Brugnera and Zanin-IBZ , São Paulo, Brazil .
2
2 Post Graduation Program, School of Dentistry, Ibirapuera University , Sao Paulo, Brazil .
3
3 Department of Restorative Dentistry of the School of Dentistry, University of Sao Paulo , Sao Paulo, Brazil .
4
4 PG student of Master Oral Laser Application, University of Liège , Liège, Belgium .
5
5 Associate Researcher of the National Institute of Science and Technology (INCT)-“Basic Optics and Applied to Life Sciences”-IFSC, University of Sao Paulo , Sao Paulo, Brazil .

Abstract

BACKGROUND:

Interdental papilla is of major importance to patients’ orofacial aesthetics, especially regarding anterior teeth as part of the smile’s harmony. Loss of gingival tissue, which constitutes interdental papilla, forms what in odontology is called black spaces. This loss, besides affecting the smile’s aesthetics, also provokes phonetic and functional damage.

OBJECTIVE:

The objective of the authors is to present the result of three clinical cases treated with an innovative technique called hemolasertherapy, which stimulates growth of gingival papilla and thus permanently fills in the black spaces.

METHODS:

The photobiomodulation therapy (PBMT) used a 660 nm diode laser (Laser Duo, MMO-São Carlos, SP, Brazil), punctual, contact mode in two steps: before the bleeding (first PBMT) and immediately after bleeding (second PBMT). Parameters used were power output: 100 mW, CW; diameter tip: 5 mm; spot area: 0.19 cm2; irradiation exposure time per point: 20 sec; 14 points per daily session; total of 2 sessions, with a 1-week interval; E: 2 J per point; E: per daily session, 28 J; irradiance per point: 0.52 W/cm2; fluence per point: 10.4 J/cm2. Total in two daily sessions: total energy: 56 J; total fluence: 294.75 J/cm, 560?sec total time. An in vitro preliminary study was simultaneously carried out to demonstrate what could happen at cellular level in hemotherapy clinical cases associated with PBMT laser application.

RESULTS:

This initial study demonstrated that the blood clot originated from the bleeding provoked in the gingival area is rich in mesenchymal stem cells. PBMT enables preservation, viability, and further differentiation, stimulating the return of gingival stem cells, which would support their survival and differentiation in the blood clot, thus favoring interdental papilla regeneration.

CONCLUSIONS:

Follow-up was done for a time span of 4-5 years and considered excellent with regard to papilla preservation.

KEYWORDS:

dentistry; periodontology; photobiomodulation; stem cells

Tissue Engineered, Skin Substitutes

J Biomater Appl. 2018 Jan 1:885328218759385. doi: 10.1177/0885328218759385. [Epub ahead of print]

Red light accelerates the formation of a human dermal equivalent.

Oliveira AC1, Morais TF1, Bernal C2, Martins VC2, Plepis AM1,2, Menezes PF3, Perussi JR1,2.

Author information

1
1 Programa de Pós-Graduação Interunidades Bioengenharia – EESC/FMRP/IQSC, 67817 Universidade São Paulo-São Carlos-SP , Brazil.
2
2 Instituto de Química de São Carlos, 67817 Universidade de São Paulo-São Carlos-SP , Brazil.
3
3 Instituto de Física de São Carlos, Universidade de São Paulo-São Carlos-SP, Brazil.

Abstract

Development of biomaterials’ substitutes and/or equivalents to mimic normal tissue is a current challenge in tissue engineering. Thus, three-dimensional cell culture using type I collagen as a polymeric matrix cell support designed to promote cell proliferation and differentiation was employed to create a dermal equivalent in vitro, as well to evaluate the photobiomodulation using red light. Polymeric matrix cell support was prepared from porcine serous collagen (1.1%) hydrolyzed for 96 h. The biomaterial exhibited porosity of 95%, a median pore of 44 µm and channels with an average distance between the walls of 78 ± 14 µm. The absorption of culture medium was 95%, and the sponge showed no cytotoxicity to Vero cells, a non-tumor cell line. Additionally, it was observed that irradiation with light at 630 nm (fluency 30J/cm-2) leads to the cellular photobiomodulation in both monolayer and human dermal equivalent (three-dimensional cell culture system). It was also verified that the cells cultured in the presence of the polymeric matrix cell support, allows differentiation and extracellular matrix secretion. Therefore, the results showed that the collagen sponge used as polymeric matrix cell support and the photobiomodulation at 630 nm are efficient for the production of a reconstructed human dermal equivalent in vitro.

J Biomed Opt. 2009 May-Jun;14(3):034002. doi: 10.1117/1.3127201.

Effect of lowlevel laser treatment of tissue-engineered skin substitutes: contraction of collagen lattices.

Ho G1, Barbenel J, Grant MH.

Author information

1
Exploit Technologies, Biomedical Sciences Division, Agency of Science and Technology (A STAR), 30 Biopolis Street, Singapore 138671, Singapore.

Abstract

Fibroblast-populated collagen lattices (FPCL) are widely used in tissue-engineered artificial skin substitutes, but their main drawback is that interaction of fibroblasts and matrix causes contraction of the lattice, reducing it to about 20% of its original area. The effect of lowlevellaser treatment (LLLT) on the behavior of 3T3 fibroblasts seeded in collagen lattices containing 20% chondroitin-6-sulphate was investigated to determine whether LLLT could control the contraction of FPCL. A He-Ne laser was used at 632.8 nm to deliver a 5-mW continuous wave with fluences from 1 to 4 J/cm(2). Laser treatment at 3 J/cm(2) increased contraction of collagen lattices in the absence of cells but decreased contraction of cell seeded lattices over a 7-day period. The effect was energy dependent and was not observed at 1, 2, or 4 J/cm(2). There was no alteration in fibroblast viability, morphology, or mitochondrial membrane potential after any laser treatments, but the distribution of actin fibers within the cells and collagen fibers in the matrices was disturbed at 3 J/cm(2). These effects contribute to the decrease in contraction observed. LLLT may offer a means to control contraction of FPCL used as artificial skin substitutes.

Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2006 Feb;20(2):172-6.

[Primary grafting research of tissue engineered oral mucosa lamina propria on skin full thickness wounds].

[Article in Chinese]
Wu Z1, Ding Y, Zhang L, Zhong S, Jiang T.

Author information

1
Department of Oral Surgery, 2nd Hospital of China Medical University, Shenyang Liaoning 110004, P R China. Lauwuzq@sina.com

Abstract

OBJECTIVE:

To study the allograft effect of two kinds of tissue engineered oral mucosa lamina propria on skin full-thickness wounds.

METHODS:

The cultured Wistar rat oral mucosa fibroblasts (OMF) were incorporated into collagen or chitosan-collagen to construct the tissue engineered oral mucosa lamina propria, and then the OMFs were labeled with BrdU. The full-thickness round skin defects were made with a round knife (diameter, 0.8 cm) on the backs of 36 Wistar rats (21-25 weeks old), which were divided into 2 experimental groups: the fibroblast-populated collagen lattices (FPCL) group (grafted by FPCLs) and the fibroblast-populated chitosan collagen lattices (FPCCL) group (grafted by FPCCLs), and the control group (only covered with gauges). All the wounds were observed by the naked eyes or the light microscope, and were measured 4, 7, 14, and 21 days postoperatively.

RESULTS:

There were no infection during the wound healing period. At 7 days after the grafting, the wounds in the 3 groups were covered by scab and/or gauze; at 14 days, the gauze and scab on the wounds in the three groups were all replaced by the new epidermis naturally except one scab each in the FPCCL group and the control groups, which was replaced at 17 days. All the centers of the new epidermis were measurable as the pink red points. At 21 days, all the new skins were smooth without hairs, and their color was similar to the normal one. At 4, 7, and 14 days, there was an indication that the wound diameters became significantly smaller in the three groups; but after the 14th day, there was no significant indication of this kind. At 7 days, the wound diameter in the FPCL group was significantly smaller than that in the FPCCL group and the control group (P < 0.01). Under the light microscope, at 4 days postoperatively, the decayed tissue on the surfaces of the recipient wounds in the FPCL group and the FPCCL group was separated from the lower granular tissue in which there were many inflammatory cells, fibroblasts, and new vessels. There was a similar phenomenon in the control group. Each skin wound in the three groups was only partly keratinocytes at 7 days postoperatively. The recipient wounds were wholly keratinocytes with when rete ridges observed at 14 and 21 days, but in the control group the wounds were keratinocytes with no rete ridges. Fibers in the new dermis were thin. The OMFs with Brdu appeared in the granular tissue and new dermis at 4, 7, 14, and 21 days postoperatively, which could be illustrated by the immunohistochemical staining. The positive OMFs and the granular tissue joined in the repair of the skin defects without any allergic reaction during the period of the wound healing.

CONCLUSION:

The oral mucosa fibroblasts as the new seed cells can join in the repair of the skin defects effectively and feasible. The fibroblast-populated collagen lattices and the fibroblast-populated chitosan collagen lattices can repair skin defects effectively and feasible, too. And the quality of the new skins was better in the two experimental groups than in the control group.

Skin Grafts

Lasers Med Sci. 2018 Jan 24. doi: 10.1007/s10103-017-2430-4. [Epub ahead of print]

Effect of low-level laser therapy on the healing process of donor site in patients with grade 3 burn ulcer after skin graft surgery (a randomized clinical trial).

Vaghardoost R1, Momeni M2, Kazemikhoo N3, Mokmeli S4, Dahmardehei M1, Ansari F5, Nilforoushzadeh MA3, Sabr Joo P1, Mey Abadi S1, Naderi Gharagheshlagh S1, Sassani S6.

Author information

1
Burn Research center, Department of Plastic and Reconstructive Surgery, Iran University of Medical Sciences, Tehran, Iran.
2
Burn Research center, Department of Plastic and Reconstructive Surgery, Iran University of Medical Sciences, Tehran, Iran. mah_momeni@yahoo.com.
3
Skin and Stem Cell Research Center, Tehran University of Medical Sciences, Tehran, Iran.
4
Canadian Optic and Laser Center, Victoria, BC, Canada.
5
Laser Application in Medical Sciences Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
6
Al Nasr Sport Club Medical Section, Dubai, United Arab Emirates.

Abstract

Skin graft is a standard therapeutic technique in patients with deep ulcers, but managing donor site after grafting is very important. Although several modern dressings are available to enhance the comfort of donor site, using techniques that accelerate wound healing may enhance patient satisfaction. Low-level laser therapy (LLLT) has been used in several medical fields, including healing of diabetic, surgical, and pressure ulcers, but there is not any report of using this method for healing of donor site in burn patients. The protocols and informed consent were reviewed according to Medical Ethics Board of Shahid Beheshti University of Medical Sciences (IR.SBMU.REC.1394.363) and Iranian Registry of Clinical Trials (IRCT2016020226069N2). Eighteen donor sites in 11 patients with grade 3 burn ulcer were selected. Donor areas were divided into 2 parts, for laser irradiation and control randomly. Laser area was irradiated by a red, 655-nm laser light, 150 mW, 2 J/cm2, on days 0 (immediately after surgery), 3, 5, and 7. Dressing and other therapeutic care for both sites were the same. The patients and the person who analyzed the results were blinded. The size of donor site reduced in both groups during the 7-day study period (P <0.01) and this reduction was significantly greater in the laser group (P =?0.01). In the present study, for the first time, we evaluate the effects of LLLT on the healing process of donor site in burn patients. The results showed that local irradiation of red laser accelerates wound healing process significantly.

KEYWORDS:

Low-level laser therapy; Skin graft; Wound healing

Lasers Med Sci. 2017 Apr;32(3):641-648. doi: 10.1007/s10103-017-2160-7. Epub 2017 Feb 2.

Photobiomodulation laser and pulsed electrical field increase the viability of the musculocutaneous flap in diabetic rats.

Leite GP1, das Neves LM1, Silva CA2, Guirro RR1, de Souza TR1, de Souza AK1, Garcia SB3, Guirro EC4,5.

Author information

1
Post-Graduate Program in Rehabilitation and Functional Performance, Ribeirão Preto Medical School-FMRP/USP, Ribeirão Preto, Brazil.
2
Post-Graduate Program in Human Movement Sciences, Methodist University of Piracicaba-UNIMEP, Piracicaba, Brazil.
3
Post-Graduate Program in Pathology, Ribeirao Preto Medical School-FMRP/USP, Ribeirão Preto, Brazil.
4
Post-Graduate Program in Rehabilitation and Functional Performance, Ribeirão Preto Medical School-FMRP/USP, Ribeirão Preto, Brazil. ecguirro@fmrp.usp.br.
5
Ribeirão Preto Medical School of the University of São Paulo (USP), Bandeirantes Avenue, 3900, Ribeirão Preto, SP, 14049-900, Brazil. ecguirro@fmrp.usp.br.

Abstract

The purpose of this study is to investigate the effect of pulsed electrical field (PEF) and photobiomodulation laser (PBM) on the viability of the TRAM flap in diabetic rats. Fifty Wistar rats were divided into five homogeneous groups: Group 1-control; Group 2-diabetics; Group 3-diabetics + PEF; Group 4-diabetic + laser 660 nm, 10 J/cm2, 0.27 J; Group 5-diabetic + laser 660 nm, 140 J/cm2, 3.9 J. The percentage of necrotic area was evaluated using software Image J®. The peripheral circulation of the flap was evaluated by infrared thermography FLIR T450sc (FLIR® Systems-Oregon USA). The thickness of the epidermis (haematoxylin-eosin), mast cell (toluidine blue), leukocytes, vascular endothelial growth factor, fibroblast and newly formed blood vessels were evaluated. For the statistical analysis, the Kruskal-Wallis test was applied followed by Dunn and ANOVA test followed by Tukey with critical level of 5% (p?<?0.05). The PEF reduced the area of necrosis, decreased the leukocytes, increased the mast cells, increased the thickness of epidermis and increased newly formed blood vessels when it was compared to the untreated diabetic group of animals. Laser 660 nm, fluence 140 J/cm2 (3.9 J) showed better results than the 10 J/cm2(0.27 J) related to reduction of the area of necrosis and the number of leukocytes, increased mast cells, increased thickness of the epidermis, increased vascular endothelial growth factor, increased fibroblast growth factor and increase of newly formed blood vessels in diabetic animals. The laser and pulsed electrical field increase the viability of the musculocutaneous flap in diabetic rats.

KEYWORDS:

Diabetes mellitus; Dosimetry; Fibroblast growth factor; Lowlevel laser therapy; Phototherapy; Physiotherapy; Vascular endothelial growth factor

Lasers Med Sci. 2017 Feb;32(2):335-341. doi: 10.1007/s10103-016-2118-1. Epub 2016 Dec 2.

Laser photobiomodulation (830 and 660 nm) in mast cells, VEGF, FGF, and CD34 of the musculocutaneous flap in rats submitted to nicotine.

das Neves LM1, Leite GP2, Marcolino AM2, Pinfildi CE3, Garcia SB4, de Araújo JE1, Guirro EC5.

Author information

1
Post-Graduate Program in Rehabilitation and Functional Performance of Ribeirão Preto Medical School of the University of São Paulo (FMRP/USP), Av. dos Bandeirantes, 3900, Ribeirão Preto, SP, 14049-900, Brazil.
2
Post-Graduate Program in Rehabilitation Sciences, Federal University of Santa Catarina – Campus Araranguá – UFSC/SC, Florianópolis, Brazil.
3
Post-Graduate Program in Interdisciplinary Health Sciences, Federal University of Sao Paulo – Santos Campus – UNIFESP/SP, Sao Paulo, Brazil.
4
Post-Graduate Program in Pathology, Ribeirão Preto Medical School – FMRP/USP, Sao Paulo, Brazil.
5
Post-Graduate Program in Rehabilitation and Functional Performance of Ribeirão Preto Medical School of the University of São Paulo (FMRP/USP), Av. dos Bandeirantes, 3900, Ribeirão Preto, SP, 14049-900, Brazil. ecguirro@fmrp.usp.br.

Abstract

The aim of this study was to investigate the effect of laser photobiomodulation (PBM) on the viability of the transverse rectus abdominis musculocutaneous (TRAM) flap in rats subjected to the action of nicotine. We evaluated 60 albino Wistar rats, divided into six groups of ten animals. Group 1 (saline) underwent the surgical technique to obtain a TRAM flap; group 2 (laser 830 nm) underwent the surgical technique and was irradiated with a laser 830 nm; group 3 (laser 660 nm) underwent the surgical technique and was irradiated with a laser 660 nm; group 4 was treated with nicotine subcutaneously (2 mg/kg/2×/day/4 weeks) and underwent surgery; group 5 (nicotine?+?laser 830 nm) was exposed to nicotine, underwent the surgical technique, and was irradiated with a laser 830 nm; group 6 (nicotine?+?laser 660 nm) was exposed to nicotine, underwent the surgical technique, and was irradiated with a laser 660 nm. The application of PBM occurred immediately after surgery and on the two following days. The percentage of necrosis was assessed using the AxioVision® software. The number of mast cells (toluidine blue staining) was evaluated, and immunohistochemistry was performed to detect vascular endothelial growth factor expression (anti-VEGF-A), fibroblasts (anti-basic FGF), and neoformed vessels (anti-CD34). PBM with a wavelength of 830 nm increased the viability of the TRAM flap, with a smaller area of necrosis, increased number of mast cells, and higher expression of VEGF and CD34. PBM increases the viability of musculocutaneous flaps treated with to nicotine.

Lasers Med Sci. 2016 Apr;31(3):497-502. doi: 10.1007/s10103-016-1896-9. Epub 2016 Feb 11.

Effects of low level laser therapy on the prognosis of split-thickness skin graft in type 3 burn of diabetic patients: a case series.

Dahmardehei M1, Kazemikhoo N2, Vaghardoost R1, Mokmeli S3, Momeni M1, Nilforoushzadeh MA4,5, Ansari F4, Amirkhani A4.

Author information

1
Burn Research Center, Iran University of Medical Sciences, Tehran, Iran.
2
Skin and Stem Cell Research Center, Tehran University of Medical Sciences, Tehran, Iran. nooshakazemi@gmail.com.
3
Canadian Optic and Laser Center, Victoria, BC, Canada.
4
Skin and Stem Cell Research Center, Tehran University of Medical Sciences, Tehran, Iran.
5
Skin Diseases and Leishmaniasis Research Center, Isfahan University of Medical Sciences, Isfahan, Iran.

Abstract

Significant populations in burn centers are diabetic burn patients. Healing process in these patients is more difficult due to diabetes complications. The gold standard treatment for patients with grade 3 burn ulcer is split-thickness skin grafting (STSG), but in the diabetic patients, the rate of graft failure and amputation is high due to impaired tissue perfusion. The technique of low level laser therapy (LLLT) improves tissue perfusion and fibroblast proliferation, increases collagen synthesis, and accelerates wound healing. The purpose of this case report is introducing a new therapeutic method for accelerating healing with better prognosis in these patients. The protocols and informed consent were reviewed according to the Medical Ethics, Board of Shahid Beheshti Medical Sciences (IR.SBMU.RAM.REC.13940.363). Diabetic type 2 patients with 13 grade 3 burn ulcers, candidate for amputation, were enrolled in the study. We used a 650-nm red laser light, 2 J/Cm for the bed of the ulcer and an 810-nm infrared laser light 6 J/Cm(2) for the margins along with intravenous laser therapy with a 660-nm red light, before and after STSG for treating grade 3 burn ulcers in 13 diabetic ulcers. The results of this study showed complete healing in the last 8 weeks for all patients who were candidates for amputation. In this case series, we present 13 cases of diabetic ulcer with type 3 burn wound, candidate for amputation, who healed completely using LLLT and STSG. This is the first time that these two techniques are combined for treatment of burn ulcer in diabetic patients. Using LLLT with STSG might be a promising treatment for burn victims especially diabetic patients.

KEYWORDS:

Burn wound; Low level laser therapy; Skin transplantation

Plast Surg (Oakv). 2015 Spring;23(1):35-9.

Inhibitory effects of low-level laser therapy on skin-flap survival in a rat model.

Baldan CS1, Masson IF1, Esteves Júnior I1, Baldan AM1, Machado AF1, Casaroto RA1, Liebano RE1.

Author information

1
Paulista University, São Paulo, Brazil.

Abstract

in English, French

BACKGROUND:

Although several studies have demonstrated the effects of low-level laser therapy (LLLT) on skin flap viability, the role of higher doses has been poorly investigated.

OBJECTIVE:

To investigate the inhibitory effect of the LLLT (?=670 nm) on the viability of random skin flaps in a rat model using an irradiation energy of 2.79 J at each point.

METHODS:

Sixteen Wistar rats were randomly assigned into two groups: sham laser irradiation (n=8); and active laser irradiation (n=8). Animals in the active laser irradiation group were irradiated with a 670 nm diode laser with an energy of 2.79 J/point, a power output 30 mW, a beam area of 0.028 cm(2), an energy density of 100 J/cm(2), an irradiance of 1.07 W/cm(2) for 93 s/point. Irradiation was performed in 12 points in the cranial skin flap portion. The total energy irradiated on the tissue was 33.48 J. The necrotic area was evaluated on postoperative day 7.

RESULTS:

The sham laser irradiation group presented a mean (± SD) necrotic area of 47.96±3.81%, whereas the active laser irradiation group presented 62.24±7.28%. There was a significant difference in skin-flap necrosis areas between groups (P=0.0002).

CONCLUSION:

LLLT (?=670 nm) increased the necrotic area of random skin flaps in rats when irradiated with an energy of 2.79 J (100 J/cm(2)).

KEYWORDS:

Low-level laser therapy; Necrosis; Rats; Surgical flaps

Minerva Stomatol. 2014 Mar;63(3):77-83.

Histological assessment of nonablative laser stimulation of tissue repair in acellular dermal grafts.

Silveira V1, Cenci R, Oliveira M, Moraes J, Etges A, Zerbinatti L.

Author information

1
Oral and maxillofacial Department Pontifícia Universidade do Rio Grande do Sul (PUCRS) Porto Alegre, RS, Brazil – gerhardtoliveira@gmail.com.

Abstract

AIM:

The objective of this study was to compare integration of AlloDerm® acellular dermal grafts in animals subjected to non-ablative laserirradiation and animals not exposed to this therapy.

METHODS:

Standardized AlloDerm® fragments measuring 5 mm² were grafted into the subcutaneous tissue overlying the calvaria in 32 Wistar rats. Laser therapy (685 nm), at a dose of 4 J/cm2 per session, was applied immediately after surgical intervention and every 48 hours thereafter for a total of four applications.

RESULTS:

Analysis of histology slides revealed significantly greater edema in the control group. There was no neutrophil infiltration in the laser-irradiated group at any point during the study period, whereas such infiltration was present in control animals at three of the four points of observation. In the laser therapy group, lymphocyte infiltration was observed from day 1, whereas in the control group, it was only apparent from day 3. Vascularization was substantially greater in the control group. In the experimental group, the AlloDerm® graft was completely replaced by fibrous tissue.

CONCLUSION:

These findings suggest that add-on non-ablative laser therapy is an effective stimulator of healing and graft integration after placement of AlloDerm® acellular dermal grafts.

Lasers Med Sci. 2013 May;28(3):755-61. doi: 10.1007/s10103-012-1130-3. Epub 2012 Jun 22.

What is better in TRAM flap survival: LLLT single or multi-irradiation?

Pinfildi CE1, Hochman BS, Nishioka MA, Sheliga TR, Neves MA, Liebano RE, Ferreira LM.

Author information

1
Department of Science of Human Movement, University Federal of São Paulo-UNIFESP, Campus Baixada Santista, Santos, São Paulo, Brazil. cepinfildi@hotmail.com

Abstract

Lowlevel laser therapy (LLLT) has been used with the aim of improving vascular perfusion of the skin and musculocutaneous flaps. This study evaluated the effect of LLLT on transverse rectus abdominis musculocutaneous flap (TRAM) viability, vascular angiogenesis, and VEGF release. Eighty-four Wistar rats were randomly divided into seven groups with 12 rats in each group. Group 1 received sham lasertreatment; group 2, 3 J/cm(2) at 1 point; group 3, 3 J/cm(2) at 24 points; group 4, 72 J/cm(2) at 1 point; group 5, 6 J/cm(2) at 1 point; group 6, 6 J/cm(2) at 24 points; and group 7, 144 J/cm(2) at 1 point. All experimental groups underwent LLLT immediately after the TRAM operation and on the following 2 days; thus, animals underwent 3 days of treatment. The percentage of skin flap necrosis area was calculated on the fourth postoperative day using the paper template method, and two skin samples were collected using a 1-cm(2) punch to evaluate alpha smooth muscle actin (1A4) and VEGF levels in blood vessels. Significant differences were found in necrosis percentage, and higher values were seen in group 1 than in the other groups. Statistically significant differences were not found among groups 3 to 7 (p<0.292). Groups 5 and 7 showed significantly higher VEGF levels compared to other groups. Groups 3 and 5 had an increase in levels of blood vessels compared to other groups. LLLT at energy densities of 6 to 144 J/cm(2) was efficient to increase angiogenesis and VEGF levels and promote viability in TRAM flaps in rats.

Rev Col Bras Cir. 2013 Jan-Feb;40(1):44-8.

Macro and microscopic analysis of island skin grafts after lowlevel laser therapy.

[Article in English, Portuguese]
da Silva EB1, Maniscalco CL, Ésper GV, Guerra RR, Kerppers II.

Author information

1
Department of Agricultural and Environmental Sciences, State University of Santa Cruz – UESC, Ilheus, Bahia State – BA, Brazil. elisangelavet@yahoo.com.br

Abstract

OBJECTIVE:

To observe the effects of low intensity laser therapy in inflammation, wound healing and epithelialization of island skin grafts.

METHODS:

Twenty rats were subjected to this grafting technique and divided subsequently into two equal groups, one treated with laser and the other control.

RESULTS:

there was less inflammation, faster healing, epithelialization and keratinization in the laser-treated animals when compared to the untreated.

CONCLUSION:

Low intensity laser therapy is helpful to island skin grafting.

Lasers Med Sci. 2012 Sep;27(5):1045-50. doi: 10.1007/s10103-011-1042-7. Epub 2011 Dec 30.

LED (660 nm) and laser (670 nm) use on skin flap viability: angiogenesis and mast cells on transition line.

Nishioka MA1, Pinfildi CE, Sheliga TR, Arias VE, Gomes HC, Ferreira LM.

Author information

1
Post Graduation Plastic Surgery, Federal University of São Paulo, R. Napoleão de Barros, 715, 4º andar, CEP 04024-900, São Paulo, SP, Brazil.

Abstract

Skin flap procedures are commonly used in plastic surgery. Failures can follow, leading to the necrosis of the flap. Therefore, many studies use LLLT to improve flap viability. Currently, the LED has been introduced as an alternative to LLLT. The objective of this study was to evaluate the effect of LLLT and LED on the viability of random skin flaps in rats. Forty-eight rats were divided into four groups, and a random skin flap (10?×?4 cm) was performed in all animals. Group 1 was the sham group; group 2 was submitted to LLLT 660 nm, 0.14 J; group 3 with LED 630 nm, 2.49 J, and group 4 with LLLT 660 nm, with 2.49 J. Irradiation was applied after surgery and repeated on the four subsequent days. On the 7th postoperative day, the percentage of flap necrosis was calculated and skin samples were collected from the viable area and from the transition line of the flap to evaluate blood vessels and mast cells. The percentage of necrosis was significantly lower in groups 3 and 4 compared to groups 1 and 2. Concerning blood vessels and mast cell numbers, only the animals in group 3 showed significant increase compared to group 1 in the skin sample of the transition line. LED and LLLT with the same total energies were effective in increasing viability of random skin flaps. LED was more effective in increasing the number of mast cells and blood vessels in the transition line of random skin flaps.

Acta Cir Bras. 2012 Feb;27(2):155-61.

The effects of different doses of 670 nm diode laser on skin flap survival in rats.

Baldan CS1, Marques AP, Schiavinato AM, Casarotto RA.

Author information

1
Physical Therapy Department, UNIP and Sao Paulo Metodista University, Brazil. cristianobaldan@yahoo.com.br

Abstract

PURPOSE:

To investigate the effects of different low-level laser therapy (LLLT) doses on random skin flap rats.

METHODS:

Forty Wistar rats were randomly divided in four groups. The control group (CG) was not irradiated. The experimental groups were irradiated with a diode laser 670 nm with different energies per point: group 2 (G2) with 0.06 J; group 3 (G3) 0.15 J and group 4 (G4) 0.57 J. The three groups were irradiated in 12 equally distributed points in the cranial skin flap portion. They were submitted to the irradiation during the immediate, first and second postoperative days. The necrosis area was evaluated in the seventh postoperative day.

RESULTS:

The CG shows 49.35% of necrosis area in the skin flap; G2, 39.14%; G3, 47.01% and G4, 29.17% respectively. There was a significantly difference when G4 was compared with CG`s skin flap necrosis area.

CONCLUSION:

The low-level laser therapy diode 670 nm with 0.57 J energy per point increases the survival in randomic skin flap rats.

Photomed Laser Surg. 2010 Aug;28(4):483-8. doi: 10.1089/pho.2009.2500.

Influence of the use of laser phototherapy (lambda660 or 790 nm) on the survival of cutaneous flaps on diabetic rats.

Santos NR1, dos Santos JN, dos Reis JA Jr, Oliveira PC, de Sousa AP, de Carvalho CM, Soares LG, Marques AM, Pinheiro AL.

Author information

1
School of Dentistry, Federal University of Bahia, Salvador, Bahia, Brazil.

Abstract

OBJECTIVE:

The aim of this study was to assess and compare the effects of laser phototherapy (LPT) on cutaneous flaps on diabetic rats.

BACKGROUND:

Diabetes mellitus is characterized by high blood glucose levels. Its main complications are delayed wound healing, an impaired blood supply, and a decrease in collagen production. Cutaneous flaps are routinely used in several surgical procedures, and most failures are related to poor blood supply. LPT has been studied using several healing models.

ANIMALS AND METHODS:

Twelve Wistar rats were randomized into three groups: group 1 (G1; diabetic animals without treatment), group 2 (G2; diabetic animals irradiated with lambda680 nm), and group 3 (G3; diabetic animals irradiated with lambda790 nm). Diabetes was induced with streptozotocin. A 2- x 8-cm cutaneous flap was raised on the dorsum of each animal, and a plastic sheet was introduced between the flap and the bed to cause poor blood supply. Nonirradiated animals acted as controls. The dose per session was 40 J/cm(2). Laser light was applied transcutaneously and fractioned on 16 contact points at the wound margins (16 x 2.5 J/cm(2)). Animal death occurred on day 8 after surgery. Specimens were taken, processed, cut, stained with eosin (HE) and sirius red, and underwent histological analysis.

RESULTS:

It is shown that accute inflammation was mostly discrete for G3. Chronic inflammation was more evident for G2. Fibroblast number was higher for G3. Angiogenesis was more evident for G3. Necrosis was more evident for G2. Statistical analysis among all groups showed significant differences (p = 0.04) on the level of acute inflammation between G1 and G3, tissue necrosis between G1 and G2 (p = 0.03), chronic inflammation between (p = 0.04), fibroblastic proliferation between G2 and G3 (p = 0.05), and neovascularization between G2 and G3 (p = 0.04).

CONCLUSION:

LPT was effective in increasing angiogenesis as seen on irradiated subjects and was more pronounced when IR laser light was used.

Photomed Laser Surg. 2010 Jun;28(3):379-84. doi: 10.1089/pho.2009.2535.

Effect of low-level laser therapy on malondialdehyde concentration in random cutaneous flap viability.

Prado R1, Neves L, Marcolino A, Ribeiro T, Pinfildi C, Ferreira L, Thomazini J, Piccinato C.

Author information

1
Department of Surgery and Anatomy, University of São Paulo-FMRP-USP, Ribeirão Preto, Brazil. paschoalrp@hotmail.com

Abstract

OBJECTIVE:

The aim of this study was to assess the effects of 830 and 670 nm laser on malondialdehyde (MDA) concentration in random skin-flap survival.

BACKGROUND DATA:

Low-level laser therapy (LLLT) has been reported to be successful in stimulating the formation of new blood vessels and activating superoxide-dismutase delivery, thus helping the inhibition of free-radical action and consequently reducing necrosis.

MATERIALS AND METHODS:

Thirty Wistar rats were used and divided into three groups, with 10 rats in each one. A random skin flap was raised on the dorsum of each animal. Group 1 was the control group; group 2 received 830 nm laser radiation; and group 3 was submitted to 670 nm laser radiation. The animals underwent laser therapy with 36 J/cm(2) energy density immediately after surgery and on the 4 days subsequent to surgery. The application site of the laser radiation was 1 point, 2.5 cm from the flap’s cranial base. The percentage of the skin-flap necrosis area was calculated 7 days postoperative using the paper-template method, and a skin sample was collected immediately after as a way of determining the MDA concentration.

RESULTS:

Statistically significant differences were found between the necrosis percentages, with higher values seen in group 1 compared with groups 2 and 3. Groups 2 and 3 did not present statistically significant differences (p > 0.05). Group 3 had a lower concentration of MDA values compared to the control group (p < 0.05).

CONCLUSION:

LLLT was effective in increasing the random skin-flap viability in rats, and the 670 nm laser was efficient in reducing the MDA concentration.

Photomed Laser Surg. 2009 Jun;27(3):411-6. doi: 10.1089/pho.2008.2320.

Effect of application site of low-level laser therapy in random cutaneous flap viability in rats.

Prado RP1, Pinfildi CE, Liebano RE, Hochman BS, Ferreira LM.

Author information

1
Master of Basic Sciences in Plastic Surgery, São Paulo Federal University, São Paulo, SP, Brazil. paschoalrp@hotmail.com

Abstract

OBJECTIVE:

This study aimed to investigate the effect of diode laser (830 nm) irradiation on the viability of ischemic random skin flaps in rats, as well as to determine the most effective site for applying laser radiation to speed healing.

BACKGROUND DATA:

Low-level laser therapy (LLLT) has recently been used to improve the viability of ischemic random skin flaps in rats.

MATERIALS AND METHODS:

Seventy Wistar rats were used and divided into seven groups of 10 rats each: group 1, sham laser treatment; group 2, which received irradiation at 1 point 5 cm from the flap’s cranial base; group 3, which received irradiation at 2 points (5 and 7.5 cm from the flap’s base); group 4, which received irradiation at 3 points (2.5, 5 and 7.5 cm from the flap’s base); group 5, which received irradiation at 1 point 2.5 cm from the flap’s base; group 6, which received irradiation at 2 points (2.5 and 5 cm from the flap’s base); and group 7, which received irradiation at 1 point 7.5 cm from the flap’s base. The animals were subjected to laser therapy at an energy density of 36 J/cm(2) for 72 sec immediately after surgery, and one time on each of the four subsequent days. The percentage of necrotic skin flap area was calculated on the seventh postoperative day using a paper template.

RESULTS:

The results showed that the rats in group 5 had the highest increase in skin flap viability, with a statistically significant difference compared to the other groups. Statistically significant differences were not seen between any of the other groups.

CONCLUSION:

The diode laser was effective in increasing skin flap viability in rats, and laser irradiation of a point 2.5 cm from the cranial base flap was found to be the most effective.

Photomed Laser Surg. 2009 Apr;27(2):337-43. doi: 10.1089/pho.2008.2295.

Effect of low-level laser therapy on mast cells in viability of the transverse rectus abdominis musculocutaneous flap.

Pinfildi CE1, Liebano RE, Hochman BS, Enokihara MM, Lippert R, Gobbato RC, Ferreira LM.

Author information

1
Department of Plastic Surgery and IMES-FAFICA, São Paulo Federal University, São Paulo, SP, Brazil. cepinfildi@hotmail.com

Abstract

OBJECTIVE:

To assess the effect of low-level laser therapy (LLLT) on viability of mast cells of the transverse rectus abdominis musculocutaneous (TRAM) flap.

BACKGROUND DATA:

LLLT has been recently used on the TRAM flap to stimulate mast cells.

MATERIALS AND METHODS:

Eighty-four Wistar rats were randomly divided into seven groups of 12 rats in each: group 1 (sham laser therapy); group 2 received 3 J/cm(2) at one point; group 3 received 3 J/cm(2) at 24 points; group 4 received 72 J/cm(2) at 1 point; group 5 received 6 J/cm(2) at 1 point; group 6 received 6 J/cm(2) at 24 points; and group 7 received 144 J/cm(2) at 1 point. All experimental groups underwent LLLT immediately after TRAM surgery and on the next two following days, for three sessions in total. The percentage of the area of skin flap necrosis was calculated on the fourth postoperative day and two samples of skin were collected from each rat with a 1-cm(2) punch to perform mast cell evaluations with toluidine blue dye.

RESULTS:

Statistically significant differences were found in the percentage of necrosis, and higher values were seen in group 1 than in all other groups. Among groups 3-7 no statistically significant differences were found (p < 0.292). For mast cells, when group 1 was compared to groups 5 (6 J/cm(2) at 1 point) and 7 (144 J/cm(2) at 1 point), it had fewer mast cells.

CONCLUSION:

LLLT at a wavelength of 670 nm was effective at reducing the necrotic area, and we found that it can stimulate mast cells growth to increase vascular perfusion.

Ann Plast Surg. 1985 Mar;14(3):278-83.

Effects of low-power diode lasers on flap survival.

Kami T, Yoshimura Y, Nakajima T, Ohshiro T, Fujino T.

Abstract

We investigated the effect of low-power laser irradiation on the survival of experimental skin flaps in rats. A gallium-aluminum-arsenide diode laser that was developed by the Japan Medical Laser Laboratory was used. The laser power was 15 mW and the wavelength 830 nm. Irradiation was carried out, either before or after flap elevation, in two groups of 20 Wistar strain rats. A third group of 20 rats served as controls. A caudally based skin flap, 3 X 9 cm, was designed on the back of each rat. Laser irradiation therapy was performed for 5 consecutive days for 6 minutes per flap per day, preoperatively in one group and postoperatively in the other. Seven days postoperatively, the survival areas of the flaps were measured and compared. The survival area was increased significantly in both groups receiving laser therapy, probably due to the observed proliferation of blood vessels around the irradiated points and an increase in blood flow.

PMID:
3994272

Aging / Longevity

 2018 Sep;33(7):1513-1519. doi: 10.1007/s10103-018-2510-0. Epub 2018 Apr 26.

Photobiomodulation effects on mRNA levels from genomic and chromosome stabilization genes in injured muscle.

Author information

1
Laboratório de Pesquisa em Células Tronco, Departamento de Histologia e Embriologia, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, Avenida 28 de Setembro, 87, fundos, Vila Isabel, Rio de Janeiro, 20551-030, Brazil.
2
Laboratório de Biomorfologia e Patologia Experimental, Universidade Severino Sombra, Avenida Expedicionário Oswaldo de Almeida Ramos 280, Vassouras, Rio de Janeiro, 27700-000, Brazil.
3
Departamento de Biofísica e Biometria, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, Avenida 28 de Setembro, 87, fundos, Vila Isabel, Rio de Janeiro, 20551-030, Brazil.
4
Departamento de Biofísica e Biometria, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, Avenida 28 de Setembro, 87, fundos, Vila Isabel, Rio de Janeiro, 20551-030, Brazil. adnfonseca@yahoo.com.br.
5
Departamento de Ciências Fisiológicas, Instituto Biomédico, Universidade Federal do Estado do Rio de Janeiro, Rua Frei Caneca, 94, Rio de Janeiro, 20211-040, Brazil. adnfonseca@yahoo.com.br.

Abstract

Muscle injuries are the most prevalent type of injury in sports. A great number of athletes have relapsed in muscle injuries not being treated properly. Photobiomodulation therapy is an inexpensive and safe technique with many benefits in muscle injury treatment. However, little has been explored about the infrared laser effects on DNA and telomeres in muscle injuries. Thus, the aim of this study was to evaluate photobiomodulation effects on mRNA relative levels from genes related to telomere and genomic stabilization in injured muscle. Wistar male rats were randomly divided into six groups: control, laser 25 mW, laser 75 mW, injury, injury laser 25 mW, and injury laser 75 mW. Photobiomodulation was performed with 904 nm, 3 J/cm2 at 25 or 75 mW. Cryoinjury was induced by two applications of a metal probe cooled in liquid nitrogen directly on the tibialis anterior muscle. After euthanasia, skeletal muscle samples were withdrawn and total RNA extracted for evaluation of mRNA levels from genomic (ATM and p53) and chromosome stabilization (TRF1 and TRF2) genes by real-time quantitative polymerization chain reaction. Data show that photobiomodulation reduces the mRNA levels from ATM and p53, as well reduces mRNA levels from TRF1 and TRF2 at 25 and 75 mW in injured skeletal muscle. In conclusion, photobiomodulation alters mRNA relative levels from genes related to genomic and telomere stabilization in injured skeletal muscle.

KEYWORDS:

DNA; Laser; Muscle; Wistar rats

Photomed Laser Surg. 2018 Mar 23. doi: 10.1089/pho.2017.4393. [Epub ahead of print]

Aging Is a Sticky Business.

Sommer AP1.

Author information

1
Ulm, Germany .

Abstract

OBJECTIVE:

The objective of this work is to put forward a mechanism by which low-level light [red-to near infrared (NIR) laser or light emitting diodes (LED)] is instrumental in the process of accelerating the healing of wounds.

BACKGROUND DATA:

Interaction modalities of low-level light with oxidatively stressed cells and tissues are the focus of intense research efforts. Several models of the light/cell-interaction mechanism have been proposed. In the most popular model, cytochrome c oxidase is believed to play the role of the principal acceptor for red-to NIR photons.

METHODS:

Using as an illustrative example the successful LED treatment of an edematous limb ulcer, the results of recent in vitro tests and complementary laboratory experiments are presented and discussed.

RESULTS:

The most plausible mechanism of biostimulatory effect of red-to NIR light consists of its impact on the nanoscopic interfacial water layers in mitochondria and the extracellular matrix (ECM) where mitochondrial reactive oxygen species (ROS) induce an increase in the viscosity of the water layers bound to the predominantly hydrophilic surfaces in the intramitochondrial space as well as the ECM, where the process progressively propagates with age. The biostimulatory effect of red-to NIR light consists of counteracting the ROS-induced elevation of interfacial water viscosities, thereby instantly restoring the normal mitochondrial function, including the synthesis of adenosine triphosphate (ATP) by the rotary motor (ATP synthase).

CONCLUSIONS:

An understanding of the mechanism of interaction of red-to NIR light with mitochondria, cells, and tissues safeguards progress in the field of low-level light therapy (LLLT) and puts us in the position to design better therapies.

KEYWORDS:

ATP; LED; ROS; interfacial water viscosity; laser; mitochondria; wound

Neurobiol Aging. 2018 Feb 26;66:131-137. doi: 10.1016/j.neurobiolaging.2018.02.019. [Epub ahead of print]

Photobiomodulation reduces gliosis in the basal ganglia of aged mice.

El Massri N1, Weinrich TW2, Kam JH2, Jeffery G2, Mitrofanis J3.

Author information

1
Department of Anatomy F13, University of Sydney, Sydney, NSW, Australia.
2
Institute of Ophthalmology, University College London, London, England.
3
Department of Anatomy F13, University of Sydney, Sydney, NSW, Australia. Electronic address: john.mitrofanis@sydney.edu.au.

Abstract

This study explored the effects of long-term photobiomodulation (PBM) on the glial and neuronal organization in the striatum of aged mice. Mice aged 12 months were pretreated with PBM (670 nm) for 20 minutes per day, commencing at 5 months old and continued for 8 months. We had 2 control groups, young at 3 months and aged at 12 months old; these mice received no treatment. Brains were aldehyde-fixed and processed for immunohistochemistry with various glial and neuronal markers. We found a clear reduction in glial cell number, both astrocytes and microglia, in the striatum after PBM in aged mice. By contrast, the number of 2 types of striatal interneurons (parvalbumin+ and encephalopsin+), together with the density of striatal dopaminergic terminals (and their midbrain cell bodies), remained unchanged after such treatment. In summary, our results indicated that long-term PBM had beneficial effects on the aging striatum by reducing glial cell number; and furthermore, that this treatment did not have any deleterious effects on the neurons and terminations in this nucleus.

KEYWORDS:

Astrocytes; Caudate-putamen complex; Interneurons; Microglia; Substantia nigra

J Biophotonics. 2017 Dec 11. doi: 10.1002/jbio.201700282. [Epub ahead of print]

Aging of lymphoid organs: Can photobiomodulation reverse age-associated thymic involution via stimulation of extrapineal melatonin synthesis and bone marrow stem cells?

Odinokov D1, Hamblin MR2,3,4.

Author information

1
Department of Biomedical Engineering, Chinese University of Hong Kong, Hong Kong.
2
Wellman Center for Photomedicine, Massachusetts General Hospital, Boston, MA, 02114, USA.
3
Department of Dermatology, Harvard Medical School, Boston, MA, 02115, USA.
4
Harvard-MIT Division of Health Sciences and Technology, Cambridge, MA, 02139, USA.

Abstract

Thymic atrophy and the subsequent reduction in T cell production are the most noticeable age-related changes affecting lymphoid organs in the immune system. In fact thymic involution has been described as “programmed aging”. New therapeutic approaches such as photobiomodulation (PBM) may reduce or reverse these changes. PBM (also known as low-level laser therapy or LLLT) involves the delivery of non-thermal levels of red or near-infrared light that are absorbed by mitochondrial chromophores, in order to prevent tissue death and stimulate healing and regeneration. PBM may reverse or prevent thymic involution due to its ability to induce extrapineal melatonin biosynthesis via cyclic AMP or NF-kB activation, or alternatively by stimulating bone marrow stem cells that can regenerate the thymus. This perspective puts forward a hypotheses that PBM can alter thymic involution, improve immune functioning in aged people, and even extend lifespan.

Neurobiol Aging. 2017 Oct;58:140-150. doi: 10.1016/j.neurobiolaging.2017.06.025. Epub 2017 Jul 6.

Transcranial low-level laser therapy improves brain mitochondrial function and cognitive impairment in D-galactose-induced aging mice.

Salehpour F1, Ahmadian N2, Rasta SH3, Farhoudi M2, Karimi P2, Sadigh-Eteghad S4.

Author information

1
Neurosciences Research Center (NSRC), Tabriz University of Medical Sciences, Tabriz, Iran; Department of Medical Physics, Tabriz University of Medical Sciences, Tabriz, Iran.
2
Neurosciences Research Center (NSRC), Tabriz University of Medical Sciences, Tabriz, Iran.
3
Department of Medical Physics, Tabriz University of Medical Sciences, Tabriz, Iran; Department of Medical Bioengineering, Tabriz University of Medical Sciences, Tabriz, Iran; School of Medical Sciences, University of Aberdeen, Aberdeen, UK.
4
Neurosciences Research Center (NSRC), Tabriz University of Medical Sciences, Tabriz, Iran. Electronic address: Saeed.sadigetegad@gmail.com.

Abstract

Mitochondrial function plays a key role in the aging-related cognitive impairment, and photoneuromodulation of mitochondria by transcranial low-level laser therapy (LLLT) may contribute to its improvement. This study focused on the transcranial LLLT effects on the D-galactose (DG)-induced mitochondrial dysfunction, apoptosis, and cognitive impairment in mice. For this purpose, red and near-infrared (NIR) laser wavelengths (660 and 810 nm) at 2 different fluencies (4 and 8 J/cm2) at 10-Hz pulsed wave mode were administrated transcranially 3 d/wk in DG-received (500 mg/kg/subcutaneous) mice model of aging for 6 weeks. Spatial and episodic-like memories were assessed by the Barnes maze and What-Where-Which (WWWhich) tasks. Brain tissues were analyzed for mitochondrial function including active mitochondria, adenosine triphosphate, and reactive oxygen species levels, as well as membrane potential and cytochrome c oxidase activity. Apoptosis-related biomarkers, namely, Bax, Bcl-2, and caspase-3 were evaluated by Western blotting method. Laser treatments at wavelengths of 660 and 810 nm at 8 J/cm2 attenuated DG-impaired spatial and episodic-like memories. Also, results showed an obvious improvement in the mitochondrial function aspects and modulatory effects on apoptotic markers in aged mice. However, same wavelengths at the fluency of 4 J/cm2 had poor effect on the behavioral and molecular indexes in aging model. This data indicates that transcranial LLLT at both of red and NIR wavelengths at the fluency of 8 J/cm2 has a potential to ameliorate aging-induced mitochondrial dysfunction, apoptosis, and cognitive impairment.

KEYWORDS:

Aging; Apoptosis; D-galactose; Episodic-like memory; Mitochondrial function; Spatial memory; Transcranial low-level laser therapy

Exp Brain Res. 2017 Oct;235(10):3081-3092. doi: 10.1007/s00221-017-5048-7. Epub 2017 Jul 25.

No evidence for toxicity after long-term photobiomodulation in normal non-human primates.

Moro C1, Torres N1, Arvanitakis K2, Cullen K2, Chabrol C1, Agay D1, Darlot F1, Benabid AL1, Mitrofanis J3.

Author information

1
University of Grenoble Alpes, CEA, LETI, CLINATEC, MINATEC Campus, 38000, Grenoble, France.
2
Department of Anatomy F13, University of Sydney, Camperdown, 2006, Australia.
3
Department of Anatomy F13, University of Sydney, Camperdown, 2006, Australia. john.mitrofanis@sydney.edu.au.

Abstract

In this study, we explored the effects of a longer term application, up to 12 weeks, of photobiomodulation in normal, naïve macaque monkeys. Monkeys (n = 5) were implanted intracranially with an optical fibre device delivering photobiomodulation (red light, 670 nm) to a midline midbrain region. Animals were then aldehyde-fixed and their brains were processed for immunohistochemistry. In general, our results showed that longer term intracranial application of photobiomodulation had no adverse effects on the surrounding brain parenchyma or on the nearby dopaminergic cell system. We found no evidence for photobiomodulation generating an inflammatory glial response or neuronal degeneration near the implant site; further, photobiomodulation did not induce an abnormal activation or mitochondrial stress in nearby cells, nor did it cause an abnormal arrangement of the surrounding vasculature (endothelial basement membrane). Finally, because of our interest in Parkinson’s disease, we noted that photobiomodulation had no impact on the number of midbrain dopaminergic cells and the density of their terminations in the striatum. In summary, we found no histological basis for any major biosafety concerns associated with photobiomodulation delivered by our intracranial approach and our findings set a key template for progress onto clinical trial on patients with Parkinson’s disease.

KEYWORDS:

670 nm; Behaviour; Macaque monkeys; Striatum; Substantia nigra; Tyrosine hydroxylase

Exp Brain Res. 2017 Oct;235(10):3081-3092. doi: 10.1007/s00221-017-5048-7. Epub 2017 Jul 25.

No evidence for toxicity after long-term photobiomodulation in normal non-human primates.

Moro C1, Torres N1, Arvanitakis K2, Cullen K2, Chabrol C1, Agay D1, Darlot F1, Benabid AL1, Mitrofanis J3.

Author information

1
University of Grenoble Alpes, CEA, LETI, CLINATEC, MINATEC Campus, 38000, Grenoble, France.
2
Department of Anatomy F13, University of Sydney, Camperdown, 2006, Australia.
3
Department of Anatomy F13, University of Sydney, Camperdown, 2006, Australia. john.mitrofanis@sydney.edu.au.

Abstract

In this study, we explored the effects of a longer term application, up to 12 weeks, of photobiomodulation in normal, naïve macaque monkeys. Monkeys (n = 5) were implanted intracranially with an optical fibre device delivering photobiomodulation (red light, 670 nm) to a midline midbrain region. Animals were then aldehyde-fixed and their brains were processed for immunohistochemistry. In general, our results showed that longer term intracranial application of photobiomodulation had no adverse effects on the surrounding brain parenchyma or on the nearby dopaminergic cell system. We found no evidence for photobiomodulation generating an inflammatory glial response or neuronal degeneration near the implant site; further, photobiomodulation did not induce an abnormal activation or mitochondrial stress in nearby cells, nor did it cause an abnormal arrangement of the surrounding vasculature (endothelial basement membrane). Finally, because of our interest in Parkinson’s disease, we noted that photobiomodulation had no impact on the number of midbrain dopaminergic cells and the density of their terminations in the striatum. In summary, we found no histological basis for any major biosafety concerns associated with photobiomodulation delivered by our intracranial approach and our findings set a key template for progress onto clinical trial on patients with Parkinson’s disease.

KEYWORDS:

670 nm; Behaviour; Macaque monkeys; Striatum; Substantia nigra; Tyrosine hydroxylase

Exp Brain Res. 2017 Jun;235(6):1861-1874. doi: 10.1007/s00221-017-4937-0. Epub 2017 Mar 15.

Photobiomodulation-induced changes in a monkey model of Parkinson’s disease: changes in tyrosine hydroxylase cells and GDNF expression in the striatum.

El Massri N1, Lemgruber AP1, Rowe IJ1, Moro C2, Torres N2, Reinhart F2, Chabrol C2, Benabid AL2, Mitrofanis J3.

Author information

1
Department of Anatomy F13, University of Sydney, Sydney, 2006, Australia.
2
University of Grenoble Alpes, CEA, LETI, CLINATEC, MINATEC Campus, 38000, Grenoble, France.
3
Department of Anatomy F13, University of Sydney, Sydney, 2006, Australia. john.mitrofanis@sydney.edu.au.

Abstract

Intracranial application of red to infrared light, known also as photobiomodulation (PBM), has been shown to improve locomotor activity and to neuroprotect midbrain dopaminergic cells in rodent and monkey models of Parkinson’s disease. In this study, we explored whether PBM has any influence on the number of tyrosine hydroxylase (TH)+cells and the expression of GDNF (glial-derived neurotrophic factor) in the striatum. Striatal sections of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine)-treated mice and monkeys and 6-hydroxydopamine (6OHDA)-lesioned rats that had PBM optical fibres implanted intracranially (or not) were processed for immunohistochemistry (all species) or western blot analysis (monkeys). In our MPTP monkey model, which showed a clear loss in striatal dopaminergic terminations, PBM generated a striking increase in striatal TH+ cell number, 60% higher compared to MPTP monkeys not treated with PBM and 80% higher than controls. This increase was not evident in our MPTP mouse and 6OHDA rat models, both of which showed minimal loss in striatal terminations. In monkeys, the increase in striatal TH+ cell number in MPTP-PBM cases was accompanied by similar increases in GDNF expression, as determined from western blots, from MPTP and control cases. In summary, these results offer insights into the mechanisms by which PBM generates its beneficial effects, potentially with the use of trophic factors, such as GDNF.

KEYWORDS:

670 nm; 6OHDA; Caudate; MPTP; Near infrared light; Putamen

 2016 Aug;31(6):1161-7. doi: 10.1007/s10103-016-1956-1. Epub 2016 May 25.

Lowlevel infrared laser modulates muscle repair and chromosome stabilization genes in myoblasts.

Author information

1
Laboratório de Pesquisa em Células Tronco, Departamento de Histologia e Embriologia, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, Avenida 28 de Setembro, 87, fundos, Vila Isabel, Rio de Janeiro, 20551030, Brazil.
2
Departamento de Biofísica e Biometria, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, Avenida 28 de Setembro, 87, fundos, 4° andar, Vila Isabel, Rio de Janeiro, 20551030, Brazil.
3
Departamento de Biofísica e Biometria, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, Avenida 28 de Setembro, 87, fundos, 4° andar, Vila Isabel, Rio de Janeiro, 20551030, Brazil. adnfonseca@yahoo.com.br.
4
Departamento de Ciências Fisiológicas, Instituto Biomédico, Universidade Federal do Estado do Rio de Janeiro, Rua Frei Caneca, 94, Rio de Janeiro, 20211040, Brazil. adnfonseca@yahoo.com.br.

Abstract

Infrared laser therapy is used for skeletal muscle repair based on its biostimulative effect on satellite cells. However, shortening of telomerelength limits regenerative potential in satellite cells, which occurs after each cell division cycle. Also, laser therapy could be more effective on non-physiologic tissues. This study evaluated lowlevel infrared laser exposure effects on mRNA expression from muscle injury repair and telomere stabilization genes in myoblasts in normal and stressful conditions. Laser fluences were those used in clinical protocols. C2C12 myoblast cultures were exposed to lowlevel infrared laser (10, 35, and 70 J/cm(2)) in standard or normal (10 %) and reduced (2 %) fetal bovine serum concentrations; total RNA was extracted for mRNA expression evaluation from muscle injury repair (MyoD and Pax7) and chromosome stabilization (TRF1 and TRF2) genes by real time quantitative polymerization chain reaction. Data show that lowlevelinfrared laser increases the expression of MyoD and Pax7 in 10 J/cm(2) fluence, TRF1 expression in all fluences, and TRF2 expression in 70 J/cm(2) fluence in both 10 and 2 % fetal bovine serum. Lowlevel infrared laser increases mRNA expression from genes related to muscle repair and telomere stabilization in myoblasts in standard or normal and stressful conditions.

KEYWORDS:

Low level laser; MyoD; Pax7; TRF1; TRF2

[Experimental study of effect of low power laser on telomere length of cells].

[Article in Chinese]

Author information

1
Department of Physiology, Guangxi Medical University, Nanning 530021, China.

Abstract

To investigate the effect of low power helium neon laser (He-Ne laser) on the telomere length of human fetal lung diploid fibroblast (2BS) cell, we used the laser (gamma = 632. 8 nm, P = 2 mW) to treat the young 2BS cells. Cell growth and proliferation was observed through MTT method after treating with low power laser. The relative telomere length of 2BS cells was detected by fluorescence real-time quantitative PCR (q-PCR). The results showed that the cells of the treated groups grew better than the untreated groups. The telomereDNA length of the old 2BS cells, treated by low power He-Ne laser when they were young, was longer than that of untreated group. The results of the present study indicated that the low power He-Ne laser might decrease shortening rate of telomere and delay the aging of cells. Therefore, this study provides the experimental basis for us to further investigate the effect of low power laser on cell aging at the gene level.

Vopr Kurortol Fizioter Lech Fiz Kult.  2011 Jul-Aug;(4):39-42.

The influence of pulsed infrared laser radiation on the hormone production in the thymus (an experimental study).

[Article in Russian]
[No authors listed]

Abstract

Local irradiation with pulsed (1500 Hz) low-energy infrared laser light of the thymus and thyroid gland region caused well-apparent stimulation of alpha-1-thymosin production in the healthy animals and normalized its level in the stressed ones. Similar stimulation of alpha-1-timosine biosynthesis was observed in an experiment with direct laser irradiation of the cultured HTSC epitheliocytes from the human thymus.

Adv Gerontol. 2010;23(4):547-53.

Induced thymus aging: radiation model and application perspective for low intensive laser radiation.

[Article in Russian]
Sevost’ianova NN, Trofimov AV, Lin’kova NS, Poliakova VO, Kvetno IM.

Abstract

The influence of gamma-radiation on morphofunctional state of thymus is rather like as natural thymus aging. However gamma-radiation model of thymus aging widely used to investigate geroprotectors has many shortcomings and limitations. Gamma-radiation can induce irreversible changes in thymus very often. These changes are more intensive in comparison with changes, which can be observed at natural thymus aging. Low intensive laser radiation can not destroy structure of thymus and its effects are rather like as natural thymus aging in comparison with gamma-radiation effects. There are many parameters of low intensive laser radiation, which can be changed to improve morphofunctional thymus characteristics in aging model. Using low intensive laser radiation in thymus aging model can be very perspective for investigations of aging immune system.

Biofizika. 2007 Jan-Feb;52(1):137-40.

Protective effect of low-power laser radiation in acute toxic stress.

[Article in Russian]
Novoselova EG, Glushkova OV, Khrenov MO, Chernenkov DA, Lunin SM, Novoselova TV, Chudnovski? VM, Iusupov VI, Fesenko EE.

Abstract

The effect of preliminary short-term irradiation with He-Ne laser light (632.8 nm, 0.2 mW/cm2) of the thymus zone projection of male NMRI mice subjected to acute toxic stress on the responses of immune cells was studied. Stress was modeled by lipopolysaccharide injection, 250 mg/100 g of body weight, which induced a significant increase in the production of several macrophage cytokines, IL-1alpha, IL-1beta, IL-6, IL-10 and TNF-alpha. A single irradiation with laser light did not provoke considerable variations in NO production in cells but induced an enhancement in the production of heat shock proteins Hsp25, Hsp70, and Hsp90. Nevertheless, when irradiation with red laser light was applied prior to toxic stress, considerable normalization of production of nearly all cytokines studied and nitric oxide was observed. Moreover, the normalization of production of heat shock proteins has been shown in these conditions. Thus, preliminary exposure of a small area of animal skin surface provoked a significant lowering in the toxic effect of lipopolysaccharide.

Izv Akad Nauk Ser Biol. 2006 Nov-Dec;(6):667-79.

Structure peculiarities of muscle regenerates and state of thymus under He-Ne laser therapy in different periods after muscle trauma.

[Article in Russian]
Buliakova NV, Azarova VS.

Abstract

We studied the gastrocnemius muscle regeneration and the reactive changes in thymus of rats under different regimens of He-Ne laser therapy of both operated legs (632.8 nm; 2.5-3.0 mW/cm(2) ). Laser radiation (10 exposures by 3 min within 1-15 days after muscle trauma, 4.5-5.4 J/cm(2) total dose per each leg) stimulated inflammatory reaction, muscle healing and favored preservation of muscle tissue in regenerates. The changes in thymus mass, its histological structure, size of cortex and thymocite mitotic index pointed to the increase of the functional load on thymus and delay of its recovery. The same dose of laser therapy of muscles within 16-30 days after trauma led to the increase of muscle tissue sclerotization in regenerates. The reactive changes in thymus were less pronounced. Threefold decrease of laser dose (10 exposures by 1 min for 1-15 days, 1.5-1.8 J/cm(2)) suppressed inflammatory reaction, impaired the muscle regeneration. The increase of functional activity in thymus was not observed.

Minim Invasive Ther Allied Technol. 2006;15(5):277-85.

Regeneration of skeletal muscles and state of thymus in gamma-irradiated rats under laser therapy of the area of muscle trauma.

Bulyakova NV, Azarova VS.

Source

A N Severtzov Institute of Ecology & Evolution, Russian Academy of Sciences, Moscow, Russia. admin@sevin.ru

Abstract

The gamma-irradiation of adult rats with a semi-lethal dose (6 Gy) suppressed the posttraumatic regeneration of skeletal muscles and brought about considerable destructive changes in the thymus. The effect of He-Ne laser radiation at a total dose 4.5-5.4 J/cm2 at each operated leg in irradiated rats stimulated the regenerative capacity of skeletal muscle tissue, the healing of skin-muscle wound, and the processes of postradiation recovery in thymus cells (a decrease of chromosome aberrations). The histological structure of regenerates had more muscle pattern. At the same time, the positive dynamics of regenerative processes in muscles was achieved by an increased functional load on the thymus. To stimulate the regeneration of irradiated muscles on the background of a more moderate load on the thymus, the prolonged period of laser therapy and fragmentary distribution of laser exposures during muscle regeneration were preferable. Wound healing improved visibly. Nor formation of chronic radiation ulcers on operated shins was observed.

Biofizika. 2006 Jan-Feb;51(1):123-35.

Effects of exposure of different skin areas to low-power laser light.

[Article in Russian]
Glushkov OV, Novoselova EG, Cherenkov DA, Novoselova TV, Khrenov MO, Lunin SM, Chudnovski? VM, Iusupov VI, Fesenko EE.

Abstract

The effect of helium-neon laser light of extremely low power of 0.2 mW/cm2 and wavelength 632.8 nm on the immune status of mice bearing solid tumors was studied. The evaluation of the status of tumor-bearing animals was provided by taking into account the number of immune cells, cytokine concentration (tumor necrosis factor, interleukin 2, production of nitric oxide, expression of heat shock proteins (Hsp70 and Hsp90), and activity of natural killers. The model of a solid tumor was formed by subcutaneous injection of Ehrlich carcinoma cells, and average life span of tumor-bearing mice achieved about 55 days. Different areas of the skin of tumor-bearing mice were subjected either to a single (1 min, dose 0.012 J/cm3) or repeated exposure to laser light (1 min, 48-h intervals, 30 days). Two different areas were irradiated: the thymus projection area or a hind limb with solid tumors. The results showed that chronic exposure of tumor-bearing mice in the thymus projection area, and especially, hind limb, reduced the resistance, which manifested itself in the acceleration of tumor growth and a tendency of mouse life span to decrease. On the contrary, a single exposure stimulated the antitumor immunity for several days after the exposure. The results show the expediency of further investigation of the immunomodulative effects of low-power laser light and the necessity of monitoring the immune system during laser therap

Laser and NSAIDs

Lasers Med Sci. 2017 Aug 9. doi: 10.1007/s10103-017-2299-2. [Epub ahead of print]

Effects of photobiomodulation therapy and topical non-steroidal anti-inflammatory drug on skeletal muscle injury induced by contusion in rats-part 2: biochemical aspects.

Tomazoni SS1, Frigo L2, Dos Reis Ferreira TC3,4, Casalechi HL3, Teixeira S5, de Almeida P6, Muscara MN5, Marcos RL6, Serra AJ6, de Carvalho PTC4,6, Leal-Junior ECP3,4.

Author information

1
Masters and Doctoral Programs in Physical Therapy, Universidade Cidade de São Paulo (UNICID), Rua Cesário Galeno, 448/475, São Paulo, SP, 05508-900, Brazil. shaiane.tomazoni@gmail.com.
2
Biological Sciences and Health Center, Cruzeiro do Sul University (UNICSUL), São Paulo, SP, Brazil.
3
Laboratory of Phototherapy in Sports and Exercise, Nove de Julho University (UNINOVE), São Paulo, SP, Brazil.
4
Postgraduate Program in Rehabilitation Sciences, Nove de Julho University (UNINOVE), São Paulo, SP, Brazil.
5
Department of Pharmacology, Institute of Biomedical Sciences, University of São Paulo, São Paulo, SP, Brazil.
6
Postgraduate Program in Biophotonics Applied to Health Sciences, Nove de Julho University (UNINOVE), São Paulo, SP, Brazil.

Abstract

Muscle injuries trigger an inflammatory process, releasing important biochemical markers for tissue regeneration. The use of non-steroidal anti-inflammatory drugs (NSAIDs) is the treatment of choice to promote pain relief due to muscle injury. NSAIDs exhibit several adverse effects and their efficacy is questionable. Photobiomodulation therapy (PBMT) has been demonstrated to effectively modulate inflammation induced from musculoskeletal disorders and may be used as an alternative to NSAIDs. Here, we assessed and compared the effects of different doses of PBMT and topical NSAIDs on biochemical parameters during an acute inflammatory process triggered by a controlled model of contusion-induced musculoskeletal injury in rats. Muscle injury was induced by trauma to the anterior tibial muscle of rats. After 1 h, rats were treated with PBMT (830 nm, continuous mode, 100 mW of power, 35.71 W/cm2; 1, 3, and 9 J; 10, 30, and 90 s) or diclofenac sodium (1 g). Our results demonstrated that PBMT, 1 J (35.7 J/cm2), 3 J (107.1 J/cm2), and 9 J (321.4 J/cm2) reduced the expression of tumor necrosis factor alpha (TNF-?) and cyclooxygenase-2 (COX-2) genes at all assessed times as compared to the injury and diclofenac groups (p < 0.05). The diclofenac group showed reduced levels of COX-2 only in relation to the injury group (p < 0.05). COX-2 protein expression remained unchanged with all therapies except with PBMT at a 3-J dose at 12 h (p < 0.05 compared to the injury group). In addition, PBMT (1, 3, and 9 J) effectively reduced levels of cytokines TNF-?, interleukin (IL)-1?, and IL-6 at all assessed times as compared to the injury and diclofenac groups (p < 0.05). Thus, PBMT at a 3-J dose was more effective than other doses of PBMT and topical NSAIDs in the modulation of the inflammatory process caused by muscle contusion injuries.

Med Oral Patol Oral Cir Bucal. 2017 Jul 1;22(4):e467-e472.

Effect of pre-operatory lowlevel laser therapy on pain, swelling, and trismus associated with third-molar surgery.

Petrini M1, Ferrante M, Trentini P, Perfetti G, Spoto G.

Author information

1
Department of Medical, Oral and Biotechnological Sciences, University of Chieti – Italy, Via Vestini 31, 66013 Chieti, Italy materialidentari.uda@gmail.com.

Abstract

BACKGROUND:

The extraction of impacted third molars is commonly associated to pain, edema, trismus, limited jaw opening and movements. The aim of this retrospective study is to verify if pre-surgical lowlevel laser therapy (LLLT) associated with the extraction of impacted lower third molars could add benefits to the postoperative symptoms respect LLLT performed only after surgery.

MATERIAL AND METHODS:

Data from 45 patients subjected to a surgical extraction of lower third molars were pooled and divided into three groups. Patients that received only routine management were inserted in the control group. Group 1, were patients that received LLLT immediately after surgery and at 24 hours. In group 2 were included patients treated with LLLT immediately before the extraction and immediately after the end of the procedure. Data were analyzed using linear regression and descriptive statistics.

RESULTS:

Both laser-treated groups were characterized by minor events of post-surgery complications of pain, edema, trismus. The use of NSAIDs in the first 24 hours was significantly inferior in Group 2.

CONCLUSIONS:

Pre-surgical LLLT treatment seems to increase the analgesic effect of LLLT. However, trismus and edema were reduced in both laser treated groups, independently from the period of irradiation.

Lasers Med Sci. 2016 Winter;7(1):45-50. doi: 10.15171/jlms.2016.10. Epub 2016 Jan 7.

Low Level Laser Therapy Versus Pharmacotherapy in Improving Myofascial Pain Disorder Syndrome.

Khalighi HR1, Mortazavi H1, Mojahedi SM2, Azari-Marhabi S1, Moradi Abbasabadi F3.

Author information

1
Department of Oral and Maxillofacial Medicine, School of Dentistry, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
2
Department of Laser, School of Dentistry, Shahid Beheshti University of Medical Sciences, Tehran,Iran.
3
Department of Oral and Maxillofacial Pathology, Faculty of Dentistry, Qom University of Medical Sciences, Qom, Iran.

Abstract

INTRODUCTION:

Temporomandibular disorders (TMD) lead to masticatory muscle pain, jaw movement disability and limitation in mouth opening. Pain is the chief complaint in 90% of the TMD patients which leads to disability and severe socioeconomic costs. The purpose of this study was to evaluate the therapeutic effects of low level laser therapy (LLLT) compared to pharmacotherapy with NSAIDs (naproxen) in myofascial pain disorder syndrome (MPDS).

METHODS:

In this randomized controlled clinical trial, 40 MPDS patients were divided into two groups. One group received naproxen 500 mg bid for 3 weeks as treatment modality and also had placebo laser sessions. The other group received active laser (diode 810 nm CW) as treatment and placebo drug. Pain intensity was measured by visual analogue scale (VAS) and maximum painless mouth opening was also measured as a functional index every session and at 2 months follow up. Data was collected and analyzed with SPSS software. Independent t test was used to analyze the data. A P < 0.05 was considered significant.

RESULTS:

Low level laser caused significant reduction in pain intensity (P < 0.05) and a significant increase in mouth opening. In naproxen group neither pain intensity nor maximum mouth opening had significant improvement. Pain relief, in subjective VAS was observed in third session in LLLT group, but did not occur in naproxen group. Maximum mouth opening increased significantly in laser group compared to the naproxen group from the eighth session.

CONCLUSION:

Treatment with LLLT caused a significant improvement in mouth opening and pain intensity in patients with MPDS. Similar improvement was not observed in naproxen group.

Lasers Med Sci. 2017 Jan;32(1):101-108. doi: 10.1007/s10103-016-2091-8. Epub 2016 Oct 10.

Effects of photobiomodulation therapy, pharmacological therapy, and physical exercise as single and/or combined treatment on the inflammatory response induced by experimental osteoarthritis.

Tomazoni SS1, Leal-Junior EC2, Pallotta RC3, Teixeira S3, de Almeida P3, Lopes-Martins RÁ4.

Author information

1
Laboratory of Pharmacology and Experimental Therapeutics, Department of Pharmacology, Institute of Biomedical Sciences, University of São Paulo (USP), Av. Prof. Lineu Prestes, 1524, Butantan, São Paulo, SP, 05508-900, Brazil. shaiane.tomazoni@gmail.com.
2
Postgraduate Program in Biophotonics Applied to Health Sciences and Post Graduate Program in Rehabilitation Sciences, Nove de Julho University (UNINOVE), São Paulo, SP, Brazil.
3
Laboratory of Pharmacology and Experimental Therapeutics, Department of Pharmacology, Institute of Biomedical Sciences, University of São Paulo (USP), Av. Prof. Lineu Prestes, 1524, Butantan, São Paulo, SP, 05508-900, Brazil.
4
Biomedical Engineering Research and Post-Graduate Center, Mogi das Cruzes University (UMC), Mogi das Cruzes, SP, Brazil.

Abstract

Osteoarthritis (OA) triggers increased levels of inflammatory markers, including prostaglandin (PG) E2 and proinflammatory cytokines. The elevation of cytokine levels is closely associated with increased articular tissue degeneration. Thus, the use of combination therapies may presumably be able to enhance the effects on the modulation of inflammatory markers. The present study aimed to evaluate and compare the effects of photobiomodulation therapy (PBMT), physical exercise, and topical nonsteroidal anti-inflammatory drug (NSAID) use on the inflammatory process after they were applied either alone or in different combinations. OA was induced by intra-articular papain injection in the knee of rats. After 21 days, the animals began treatment with a topical NSAID and/or with physical exercise and/or PBMT. Treatments were performed three times a week for eight consecutive weeks, totaling 24 therapy sessions. Analysis of real-time polymerase chain reaction (RT-PCR) gene expression; interleukin (IL)-1?, IL-6, and tumor necrosis factor alpha (TNF-?) protein expression; and PGE2 levels by enzyme-linked immunosorbent assay (ELISA) was conducted. Our results showed that PBMT alone and Exerc + PBMT significantly reduced IL-1? gene expression (p?<?0.05) while no treatment changed both IL-6 and TNF-? gene expression. Treatment with NSAID alone, PBMT alone, Exerc + PBMT, and NSAID + PBMT reduced IL-1? protein expression (p<0.05). All therapies significantly reduced IL-6 and TNF-? protein expression (p<0.05) compared with the OA group. Similarly, all therapies, except Exerc, reduced the levels of PGE2 (p?<0.05) compared with the OA group. The results from the present study indicate that treatment with PBMT is more effective in modulating the inflammatory process underlying OA when compared with the other therapies tested.

Lasers Med Sci. 2014 Mar;29(2):653-8. doi: 10.1007/s10103-013-1377-3. Epub 2013 Jun 30.

What is the best treatment to decrease pro-inflammatory cytokine release in acute skeletal muscle injury induced by trauma in rats: low-level laser therapy, diclofenac, or cryotherapy?

de Almeida P1, Tomazoni SS, Frigo L, de Carvalho Pde T, Vanin AA, Santos LA, Albuquerque-Pontes GM, De Marchi T, Tairova O, Marcos RL, Lopes-Martins RÁ, Leal-Junior EC.

Author information

1
Postgraduate Program in Rehabilitation Sciences, Universidade Nove de Julho (UNINOVE), São Paulo, SP, Brazil.

Abstract

Currently, treatment of muscle injuries represents a challenge in clinical practice. In acute phase, the most employed therapies are cryotherapy and nonsteroidal anti-inflammatory drugs. In the last years, low-level laser therapy (LLLT) has becoming a promising therapeutic agent; however, its effects are not fully known. The aim of this study was to analyze the effects of sodium diclofenac (topical application), cryotherapy, and LLLT on pro-inflammatory cytokine levels after a controlled model of muscle injury. For such, we performed a single trauma in tibialis anterior muscle of rats. After 1 h, animals were treated with sodium diclofenac (11.6 mg/g of solution), cryotherapy (20 min), or LLLT (904 nm; superpulsed; 700 Hz; 60 mW mean output power; 1.67 W/cm(2); 1, 3, 6 or 9 J; 17, 50, 100 or 150 s). Assessment of interleukin-1? and interleukin-6 (IL-1? and IL-6) and tumor necrosis factor-alpha (TNF-?) levels was performed at 6 h after trauma employing enzyme-linked immunosorbent assay method. LLLT with 1 J dose significantly decreased (p?<?0.05) IL-1?, IL-6, and TNF-? levels compared to non-treated injured group as well as diclofenac and cryotherapy groups. On the other hand, treatment with diclofenac and cryotherapy does not decrease pro-inflammatory cytokine levels compared to the non-treated injured group. Therefore, we can conclude that 904 nm LLLT with 1 J dose has better effects than topical application of diclofenac or cryotherapy in acute inflammatory phase after muscle trauma.

Angle Orthod. 2010 Sep;80(5):925-32. doi: 10.2319/010410-10.1.

Interventions for pain during fixed orthodontic appliance therapy. A systematic review.

Xiaoting L1, Yin T, Yangxi C.

Author information

1
State Key Laboratory of Oral Disease and Department of Orthodontics, West China School of Dentistry, Sichuan University, Chengdu, China.

Abstract

OBJECTIVE:

To compare the different methods of pain control intervention during fixed orthodontic appliance therapy.

MATERIALS AND METHODS:

A computerized literature search was performed in MEDLINE (1966-2009), The Cochrane Library (Issue 4, 2009), EMBASE (1984-2009), and CNKI (1994-2009) to collect randomized controlled trials (RCTs) for pain reduction during orthodontic treatment. Data were independently extracted by two reviewers and a quality assessment was carried out. The Cochrane Collaboration’s RevMan5 software was used for data analysis. The Cochrane Oral Health Group’s statistical guidelines were followed.

RESULTS:

Twenty-six RCTs were identified and six trials including 388 subjects were included. Meta-analysis showed that ibuprofen had a pain control effect at 6 hours and at 24 hours after archwire placement compared with the placebo group. The standard mean difference was -0.47 and -0.48, respectively. There was no difference in pain control between ibuprofen, acetaminophen, and aspirin. Other analgesics such as tenoxicam and valdecoxib had relatively lower visual analog scale (VAS) scores in pain perception. Lowlevel laser therapy (LLLT) was also an effective approach for pain relief with VAS scores of 3.30 in the LLLT group and 7.25 in the control group.

CONCLUSIONS:

Analgesics are still the main treatment modality to reduce orthodontic pain despite their side effects. Some long-acting nonsteroidal anti-inflammatory drugs (NSAIDs) and cyclo-oxygenase enzyme (COX-2) inhibitors are recommended for their comparatively lesser side effects. Their preemptive use is promising. Other approaches such as LLLT have aroused researchers’ attention.

Photomed Laser Surg. 2010 Aug;28(4):553-60. doi: 10.1089/pho.2009.2576.

Acute low back pain with radiculopathy: a double-blind, randomized, placebo-controlled study.

Konstantinovic LM1, Kanjuh ZM, Milovanovic AN, Cutovic MR, Djurovic AG, Savic VG, Dragin AS, Milovanovic ND.

Author information

Abstract

OBJECTIVE:

The aim of this study was to investigate the clinical effects of lowlevel laser therapy (LLLT) in patients with acute low back pain (LBP) with radiculopathy.

BACKGROUND DATA:

Acute LBP with radiculopathy is associated with pain and disability and the important pathogenic role of inflammation. LLLT has shown significant anti-inflammatory effects in many studies.

MATERIALS AND METHODS:

A randomized, double-blind, placebo-controlled trial was performed on 546 patients. Group A (182 patients) was treated with nimesulide 200 mg/day and additionally with active LLLT; group B (182 patients) was treated only with nimesulide; and group C (182 patients) was treated with nimesulide and placebo LLLT. LLLT was applied behind the involved spine segment using a stationary skin-contact method. Patients were treated 5 times weekly, for a total of 15 treatments, with the following parameters: wavelength 904 nm; frequency 5000 Hz; 100-mW average diode power; power density of 20 mW/cm(2) and dose of 3 J/cm(2); treatment time 150 sec at whole doses of 12 J/cm(2). The outcomes were pain intensity measured with a visual analog scale (VAS); lumbar movement, with a modified Schober test; pain disability, with Oswestry disability score; and quality of life, with a 12-item short-form health survey questionnaire (SF-12). Subjects were evaluated before and after treatment. Statistical analyses were done with SPSS 11.5.

RESULTS:

Statistically significant differences were found in all outcomes measured (p < 0.001), but were larger in group A than in B (p < 0.0005) and C (p < 0.0005). The results in group C were better than in group B (p < 0.0005).

CONCLUSIONS:

The results of this study show better improvement in acute LBP treated with LLLT used as additional therapy.

J Oral Rehabil. 2008 Dec;35(12):925-33. doi: 10.1111/j.1365-2842.2008.01891.x.

Lowlevel laser therapy improves bone repair in rats treated with anti-inflammatory drugs.

Ribeiro DA1, Matsumoto MA.

Author information

1
Department of Biosciences, Federal University of Sao Paulo, UNIFESP, Santos, SP, Brazil. daribeiro@unifesp.br

Abstract

Nowadays, selective cyclooxygenase-2 non-steroidal anti-inflammatory drugs have been largely used in surgical practice for reducing oedema and pain. However, the association between these drugs and laser therapy is not known up to now. Herein, the aim of this study was to evaluate the action of anti-COX-2 selective drug (celecoxib) on bone repair associated with laser therapy. A total of 64 rats underwent surgical bone defects in their tibias, being randomly distributed into four groups: Group 1) negative control; Group 2) animals treated with celecoxib; Group 3) animals treated with lowlevel power laser and Group 4) animals treated with celecoxib and lowlevel power laser. The animals were killed after 48 h, 7, 14 and 21 days. The tibias were removed for morphological, morphometric and immunohistochemistry analysis for COX-2. Statistical significant differences (P < 0.05) were observed in the quality of bone repair and quantity of formed bone between groups at 14 days after surgery for Groups 3 and 4. COX-2 immunoreactivity was more intense in bone cells for intermediate periods evaluated in the laser-exposed groups. Taken together, such results suggest that lowlevel laser therapy is able to improve bone repair in the tibia of rats as a result of an up-regulation for cyclooxygenase-2 expression in bone cells.

Clin Orthop Relat Res. 2008 Jul;466(7):1539-54. doi: 10.1007/s11999-008-0260-1. Epub 2008 Apr 30.

Treatment of tendinopathy: what works, what does not, and what is on the horizon.

Andres BM1, Murrell GA.

Author information

1
Orthopaedic Research Institute, St George Hospital, University of New South Wales, Level 2 Research and Education Building, 4-10 South Street, Kogarah, Sydney, NSW, 2217, Australia. bandres@yahoo.com

Abstract

Tendinopathy is a broad term encompassing painful conditions occurring in and around tendons in response to overuse. Recent basic science research suggests little or no inflammation is present in these conditions. Thus, traditional treatment modalities aimed at controlling inflammation such as corticosteroid injections and nonsteroidal antiinflammatory medications (NSAIDS) may not be the most effective options. We performed a systematic review of the literature to determine the best treatment options for tendinopathy. We evaluated the effectiveness of NSAIDS, corticosteroid injections, exercise-based physical therapy, physical therapy modalities, shock wave therapy, sclerotherapy, nitric oxide patches, surgery, growth factors, and stem cell treatment. NSAIDS and corticosteroids appear to provide pain relief in the short term, but their effectiveness in the long term has not been demonstrated. We identified inconsistent results with shock wave therapy and physical therapy modalities such as ultrasound, iontophoresis and lowlevel laser therapy. Current data support the use of eccentric strengthening protocols, sclerotherapy, and nitric oxide patches, but larger, multicenter trials are needed to confirm the early results with these treatments. Preliminary work with growth factors and stem cells is promising, but further study is required in these fields. Surgery remains the last option due to the morbidity and inconsistent outcomes. The ideal treatment for tendinopathy remains unclear.

Cognitive Enhancement

Front Cell Neurosci 2019 Mar 19;13:74. doi: 10.3389/fncel.2019.00074. eCollection 2019.

Photobiomodulation and Coenzyme Q10 Treatments Attenuate Cognitive Impairment Associated With Model of Transient Global Brain Ischemia in Artificially Aged Mice.

Author information

1
Neurosciences Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
2
Department of Medical Physics, Tabriz University of Medical Sciences, Tabriz, Iran.
3
ProNeuroLIGHT LLC, Phoenix, AZ, United States.
4
Higher Educational Institute of Rab-Rashid, Tabriz, Iran.
5
Department of Medical Bioengineering, Tabriz University of Medical Sciences, Tabriz, Iran.
6
School of Medical Sciences, University of Aberdeen, Aberdeen, United Kingdom.
7
Wellman Center for Photomedicine, Massachusetts General Hospital, Boston, MA, United States.
8
Department of Dermatology, Harvard Medical School, Boston, MA, United States.
9
Harvard-MIT Health Sciences and Technology, Cambridge, MA, United States.
10
Departments of Clinical Research and Nuclear Medicine, Odense University Hospital, University of Southern Denmark, Odense, Denmark.
11
Department of Neuroscience, University of Copenhagen, Copenhagen, Denmark.
12
Department of Neurology & Neurosurgery, McGill University, Montreal, QC, Canada.
13
Department of Radiology and Radiological Science, Johns Hopkins University, Baltimore, MD, United States.

Abstract

Disturbances in mitochondrial biogenesis and bioenergetics, combined with neuroinflammation, play cardinal roles in the cognitive impairment during aging that is further exacerbated by transient cerebral ischemia. Both near-infrared (NIR) photobiomodulation (PBM) and Coenzyme Q10 (CoQ10) administration are known to stimulate mitochondrial electron transport that potentially may reverse the effects of cerebral ischemia in aged animals. We tested the hypothesis that the effects of PBM and CoQ10, separately or in combination, improve cognition in a mouse model of transient cerebral ischemia superimposed on a model of aging. We modeled aging by 6-week administration of D-galactose (500 mg/kg subcutaneous) to mice. We subsequently induced transient cerebral ischemia by bilateral occlusion of the common carotid artery (BCCAO). We treated the mice with PBM (810 nm transcranial laser) or CoQ10 (500 mg/kg by gavage), or both, for 2 weeks after surgery. We assessed cognitive function by the Barnes and Lashley III mazes and the What-Where-Which (WWWhich) task. PBM or CoQ10, and both, improved spatial and episodic memory in the mice. Separately and together, the treatments lowered reactive oxygen species and raised ATP and general mitochondrial activity as well as biomarkers of mitochondrial biogenesis, including SIRT1, PGC-1?, NRF1, and TFAM. Neuroinflammatory responsiveness declined, as indicated by decreased iNOS, TNF-?, and IL-1? levels with the PBM and CoQ10 treatments. Collectively, the findings of this preclinical study imply that the procognitive effects of NIR PBM and CoQ10treatments, separately or in combination, are beneficial in a model of transient global brain ischemia superimposed on a model of aging in mice.

KEYWORDS:

Coenzyme Q10; aging; global ischemia; learning and memory; mitochondrial biogenesis; neuroinflammation; transcranial photobiomodulation.

Lasers Med Sci. 2017 May 2. doi: 10.1007/s10103-017-2221-y. [Epub ahead of print]

Beneficial neurocognitive effects of transcranial laser in older adults.

Vargas E1, Barrett DW1, Saucedo CL1, Huang LD2, Abraham JA2, Tanaka H3, Haley AP1, Gonzalez-Lima F4.

Author information

1
Department of Psychology and Institute for Neuroscience, University of Texas at Austin, Austin, TX, 78712, USA.
2
Department of Electrical Engineering, University of Texas at Austin, Austin, TX, 78712, USA.
3
Department of Kinesiology and Health Education, University of Texas at Austin, Austin, TX, 78712, USA.
4
Department of Psychology and Institute for Neuroscience, University of Texas at Austin, Austin, TX, 78712, USA. gonzalezlima@utexas.edu.

Abstract

Transcranial infrared laser stimulation (TILS) at 1064 nm, 250 mW/cm2 has been proven safe and effective for increasing neurocognitive functions in young adults in controlled studies using photobiomodulation of the right prefrontal cortex. The objective of this pilot study was to determine whether there is any effect from TILS on neurocognitive function in older adults with subjective memory complaint at risk for cognitive decline (e.g., increased carotid artery intima-media thickness or mild traumatic brain injury). We investigated the cognitive effects of TILS in older adults (ages 49-90, n = 12) using prefrontal cortex measures of attention (psychomotor vigilance task (PVT)) and memory (delayed match to sample (DMS)), carotid artery intima-media thickness (measured by ultrasound), and evaluated the potential neural mechanisms mediating the cognitive effects of TILS using exploratory brain studies of electroencephalography (EEG, n = 6) and functional magnetic resonance imaging (fMRI, n = 6). Cognitive performance, age, and carotid artery intima-media thickness were highly correlated, but all participants improved in all cognitive measures after TILS treatments. Baseline vs. chronic (five weekly sessions, 8 min each) comparisons of mean cognitive scores all showed improvements, significant for PVT reaction time (p < 0.001), PVT lapses (p < 0.001), and DMS correct responses (p < 0.05). The neural studies also showed for the first time that TILS increases resting-state EEG alpha, beta, and gamma power and promotes more efficient prefrontal blood-oxygen-level-dependent (BOLD)-fMRI response. Importantly, no adverse effects were found. These preliminary findings support the use of TILS for larger randomized clinical trials with this non-invasive approach to augment neurocognitive function in older people to combat aging-related and vascular disease-related cognitive decline.

BBA Clin. 2016 Dec; 6: 113–124.
Published online 2016 Oct 1. doi:  10.1016/j.bbacli.2016.09.002
PMCID: PMC5066074

Shining light on the head: Photobiomodulation for brain disorders

Wellman Center for Photomedicine, Massachusetts General Hospital, Boston, MA 02114, USA
Department of Dermatology, Harvard Medical School, Boston, MA 02115, USA
Harvard-MIT Division of Health Sciences and Technology, Cambridge, MA 02139, USA
Michael R. Hamblin: ude.dravrah.hgm.xileh@nilbmaH
?Wellman Center for Photomedicine, Massachusetts General Hospital, Boston, MA 02114, USA.Wellman Center for PhotomedicineMassachusetts General HospitalBostonMA02114USA ude.dravrah.hgm.xileh@nilbmaH
Author information ? Article notes ? Copyright and License information ?
Received 2016 Sep 2; Revised 2016 Sep 27; Accepted 2016 Sep 29.

Abstract

Photobiomodulation (PBM) describes the use of red or near-infrared light to stimulate, heal, regenerate, and protect tissue that has either been injured, is degenerating, or else is at risk of dying. One of the organ systems of the human body that is most necessary to life, and whose optimum functioning is most worried about by humankind in general, is the brain. The brain suffers from many different disorders that can be classified into three broad groupings: traumatic events (stroke, traumatic brain injury, and global ischemia), degenerative diseases (dementia, Alzheimer’s and Parkinson’s), and psychiatric disorders (depression, anxiety, post traumatic stress disorder). There is some evidence that all these seemingly diverse conditions can be beneficially affected by applying light to the head. There is even the possibility that PBM could be used for cognitive enhancement in normal healthy people. In this transcranial PBM (tPBM) application, near-infrared (NIR) light is often applied to the forehead because of the better penetration (no hair, longer wavelength). Some workers have used lasers, but recently the introduction of inexpensive light emitting diode (LED) arrays has allowed the development of light emitting helmets or “brain caps”. This review will cover the mechanisms of action of photobiomodulation to the brain, and summarize some of the key pre-clinical studies and clinical trials that have been undertaken for diverse brain disorders.

Keywords: Photobiomodulation, Low level laser (light) therapy, Ischemic stroke, Traumatic brain injury, Alzheimer’s disease, Parkinson’s disease, Major depression, Cognitive enhancement

Graphical abstract

Image 2

1.?Introduction

Photobiomodulation (PBM) as it is known today (the beneficial health benefits of light therapy had been known for some time before), was accidently discovered in 1967, when Endre Mester from Hungary attempted to repeat an experiment recently published by McGuff in Boston, USA [1]. McGuff had used a beam from the recently discovered ruby laser [2], to destroy a cancerous tumor that had been experimentally implanted into a laboratory rat. However (unbeknownst to Mester) the ruby laser that had been built for him, was only a tiny fraction of the power of the laser that had previously been used by McGuff. However, instead of curing the experimental tumors with his low-powered laser, Mester succeeded in stimulating hair regrowth and wound healing in the rats, in the sites where the tumors had been implanted [3], [4]. This discovery led to a series of papers describing what Mester called “laser biostimulation”, and soon became known as “low level laser therapy” (LLLT) [5], [6], [7].

LLLT was initially primarily studied for stimulation of wound healing, and reduction of pain and inflammation in various orthopedic conditions such as tendonitis, neck pain, and carpal tunnel syndrome [8]. The advent of light emitting diodes (LED) led to LLLT being renamed as “low level light therapy”, as it became more accepted that the use of coherent lasers was not absolutely necessary, and a second renaming occurred recently [9] when the term PBM was adopted due to uncertainties in the exact meaning of “low level”.

2.?Mechanisms of action of photobiomodulation

2.1. Mitochondria and cytochrome c oxidase

The most well studied mechanism of action of PBM centers around cytochrome c oxidase (CCO), which is unit four of the mitochondrial respiratory chain, responsible for the final reduction of oxygen to water using the electrons generated from glucose metabolism [10]. The theory is that CCO enzyme activity may be inhibited by nitric oxide (NO) (especially in hypoxic or damaged cells). This inhibitory NO can be dissociated by photons of light that are absorbed by CCO (which contains two heme and two copper centers with different absorption spectra) [11]. These absorption peaks are mainly in the red (600–700 nm) and near-infrared (760–940 nm) spectral regions. When NO is dissociated, the mitochondrial membrane potential is increased, more oxygen is consumed, more glucose is metabolized and more ATP is produced by the mitochondria.

2.2. Reactive oxygen species, nitric oxide, blood flow

It has been shown that there is a brief increase in reactive oxygen species (ROS) produced in the mitochondria when they absorb the photons delivered during PBM. The idea is that this burst of ROS may trigger some mitochondrial signaling pathways leading to cytoprotective, anti-oxidant and anti-apoptotic effects in the cells [12]. The NO that is released by photodissociation acts as a vasodilator as well as a dilator of lymphatic flow. Moreover NO is also a potent signaling molecule and can activate a number of beneficial cellular pathways [13]. Fig. 2 illustrates these mechanisms.

Fig. 2

Tissue specific processes that occur after PBM and benefit a range of brain disorders. BDNF, brain-derived neurotrophic factor; LLLT, low level light therapy; NGF, nerve growth factor; NT-3, neurotrophin 3; PBM, photobiomodulation; SOD, superoxide dismutase.

2.3. Light sensitive ion channels and calcium

It is quite clear that there must be some other type of photoacceptor, in addition to CCO, as is clearly demonstrated by the fact that wavelengths substantially longer than the red/NIR wavelengths discussed above, can also produce beneficial effects is some biological scenarios. Wavelengths such as 980 nm [14], [15], 1064 nm laser [16], and 1072 nm LED [17], and even broad band IR light [18] have all been reported to carry out PBM type effects. Although the photoacceptor for these wavelengths has by no means been conclusively identified, the leading hypothesis is that it is primarily water (perhaps nanostructured water) located in heat or light sensitive ion channels. Clear changes in intracellular calcium can be observed, that could be explained by light-mediated opening of calcium ion channels, such as members of the transient receptor potential (TRP) super-family [19]. TRP describes a large family of ion channels typified by TRPV1, recently identified as the biological receptor for capsaicin (the active ingredient in hot chili peppers) [20]. The biological roles of TRP channels are multifarious, but many TRP channels are involved in heat sensing and thermoregulation [21].

2.4. Signaling mediators and activation of transcription factors

Most authors suggest that the beneficial effects of tPBM on the brain can be explained by increases in cerebral blood flow, greater oxygen availability and oxygen consumption, improved ATP production and mitochondrial activity [22], [23], [24]. However there are many reports that a brief exposure to light (especially in the case of experimental animals that have suffered some kind of acute injury or traumatic insult) can have effects lasting days, weeks or even months [25]. This long-lasting effect of light can only be explained by activation of signaling pathways and transcription factors that cause changes in protein expression that last for some considerable time. The effects of PBM on stimulating mitochondrial activity and blood flow is of itself, unlikely to explain long-lasting effects. A recent review listed no less than fourteen different transcription factors and signaling mediators, that have been reported to be activated after light exposure [10].

Fig. 1 illustrates two of the most important molecular photoreceptors or chromophores (cytochrome c oxidase and heat-gated ion channels) inside neuronal cells that absorb photons that penetrate into the brain. The signaling pathways and activation of transcription factors lead to the eventual effects of PBM in the brain.

Fig. 1

Molecular and intracellular mechanisms of transcranial low level laser (light) or photobiomodulation. AP1, activator protein 1; ATP, adenosine triphosphate; Ca2 +, calcium ions; cAMP, cyclic adenosine monophosphate; NF-kB, nuclear factor kappa

Fig. 2 illustrates some more tissue specific mechanisms that lead on from the initial photon absorption effects explained in Fig. 1. A wide variety of processes can occur that can benefit a correspondingly wide range of brain disorders. These processes can be divided into short-term stimulation (ATP, blood flow, lymphatic flow, cerebral oxygenation, less edema). Another group of processes center around neuroprotection (upregulation of anti-apoptotic proteins, less excitotoxity, more antioxidants, less inflammation). Finally a group of processes that can be grouped under “help the brain to repair itself” (neurotrophins, neurogenesis and synaptogenesis).

2.5. Biphasic dose response and effect of coherence

The biphasic dose response (otherwise known as hormesis, and reviewed extensively by Calabrese et al. [26]) is a fundamental biological law describing how different biological systems can be activated or stimulated by low doses of any physical insult or chemical substance, no matter how toxic or damaging this insult may be in large doses. The most well studied example of hormesis is that of ionizing radiation, where protective mechanisms are induced by very low exposures, that can not only protect against subsequent large doses of ionizing radiation, but can even have beneficial effects against diseases such as cancer using whole body irradiation [27].

There are many reports of PBM following a biphasic dose response (sometimes called obeying the Arndt-Schulz curve [28], [29]. A low dose of light is beneficial, but raising the dose produces progressively less benefit until eventually a damaging effect can be produced at very high light [30]. It is often said in this context that “more does not mean more”.

Another question that arises in the field of PBM is whether the coherent monochromatic lasers that were used in the original discovery of the effect, and whose use continued for many years, are superior to the rather recent introduction of LEDs, that are non-coherent and have a wider band-spread (generally 30 nm full-width half-maximum). Although there are one or two authors who continue to believe that coherent lasers are superior [31], most commentators feel that other parameters such as wavelength, power density, energy density and total energy are the most important determinants of efficacy [8].

3.?Tissue optics, direct versus systemic effects, light sources

3.1. Light penetration into the brain

Due to the growing interest in PBM of the brain, several tissue optics laboratories have investigated the penetration of light of different wavelengths through the scalp and the skull, and to what depths into the brain this light can penetrate. This is an intriguing question to consider, because at present it is unclear exactly what threshold of power density in mW/cm2 is required in the b5rain to have a biological effect. There clearly must be a minimum value below which the light can be delivered for an infinite time without doing anything, but whether this is in the region of ?W/cm2 or mW/cm2 is unknown at present.

Functional near-infrared spectroscopy (fNIRS) using 700–900 nm light has been established as a brain imaging technique that can be compared to functional magnetic resonance imaging (fMRI) [32]. Haeussinger et al. estimated that the mean penetration depth (5% remaining intensity) of NIR light through the scalp and skull was 23:6 + 0:7 mm [33]. Other studies have found comparable results with variations depending on the precise location on the head and wavelength [34], [35].

Jagdeo et al. [36] used human cadaver heads (skull with intact soft tissue) to measure penetration of 830 nm light, and found that penetration depended on the anatomical region of the skull (0.9% at the temporal region, 2.1% at the frontal region, and 11.7% at the occipital region). Red light (633 nm) hardly penetrated at all. Tedord et al. [37] also used human cadaver heads to compare penetration of 660 nm, 808 nm, and 940 nm light. They found that 808 nm light was best and could reach a depth in the brain of 40–50 mm. Lapchak et al. compared the transmission of 810 nm light through the skulls of four different species, and found mouse transmitted 40%, while for rat it was 21%, rabbit it was 11.3 and for human skulls it was only 4.2% [38]. Pitzschke and colleagues compared penetration of 670 nm and 810 nm light into the brain when delivered by a transcranial or a transphenoidal approach, and found that the best combination was 810 nm delivered transphenoidally [39]. In a subsequent study these authors compared the effects of storage and processing (frozen or formalin-fixed) on the tissue optical properties of rabbit heads [40]. Yaroslavsky et al. examined light penetration of different wavelengths through different parts of the brain tissue (white brain matter, gray brain matter, cerebellum, and brainstem tissues, pons, thalamus). Best penetration was found with wavelengths between 1000 and 1100 nm [41].

Henderson and Morries found that between 0.45% and 2.90% of 810 nm or 980 nm light penetrated through 3 cm of scalp, skull and brain tissue in ex vivo lamb heads [42].

3.2. Systemic effects

It is in fact very likely that the beneficial effects of PBM on the brain cannot be entirely explained by penetration of photons through the scalp and skull into the brain itself. There have been some studies that have explicitly addressed this exact issue. In a study of PBM for Parkinson’s disease in a mouse model [43]. Mitrofanis and colleagues compared delivering light to the mouse head, and also covered up the head with aluminum foil so that they delivered light to the remainder of the mouse body. They found that there was a highly beneficial effect on neurocognitive behavior with irradiation to the head, but nevertheless there was also a statistically significant (although less pronounced benefit, referred to by these authors as an ‘abscopal effect”) when the head was shielded from light [44]. Moreover Oron and co-workers [45] have shown that delivering NIR light to the mouse tibia (using either surface illumination or a fiber optic) resulted in improvement in a transgenic mouse model of Alzheimer’s disease (AD). Light was delivered weekly for 2 months, starting at 4 months of age (progressive stage of AD). They showed improved cognitive capacity and spatial learning, as compared to sham-treated AD mice. They proposed that the mechanism of this effect was to stimulate c-kit-positive mesenchymal stem cells (MSCs) in autologous bone marrow (BM) to enhance the capacity of MSCs to infiltrate the brain, and clear ?-amyloid plaques [46]. It should be noted that the calvarial bone marrow of the skull contains substantial numbers of stem cells [47].

3.3. Laser acupuncture

Laser acupuncture is often used as an alternative or as an addition to traditional Chinese acupuncture using needles [48]. Many of the applications of laser acupuncture have been for conditions that affect the brain [49] such as Alzheimer’s disease [50] and autism [51] that have all been investigated in animal models. Moreover laser acupuncture has been tested clinically [52].

3.4. Light sources

A wide array of different light sources (lasers and LEDs) have been employed for tPBM. One of the most controversial questions which remains to be conclusively settled, is whether a coherent monochromatic laser is superior to non-coherent LEDs typically having a 30 nm band-pass (full width half maximum). Although wavelengths in the NIR region (800–1100 nm) have been the most often used, red wavelengths have sometimes been used either alone, or in combination with NIR. Power levels have also varied markedly from Class IV lasers with total power outputs in the region of 10 W [53], to lasers with more modest power levels (circa 1 W). LEDs can also have widely varying total power levels depending on the size of the array and the number and power of the individual diodes. Power densities can also vary quite substantially from the Photothera laser [54] and other class IV lasers , which required active cooling (~ 700 mW/cm2) to LEDs in the region of 10–30 mW/cm2.

3.5. Usefulness of animal models when testing tPBM for brain disorders

One question that is always asked in biomedical research, is how closely do the laboratory models of disease (which are usually mice or rats) mimic the human disease for which new treatments are being sought? This is no less critical a question when the areas being studied include brain disorders and neurology. There now exist a plethora of transgenic mouse models of neurological disease [55], [56]. However in the present case, where the proposed treatment is almost completely free of any safety concerns, or any reported adverse side effects, it can be validly questioned as to why the use of laboratory animal models should be encouraged. Animal models undoubtedly have disadvantages such as failure to replicate all the biological pathways found in human disease, difficulty in accurately measuring varied forms of cognitive performance, small size of mice and rats compared to humans, short lifespan affecting the development of age related diseases, and lack of lifestyle factors that adversely affect human diseases. Nevertheless, small animal models are less expensive, and require much less time and effort to obtain results than human clinical trials, so it is likely they will continue to be used to test tPBM for the foreseeable future.

4.?PBM for stroke

4.1. Animal models

Perhaps the most well-investigated application of PBM to the brain, lies in its possible use as a treatment for acute stroke [57]. Animal models such as rats and rabbits, were first used as laboratory models, and these animals had experimental strokes induced by a variety of methods and were then treated with light (usually 810 nm laser) within 24 h of stroke onset [58]. In these studies intervention by tLLLT within 24 h had meaningful beneficial effects. For the rat models, stroke was induced by middle cerebral artery occlusion (MCAO) via an insertion of a filament into the carotid artery or via craniotomy [59], [60]. Stroke induction in the “rabbit small clot embolic model” (RSCEM) was by injection of a preparation of small blood clots (made from blood taken from a second donor rabbit) into a catheter placed in the right internal carotid artery [61]. These studies and the treatments and results are listed in Table 1.

Table 1

Reports of transcranial LLLT used for stroke in animal models.

CW, continuous wave; LLLT, low level light therapy; MCAO, middle cerebral artery occlusion; NOS, nitric oxide synthase; RSCEM, rabbit small clot embolic model; TGF?1, transforming growth factor ?1.

4.2. Clinical trials for acute stroke

Treatment of acute stroke was addressed in a series of three clinical trials called “Neurothera Effectiveness and Safety Trials” (NEST-1 [65], NEST-2 [66], and NEST-3 [67]) using an 810 nm laser applied to the shaved head within 24 h of patients suffering an ischemic stroke. The first study, NEST-1, enrolled 120 patients between the ages of 40 to 85 years of age with a diagnosis of ischemic stroke involving a neurological deficit that could be measured. The purpose of this first clinical trial was to demonstrate the safety and effectiveness of laser therapy for stroke within 24 h [65]. tPBM significantly improved outcome in human stroke patients, when applied at ~ 18 h post-stroke, over the entire surface of the head (20 points in the 10/20 EEG system) regardless of stroke [65]. Only one laser treatment was administered, and 5 days later, there was significantly greater improvement in the Real- but not in the Sham-treated group (p < 0.05, NIH Stroke Severity Scale). This significantly greater improvement was still present at 90 days post-stroke, where 70% of the patients treated with Real-LLLT had a successful outcome, while only 51% of Sham-controls did. The second clinical trial, NEST-2, enrolled 660 patients, aged 40 to 90, who were randomly assigned to one of two groups (331 to LLLT, 327 to sham) [68]. Beneficial results (p < 0.04) were found for the moderate and moderate-severe (but not for the severe) stroke patients, who received the Real laser protocol [68]. These results suggested that the overall severity of the individual stroke should be taken into consideration in future studies, and very severe patients are unlikely to recover with any kind of treatment. The last clinical trial, NEST-3, was planned for 1000 patients enrolled. Patients in this study were not to receive tissue plasminogen activator, but the study was prematurely terminated by the DSMB for futility (an expected lack of statistical significance) [67]. NEST-1 was considered successful, even though as a phase 1 trial, it was not designed to show efficacy. NEST-2 was partially successful when the patients were stratified, to exclude very severe strokes or strokes deep within the brain [66]. There has been considerable discussion in the scientific literature on precisely why the NEST-3 trial failed [69]. Many commentators have wondered how could tPBM work so well in the first trial, in a sub-group in the second trial, and fail in the third trial. Lapchak’s opinion is that the much thicker skull of humans compared to that of the other animals discussed above (mouse, rat and rabbit), meant that therapeutically effective amounts of light were unlikely to reach the brain [69]. Moreover the time between the occurrence of a stroke and initiation of the PBMT may be an important factor. There are reports in the literature that neuroprotection must be administered as soon as possible after a stroke [70], [71]. Furthermore, stroke trials in particular should adhere to the RIGOR (rigorous research) guidelines and STAIR (stroke therapy academic industry roundtable) criteria [72]. Other contributory causes to the failure of NEST-3 may have been included the decision to use only one single tPBM treatment, instead of a series of treatments. Moreover, the optimum brain areas to be treated in acute stroke remain to be determined. It is possible that certain areas of the brain that have sustained ischemic damage should be preferentially illuminated and not others.

4.3. Chronic stroke

Somewhat surprisingly, there have not as yet been many trials of PBM for rehabilitation of stroke patients with only the occasional report to date. Naeser reported in an abstract the use of tPBM to treat chronic aphasia in post-stroke patients [73]. Boonswang et al. [74] reported a single patient case in which PBM was used in conjunction with physical therapy to rehabilitate chronic stroke damage. However the findings that PBM can stimulate synaptogenesis in mice with TBI, does suggest that tPBM may have particular benefits in rehabilitation of stroke patients. Norman Doidge, in Toronto, Canada has described the use of PBM as a component of a neuroplasticity approach to rehabilitate chronic stroke patients [75].

5. PBM for traumatic brain injury (TBI)

5.1. Mouse and rat models

There have been a number of studies looking at the effects of PBM in animal models of TBI. Oron’s group was the first [76] to demonstrate that a single exposure of the mouse head to a NIR laser (808 nm) a few hours after creation of a TBI lesion could improve neurological performance and reduce the size of the brain lesion. A weight-drop device was used to induce a closed-head injury in the mice. An 808 nm diode laser with two energy densities (1.2–2.4 J/cm2 over 2 min of irradiation with 10 and 20 mW/cm2) was delivered to the head 4 h after TBI was induced. Neurobehavioral function was assessed by the neurological severity score (NSS). There were no significant difference in NSS between the power densities (10 vs 20 mW/cm2) or significant differentiation between the control and laser treated group at early time points (24 and 48 h) post TBI. However, there was a significant improvement (27% lower NSS score) in the PBM group at times of 5 days to 4 weeks. The laser treated group also showed a smaller loss of cortical tissue than the sham group [76].

Hamblin’s laboratory then went on (in a series of papers [76]) to show that 810 nm laser (and 660 nm laser) could benefit experimental TBI both in a closed head weight drop model [77], and also in controlled cortical impact model in mice [25]. Wu et al. [77] explored the effect that varying the laser wavelengths of LLLT had on closed-head TBI in mice. Mice were randomly assigned to LLLT treated group or to sham group as a control. Closed-head injury (CHI) was induced via a weight drop apparatus. To analyze the severity of the TBI, the neurological severity score (NSS) was measured and recorded. The injured mice were then treated with varying wavelengths of laser (665, 730, 810 or 980 nm) at an energy level of 36 J/cm2 at 4 h directed onto the scalp. The 665 nm and 810 nm groups showed significant improvement in NSS when compared to the control group at day 5 to day 28. Results are shown in Fig. 3. Conversely, the 730 and 980 nm groups did not show a significant improvement in NSS and these wavelengths did not produce similar beneficial effects as in the 665 nm and 810 nm LLLT groups [77]. The tissue chromophore cytochrome c oxidase (CCO) is proposed to be responsible for the underlying mechanism that produces the many PBM effects that are the byproduct of LLLT. COO has absorption bands around 665 nm and 810 nm while it has low absorption bands at the wavelength of 730 nm [78]. It should be noted that this particular study found that the 980 nm did not produce the same positive effects as the 665 nm and 810 nm wavelengths did; nevertheless previous studies did find that the 980 nm wavelength was an active one for LLLT. Wu et al. proposed that these dissimilar results may be due to the variance in the energy level, irradiance, etc. between the other studies and this particular study [77].

Fig. 3

tPBM for TBI in a mouse model. Mice received a closed head injury and 4 hours later a single exposure of the head to one of four different lasers (36 J/cm2 delivered at 150 mW/cm2 over 4 min with spot size 1-cm diameter)

Ando et al. [25] used the 810 nm wavelength laser parameters from the previous study and varied the pulse modes of the laser in a mouse model of TBI. These modes consisted of either pulsed wave at 10 Hz or at 100 Hz (50% duty cycle) or continuous wave laser. For the mice, TBI was induced with a controlled cortical impact device via open craniotomy. A single treatment with an 810 nm Ga-Al-As diode laser with a power density of 50 mW/m2 and an energy density of 36 J/cm2 was given via tLLLT to the closed head in mice for a duration of 12 min at 4 h post CCI. At 48 h to 28 days post TBI, all laser treated groups had significant decreases in the measured neurological severity score (NSS) when compared to the control (Fig. 4A). Although all laser treated groups had similar NSS improvement rates up to day 7, the PW 10 Hz group began to show greater improvement beyond this point as seen in Fig. 4. At day 28, the forced swim test for depression and anxiety was used and showed a significant decrease in the immobility time for the PW 10 Hz group. In the tail suspension test which measures depression and anxiety, there was also a significant decrease in the immobility time at day 28, and this time also at day 1, in the PW 10 Hz group.

Fig. 4

tPBM for controlled cortical impact TBI in a mouse model. (A) Mice received a single exposure (810 nm laser, 36 J/cm2 delivered at 50 mW/cm2 over 12 min) [121]. (B) Mice received 3 daily exposures starting 4 h post-TBI

Studies using immunofluorescence of mouse brains showed that tPBM increased neuroprogenitor cells in the dentate gyrus (DG) and subventricular zone at 7 days after the treatment [79]. The neurotrophin called brain derived neurotrophic factor (BDNF) was also increased in the DG and SVZ at 7 days , while the marker (synapsin-1) for synaptogenesis and neuroplasticity was increased in the cortex at 28 days but not in the DG, SVZ or at 7 days [80] (Fig. 4B). Learning and memory as measured by the Morris water maze was also improved by tPBM [81]. Whalen’s laboratory [82] and Whelan’s laboratory [83] also successfully demonstrated therapeutic benefits of tPBM for TBI in mice and rats respectively.

Zhang et al. [84] showed that secondary brain injury occurred to a worse degree in mice that had been genetically engineered to lack “Immediate Early Response” gene X-1 (IEX-1) when exposed to a gentle head impact (this injury is thought to closely resemble mild TBI in humans). Exposing IEX-1 knockout mice to LLLT 4 h post injury, suppressed proinflammatory cytokine expression of interleukin (IL)-I? and IL-6, but upregulated TNF-?. The lack of IEX-1 decreased ATP production, but exposing the injured brain to LLLT elevated ATP production back to near normal levels.

Dong et al. [85] even further improved the beneficial effects of PBM on TBI in mice, by combining the treatment with metabolic substrates such as pyruvate and/or lactate. The goal was to even further improve mitochondrial function. This combinatorial treatment was able to reverse memory and learning deficits in TBI mice back to normal levels, as well as leaving the hippocampal region completely protected from tissue loss; a stark contrast to that found in control TBI mice that exhibited severe tissue loss from secondary brain injury.

5.2. TBI in humans

Margaret Naeser and collaborators have tested PBM in human subjects who had suffered TBI in the past [86]. Many sufferers from severe or even moderate TBI, have very long lasting and even life-changing sequelae (headaches, cognitive impairment, and difficulty sleeping) that prevent them working or living any kind or normal life. These individuals may have been high achievers before the accident that caused damage to their brain [87]. Initially Naeser published a report [88] describing two cases she treated with PBM applied to the forehead twice a week. A 500 mW continuous wave LED source (mixture of 660 nm red and 830 nm NIR LEDs) with a power density of 22.2 mW/cm2 (area of 22.48 cm2), was applied to the forehead for a typical duration of 10 min (13.3 J/cm2). In the first case study the patient reported that she could concentrate on tasks for a longer period of time (the time able to work at a computer increased from 30 min to 3 h). She had a better ability to remember what she read, decreased sensitivity when receiving haircuts in the spots where LLLT was applied, and improved mathematical skills after undergoing LLLT. The second patient had statistically significant improvements compared to prior neuropsychological tests after 9 months of treatment. The patient had a 2 standard deviation (SD) increase on tests of inhibition and inhibition accuracy (9th percentile to 63rd percentile on the Stroop test for executive function and a 1 SD increase on the Wechsler Memory scale test for the logical memory test (83rd percentile to 99th percentile) [89].

Naeser et al. then went on to report a case series of a further eleven patients [90]. This was an open protocol study that examined whether scalp application of red and near infrared (NIR) light could improve cognition in patients with chronic, mild traumatic brain injury (mTBI). This study had 11 participants ranging in age from 26 to 62 (6 males, 5 females) who suffered from persistent cognitive dysfunction after mTBI. The participants’ injuries were caused by motor vehicle accidents, sports related events and for one participant, an improvised explosive device (IED) blast. tLLLT consisted of 18 sessions (Monday, Wednesday, and Friday for 6 weeks) and commenced anywhere from 10 months to 8 years post-TBI. A total of 11 LED clusters (5.25 cm in diameter, 500 mW, 22.2 mW/cm2, 13 J/cm2) were applied for about 10 min per session (5 or 6 LED placements per set, Set A and then Set B, in each session). Neuropsychological testing was performed pre-LED application and 1 week, 1 month and 2 months after the final treatment. Naeser and colleagues found that there was a significant positive linear trend observed for the Stroop Test for executive function, in trial 2 inhibition (p = 0.004); Stroop, trial 4 inhibition switching (p = 0.003); California Verbal Learning Test (CVLT)-II, total trials 1–5 (p = 0.003); CVLT-II, long delay free recall (p = 0.006). Improved sleep and fewer post-traumatic stress disorder (PTSD) symptoms, if present beforehand, were observed after treatment. Participants and family members also reported better social function and a better ability to perform interpersonal and occupational activities. Although these results were significant, further placebo-controlled studies will be needed to ensure the reliability of this these data [90].

Henderson and Morries [91] used a high-power NIR laser (10–15 W at 810 and 980 nm) applied to the head to treat a patient with moderate TBI. The patient received 20 NIR applications over a 2-month period. They carried out anatomical magnetic resonance imaging (MRI) and perfusion single-photon emission computed tomography (SPECT). The patient showed decreased depression, anxiety, headache, and insomnia, whereas cognition and quality of life improved, accompanied by changes in the SPECT imaging.

6. PBM for Alzheimer’s disease (AD)

6.1. Animal models

There was a convincing study [92] carried out in an A?PP transgenic mouse of AD. tPBM (810 nm laser) was administered at different doses 3 times/week for 6 months starting at 3 months of age. The numbers of A? plaques were significantly reduced in the brain with administration of tPBM in a dose-dependent fashion. tPBM mitigated the behavioral effects seen with advanced amyloid deposition and reduced the expression of inflammatory markers in the transgenic mice. In addition, TLT showed an increase in ATP levels, mitochondrial function, and c-fos expression suggesting that there was an overall improvement in neurological function.

6.2. Humans

There has been a group of investigators in Northern England who have used a helmet built with 1072 nm LEDs to treat AD, but somewhat surprisingly no peer-reviewed publications have described this approach [93]. However a small pilot study (19 patients) that took the form of a randomized placebo-controlled trial investigated the effect of the Vielight Neuro system (see Fig. 5A) (a combination of tPBM and intranasal PBM) on patients with dementia and mild cognitive impairment [94]. This was a controlled single blind pilot study in humans to investigate the effects of PBM on memory and cognition. 19 participants with impaired memory/cognition were randomized into active and sham treatments over 12 weeks with a 4-week no-treatment follow-up period. They were assessed with MMSE and ADAS-cog scales. The protocol involved in-clinic use of a combined transcranial-intranasal PBM device; and at-home use of an intranasal-only PBM device and participants/ caregivers noted daily experiences in a journal. Active participants with moderate to severe impairment (MMSE scores 5–24) showed significant improvements (5-points MMSE score) after 12 weeks. There was also a significant improvement in ADAS-cog scores (see Fig. 5B). They also reported better sleep, fewer angry outbursts and decreased anxiety and wandering. Declines were noted during the 4-week no-treatment follow-up period. Participants with mild impairment to normal (MMSE scores of 25 to 30) in both the active and sham sub-groups showed improvements. No related adverse events were reported.

Fig. 5

tPBM for Alzheimer’s disease. (A) Nineteen patients were randomized to receive real or sham tPBM (810 nm LED, 24.6 J/cm2 at 41 mW/cm2). (B) Significant decline in ADAS-cog (improved cognitive performance) in real but not sham (unpublished

An interesting paper from Russia [95] described the use of intravascular PBM to treat 89 patients with AD who received PBM (46 patients) or standard treatment with memantine and rivastigmine (43 patients). The PBM consisted of threading a fiber-optic through a cathéter in the fémoral artery and advancing it to the distal site of the anterior and middle cerebral arteries and delivering 20 mW of red laser for 20–40 min. The PBM group had improvement in cerebral microcirculation leading to permanent (from 1 to 7 years) reduction in dementia and cognitive recovery.

7. Parkinson’s disease

The majority of studies on PBM for Parkinson’s disease have been in animal models and have come from the laboratory of John Mitrofanis in Australia [96]. Two basic models of Parkinson’s disease were used. The first employed administration of the small molecule (MPTP or 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) to mice [97]. MPTP was discovered as an impurity in an illegal recreational drug to cause Parkinson’s like symptoms (loss of substantia nigra cells) in young people who had taken this drug [98]. Mice were treated with tPBM (670-nm LED, 40 mW/cm2, 3.6 J/cm2) 15 min after each MPTP injection repeated 4 times over 30 h. There were significantly more (35%–45%) dopaminergic cells in the brains of the tPBM treated mice [97]. A subsequent study showed similar results in a chronic mouse model of MPTP-induced Parkinson’s disease [99]. They repeated their studies in another mouse model of Parkinson’s disease, the tau transgenic mouse strain (K3) that has a progressive degeneration of dopaminergic cells in the substantia nigra pars compacta (SNc) [100]. They went on to test a surgically implanted intracranial fiber designed to deliver either 670 nm LED (0.16 mW) or 670 nm laser (67 mW) into the lateral ventricle of the brain in MPTP-treated mice [101]. Both low power LED and high power laser were effective in preserving SNc cells, but the laser was considered to be unsuitable for long-term use (6 days) due to excessive heat production. As mentioned above, these authors also reported a protective effect of abscopal light exposure (head shielded) in this mouse model [43]. Recently this group has tested their implanted fiber approach in a model of Parkinson’s disease in adult Macaque monkeys treated with MPTP [102]. Clinical evaluation of Parkinson’s symptoms (posture, general activity, bradykinesia, and facial expression) in the monkeys were improved at low doses of light (24 J or 35 J) compared to high doses (125 J) [103].

The only clinical report of PBM for Parkinson’s disease in humans was an abstract presented in 2010 [104]. Eight patients between 18 and 80 years with late stage PD participated in a non-controlled, non-randomized study. Participants received tPBM treatments of the head designed to deliver light to the brain stem, bilateral occipital, parietal, temporal and frontal lobes, and treatment along the sagittal suture. A Visual Analog Scale (VAS), was used to record the severity of their symptoms of balance, gait, freezing, cognitive function, rolling in bed, and difficulties with speech pre-procedure and at study endpoint with 10 being most severe and 0 as no symptom. Compared with baseline, all participants demonstrated a numerical improvement in the VAS from baseline to study endpoint. A statistically significant reduction in VAS rating for gait and cognitive function was observed with average mean change of —1.87 (p < 0.05) for gait and a mean reduction of —2.22 (p < 0.05) for cognitive function. Further, freezing and difficulty with speech ratings were significantly lower (mean reduction of 1.28 (p < 0.05) for freezing and 2.22 (p < 0.05) for difficulty with speech).

8. PBM for psychiatric disorders

8.1. Animal models

A common and well-accepted animal model of depression is called “chronic mild stress” [105]. After exposure to a series of chronic unpredictable mild stressors, animals develop symptoms seen in human depression, such as anhedonia (loss of the capacity to experience pleasure, a core symptom of major depressive disorder), weight loss or slower weight gain, decrease in locomotor activity, and sleep disorders [106]. Wu et al. used Wistar rats to show that after 5 weeks of chronic stress, application of tPBM 3 times a week for 3 weeks (810 nm laser, 100 Hz with 20% duty cycle, 120 J/cm2) gave significant improvement in the forced swimming test (FST) [107]. In a similar study Salehpour et al. [108] compared the effects of two different lasers (630 m nm at 89 mW/cm2, and 810 nm at 562 mW/cm2, both pulsed at 10 Hz, 50% duty cycle). The 810 nm laser proved better than the 630 nm laser in the FST, in the elevated plus maze and also reduced blood cortisol levels.

8.2. Depression and anxiety

The first clinical study in depression and anxiety was published by Schiffer et al. in 2009 [109]. They used a fairly small area 1 W 810 nm LED array (see Fig. 6A) applied to the forehead in patients with major depression and anxiety. They found improvements in the Hamilton depression rating scale (HAM-D) (see Fig. 6B), and the Hamilton anxiety rating scale (HAM-A), 2 weeks after a single treatment. They also found increases in frontal pole regional cerebral blood flow (rCBF) during the light delivery using a commercial NIR spectroscopy device. Cassano and co-workers [110] used tPBM with an 810 nm laser (700 mW/cm2and a fluence of 84  J/cm2 delivered per session for 6 sessions in patients with major depression. Baseline mean HAM-D17 scores decreased from 19.8 ± 4.4 (SD) to 13 ± 5.35 (SD) after treatment (p = 0.004).

Fig. 6

tPBM for major depression and anxiety in humans. (A) Ten patients received a single exposure to the forehead (810 LED, 60 J/cm2delivered at 250 mW/cm2). (B) Mean Hamilton score for depression at baseline and at two weeks post-treatment

9. Cognitive enhancement

From what we have seen above, it need come as no surprise, to learn that there are several reports about cognitive enhancement in normal people or healthy animals using PBM. The first report was in middle aged (12 months) CD1 female mice [111]. Exposure of the mice to 1072 nm LED arrays led to improved performance in a 3D maze compared to sham treated age-matched controls. Francisco Gonzalez-Lima at the University of Texas Austin, has worked in this area for some time [112]. Working in rats they showed that transcranial PBM (9 mW/cm2 with 660 nm LED array) induced a dose-dependent increase in oxygen consumption of 5% after 1 J/cm2 and 16% after 5 J/cm2 [113]. They also found that tPBM reduced fear renewal and prevented the reemergence of extinguished conditioned fear responses [113]. In normal human volunteers they used transcranial PBM (1064 nm laser, 60 J/cm2 at 250 mW/cm2) delivered to the forehead in a placebo-controlled, randomized study, to influence cognitive tasks related to the prefrontal cortex, including a psychomotor vigilance task (PVT), a delayed match-to-sample (DMS) memory task, and the positive and negative affect schedule (PANAS-X) to show improved mood [16]. Subsequent studies in normal humans showed that tPBM with 1064 nm laser could improve performance in the Wisconsin Card Sorting Task (considered the gold standard test for executive function) [114]. They also showed that tPBM to the right forehead (but not the left forehead) had better effects on improving attention bias modification (ABM) in humans with depression [115].

A study by Salgado et al. used transcranial LED PBM on cerebral blood flow in healthy elderly women analyzed by transcranial Doppler ultrasound (TCD) of the right and left middle cerebral artery and basilar artery. Twenty-five non-institutionalized elderly women (mean age 72 years old), with cognitive status > 24, were assessed using TCD before and after transcranial LED therapy. tPBM (627 nm, 70 mW/cm2, 10 J/cm2) was performed at four points of the frontal and parietal region for 30 s each twice a week for 4 weeks. There was a significant increase in the systolic and diastolic velocity of the left middle cerebral artery (25 and 30%, respectively) and the basilar artery (up to 17 and 25%), as well as a decrease in the pulsatility index and resistance index values of the three cerebral arteries analyzed [116].

10. Conclusion

Many investigators believe that PBM for brain disorders will become one of the most important medical applications of light therapy in the coming years and decades. Despite the efforts of “Big Pharma”, prescription drugs for psychiatric disorders are not generally regarded very highly (either by the medical profession or by the public), and many of these drugs perform little better than placebos in different trials, and moreover can also have major side-effects [117]. Moreover it is well accepted that with the overall aging of the general population, together with ever lengthening life spans, that dementia, Alzheimer’s, and Parkinson’s diseases will become a global health problem [118], [119]. Even after many years of research, no drug has yet been developed to benefit these neurodegenerative disorders. A similar state of play exists with drugs for stroke (with the exception of clot-busting enzymes) and TBI. New indications for tPBM such as global ischemia (brain damage after a heart attack), post-operative cognitive dysfunction [120], and neurodevelopmental disorders such as autism spectrum disorder may well emerge. Table 2 shows the wide range of brain disorders and diseases that may eventually be treated by some kind of tPBM, whether that be an office/clinic based procedure or a home-use based device. If inexpensive LED helmets can be developed and successfully marketed as home use devices, then we are potentially in a position to benefit large numbers of patients (to say nothing of healthy individuals). Certainly the advent of the Internet has made it much easier for knowledge about this kind of home treatment to spread (almost by word of mouth so to speak).

Table 2

List of brain disorders that may in principle be treated by tPBM.

Conflict of interest statement

The author declares no conflict of interest.

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Acknowledgments

MRH was supported by the US NIH grants R01AI050875 and R21AI121700, the Air Force Office of Scientific Research grant FA9550-13-1-0068, the US Army Medical Research Acquisition Activity grant W81XWH-09-1-0514, and by the US Army Medical Research and Materiel Command grant W81XWH-13-2-0067.

Footnotes

The Transparency document associated with this article can be found, in online version.

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Lasers Med Sci. 2016 Aug;31(6):1151-60. doi: 10.1007/s10103-016-1962-3. Epub 2016 May 25.

Cognitive enhancement by transcranial laser stimulation and acute aerobic exercise.

Hwang J1, Castelli DM1, Gonzalez-Lima F2.

Author information

1
Department of Kinesiology and Health Education, University of Texas at Austin, Austin, TX, 78712, USA.
2
Department of Psychology and Institute for Neuroscience, University of Texas at Austin, 108 E. Dean Keeton Stop A8000, Austin, TX, 78712, USA. gonzalezlima@utexas.edu.

Abstract

This is the first randomized, controlled study comparing the cognitive effects of transcranial laser stimulation and acute aerobic exercise on the same cognitive tasks. We examined whether transcranial infrared laser stimulation of the prefrontal cortex, acute high-intensity aerobic exercise, or the combination may enhance performance in sustained attention and working memory tasks. Sixty healthy young adults were randomly assigned to one of the following four treatments: (1) lowlevel laser therapy (LLLT) with infrared laser to two forehead sites while seated (total 8 min, 1064 nm continuous wave, 250 mW/cm(2), 60 J/cm(2) per site of 13.6 cm(2)); (2) acute exercise (EX) of high-intensity (total 20 min, with 10-min treadmill running at 85-90 % VO2max); (3) combined treatment (LLLT + EX); or (4) sham control (CON). Participants were tested for prefrontal measures of sustained attention with the psychomotor vigilance task (PVT) and working memory with the delayed match-to-sample task (DMS) before and after the treatments. As compared to CON, both LLLT and EX reduced reaction time in the PVT [F(1.56)?=?4.134, p?=?0.01, ? (2) ?=?0.181] and increased the number of correct responses in the DMS [F(1.56)?=?4.690, p?=?0.005, ? (2) ?=?0.201], demonstrating a significant enhancing effect of LLLT and EX on cognitive performance. LLLT + EX effects were similar but showed no significantly greater improvement on PVT and DMS than LLLT or EX alone. The transcranial infrared laser stimulation and acute aerobic exercise treatments were similarly effective for cognitiveenhancement, suggesting that they augment prefrontal cognitive functions similarly.

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Curr Alzheimer Res. 2015;12(9):860-9.

Cognitive Improvement by Photic Stimulation in a Mouse Model of Alzheimer’s Disease.

Zhang Y, Wang F, Luo X, Wang L, Sun P, Wang M, Jiang Y, Zou J, Uchiumi O, Yamamoto R, Sugai T, Yamamoto K, Kato N1.

Author information

  • 1Department of Physiology, Kanazawa Medical University, Ishikawa 920-0293, Japan. kato@kanazawa-med.ac.jp.

Abstract

We previously reported that activity of the large conductance calcium-activated potassium (big-K, BK) channel is suppressed by intracellular A? in cortical pyramidal cells, and that this suppression was reversed by expression of the scaffold protein Homer1a in 3xTg Alzheimer’s disease model mice. Homer1a is known to be expressed by physiological photic stimulation (PS) as well. The possibility thus arises that PS also reverses A?-induced suppression of BK channels, and thereby improves cognition in 3xTg mice. This possibility was tested here. Chronic application of 6-hour-long PS (frequency, 2 Hz; duty cycle, about 1/10; luminance, 300 lx) daily for 4 weeks improved contextual and tone-dependent fear memory in 3xTg mice and, to a lesser extent, Morris water maze performance as well. Hippocampal long-term potentiation was also enhanced after PS. BK channel activity in cingulate cortex pyramidal cells and lateral amygdalar principal cells, suppressed in 3xTg mice, were facilitated. In parallel, neuronal excitability, elevated in 3xTg mice, was recovered to the control level. Gene expression of BK channel, as well as that of the scaffold protein Homer1a, was found decreased in 3xTg mice and reversed by PS. It is known that Homer1a is an activity-dependently inducible immediate early gene product. Consistently, our previous findings showed that Homer1a induced by electrical stimulation facilitated BK channels. By using Homer1a knockouts, we showed that the present PS-induced BK channel facilitation is mediated by Homer1a expression. We thus propose that PS might be potentially useful as a non-invasive therapeutic measure against Alzheimer’s disease.

BMC Neurosci. 2016 May 18;17(1):21. doi: 10.1186/s12868-016-0259-6.

Comparative assessment of phototherapy protocols for reduction of oxidative stress in partially transected spinal cord slices undergoing secondary degeneration.

Ashworth BE1,2, Stephens E1,2, Bartlett CA1, Serghiou S3, Giacci MK1, Williams A3, Hart NS1,4, Fitzgerald M5.

Author information

  • 1Experimental and Regenerative Neurosciences, School of Animal Biology, The University of Western Australia, Crawley, WA, Australia.
  • 2Department of Biology and Biochemistry, The University of Bath, Bath, UK.
  • 3Centre for Regenerative Medicine, University of Edinburgh, Edinburgh, UK.
  • 4Department of Biological Sciences, Macquarie University, Sydney, NSW, 2109, Australia.
  • 5Experimental and Regenerative Neurosciences, School of Animal Biology, The University of Western Australia, Crawley, WA, Australia. lindy.fitzgerald@uwa.edu.au.

Abstract

BACKGROUND:

Red/near-infrared light therapy (R/NIR-LT) has been developed as a treatment for a range of conditions, including injury to the central nervous system (CNS). However, clinical trials have reported variable or sub-optimal outcomes, possibly because there are few optimized treatment protocols for the different target tissues. Moreover, the low absolute, and wavelength dependent, transmission of light by tissues overlying the target site make accurate dosing problematic.

RESULTS:

In order to optimize light therapy treatment parameters, we adapted a mouse spinal cord organotypic culture model to the rat, and characterized myelination and oxidative stress following a partial transection injury. The ex vivo model allows a more accurate assessment of the relative effect of different illumination wavelengths (adjusted for equal quantal intensity) on the target tissue. Using this model, we assessed oxidative stress following treatment with four different wavelengths of light: 450 nm (blue); 510 nm (green); 660 nm (red) or 860 nm (infrared) at three different intensities: 1.93 × 10(16) (low); 3.85 × 10(16) (intermediate) and 7.70 × 10(16) (high) photons/cm(2)/s. We demonstrate that the most effective of the tested wavelengths to reduce immunoreactivity of the oxidative stress indicator 3-nitrotyrosine (3NT) was 660 nm. 860 nm also provided beneficial effects at all tested intensities, significantly reducing oxidative stress levels relative to control (p ? 0.05).

CONCLUSIONS:

Our results indicate that R/NIR-LT is an effective antioxidant therapy, and indicate that effective wavelengths and ranges of intensities of treatment can be adapted for a variety of CNS injuries and conditions, depending upon the transmission properties of the tissue to be treated.

J Exp Neurosci. 2016 Feb 1;10:1-19. doi: 10.4137/JEN.S33444. eCollection 2016.

Neuroprotective Effects Against POCD by Photobiomodulation: Evidence from Assembly/Disassembly of the Cytoskeleton.

Liebert AD1, Chow RT2, Bicknell BT3, Varigos E4.
Author information
1University of Sydney, Sydney, NSW, Australia.
2Brain and Mind Institute, University of Sydney, Sydney, NSW, Australia.
3Australian Catholic University, Sydney, NSW, Australia.
4Olympic Park Clinic, Melbourne, VIC, Australia.
Abstract
Postoperative cognitive dysfunction (POCD) is a decline in memory following anaesthesia and surgery in elderly patients. While often reversible, it consumes medical resources, compromises patient well-being, and possibly accelerates progression into Alzheimer’s disease. Anesthetics have been implicated in POCD, as has neuroinflammation, as indicated by cytokine inflammatory markers. Photobiomodulation (PBM) is an effective treatment for a number of conditions, including inflammation. PBM also has a direct effect on microtubule disassembly in neurons with the formation of small, reversible varicosities, which cause neural blockade and alleviation of pain symptoms. This mimics endogenously formed varicosities that are neuroprotective against damage, toxins, and the formation of larger, destructive varicosities and focal swellings. It is proposed that PBM may be effective as a preconditioning treatment against POCD; similar to the PBM treatment, protective and abscopal effects that have been demonstrated in experimental models of macular degeneration, neurological, and cardiac conditions.
Front Neurosci. 2016 Jan 11;9:500. doi: 10.3389/fnins.2015.00500. eCollection 2015.

Turning On Lights to Stop Neurodegeneration: The Potential of Near Infrared Light Therapy in Alzheimer’s and Parkinson’s Disease.

Johnstone DM1, Moro C2, Stone J1, Benabid AL2, Mitrofanis J2.
Author information
1Department of Physiology, University of Sydney Sydney, NSW, Australia.
2University Grenoble Alpes, CEA, LETI, CLINATEC, MINATEC Campus Grenoble, France.
Abstract
Alzheimer’s and Parkinson’s disease are the two most common neurodegenerative disorders. They develop after a progressive death of many neurons in the brain. Although therapies are available to treat the signs and symptoms of both diseases, the progression of neuronal death remains relentless, and it has proved difficult to slow or stop. Hence, there is a need to develop neuroprotective or disease-modifying treatments that stabilize this degeneration. Red to infrared light therapy (? = 600-1070 nm), and in particular light in the near infrared (NIr) range, is emerging as a safe and effective therapy that is capable of arresting neuronal death. Previous studies have used NIr to treat tissue stressed by hypoxia, toxic insult, genetic mutation and mitochondrial dysfunction with much success. Here we propose NIr therapy as a neuroprotective or disease-modifying treatment for Alzheimer’s and Parkinson’s patients.
J Mol Neurosci. 2014 Jul 4. [Epub ahead of print]

Low-Level Laser Therapy Ameliorates Disease Progression in a Mouse Model of Alzheimer’s Disease.

Farfara D1, Tuby H, Trudler D, Doron-Mandel E, Maltz L, Vassar RJ, Frenkel D, Oron U.

Author information

  • 1Department of Neurobiology, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.

Abstract

Low-level laser therapy (LLLT) has been used to treat inflammation, tissue healing, and repair processes. We recently reported that LLLT to the bone marrow (BM) led to proliferation of mesenchymal stem cells (MSCs) and their homing in the ischemic heart suggesting its role in regenerative medicine. The aim of the present study was to investigate the ability of LLLT to stimulate MSCs of autologous BM in order to affect neurological behavior and ?-amyloid burden in progressive stages of Alzheimer’s disease (AD) mouse model. MSCs from wild-type mice stimulated with LLLT showed to increase their ability to maturate towards a monocyte lineage and to increase phagocytosis activity towards soluble amyloid beta (A?). Furthermore, weekly LLLT to BM of AD mice for 2 months, starting at 4 months of age (progressive stage of AD), improved cognitive capacity and spatial learning, as compared to sham-treated AD mice. Histology revealed a significant reduction in A? brain burden. Our results suggest the use of LLLT as a therapeutic application in progressive stages of AD and imply its role in mediating MSC therapy in brain amyloidogenic diseases.

Front Syst Neurosci. 2014; 8: 36.
Published online 2014 Mar 14. doi:  10.3389/fnsys.2014.00036
PMCID: PMC3953713

Augmentation of cognitive brain functions with transcranial lasers

F. Gonzalez-Lima* and Douglas W. Barrett
Department of Psychology and Institute for Neuroscience, University of Texas at Austin, Austin, TX, USA
*Correspondence: ude.saxetu@amilzelaznog
This article was submitted to the journal Frontiers in Systems Neuroscience.
Edited by: Mikhail Lebedev, Duke University, USA
Reviewed by: Julio C. Rojas, University of Texas Southwestern Medical Center, USA; John Mitrofanis, University of Sydney, Australia
Author information ? Article notes ? Copyright and License information ?
Received 2014 Jan 31; Accepted 2014 Feb 27.
Keywords: cognitive enhancement, cytochrome oxidase, low-level light therapy, brain stimulation, photoneuromodulation

Discovering that transcranial infrared laser stimulation produces beneficial effects on frontal cortex functions such as sustained attention, working memory, and affective state has been groundbreaking. Transcranial laser stimulation with low-power density (mW/cm2) and high-energy density (J/cm2) monochromatic light in the near-infrared wavelengths modulates brain functions and may produce neurotherapeutic effects in a nondestructive and non-thermal manner (Lampl, 2007; Hashmi et al., 2010). Barrett and Gonzalez-Lima (2013) provided the first controlled study showing that transcranial laser stimulation improves human cognitive and emotional brain functions. But for the field of low-level light/laser therapy (LLLT), development of a model of how luminous energy from red-to-near-infrared wavelengths modulates bioenergetics began with in vitro and in vivo discoveries in the last 40 years. Previous LLLT reviews have provided extensive background about historical developments, principles and applications (Rojas and Gonzalez-Lima, 2011, 2013; Chung et al., 2012). The purpose of this paper is to provide an update on LLLT’s neurochemical mechanisms supporting transcranial laser stimulation for cognitive-enhancing applications. We will explain first LLLT’s action on brain bioenergetics, briefly describe its bioavailability and dose-response, and finish with its beneficial effects on cognitive functions. Although our focus is on prefrontal-related cognitive functions, in principle LLLT should be able to modulate other brain functions. For example, stimulating different brain regions should affect different functions related to sensory and motor systems.

Brain bioenergetics

The way that near-infrared lasers and light-emitting diodes (LEDs) interact with brain function is based on bioenergetics, a mechanism that is fundamentally different than that of other brain stimulation methods such as electric and magnetic stimulation. LLLT has been found to modulate the function of neurons in cell cultures, brain function in animals, and cognitive and emotional functions in healthy persons and clinical conditions. Photoneuromodulation involves the absorption of photons by specific molecules in neurons that activate bioenergetic signaling pathways after exposure to red-to-near-infrared light. The 600–1150 nm wavelengths allow better tissue penetration by photons because light is scattered at lower wavelengths and absorbed by water at higher wavelengths (Hamblin and Demidova, 2006). Over 25 years ago, it was found that molecules that absorb LLLT wavelengths are part of the mitochondrial respiratory enzyme cytochrome oxidase in different oxidation states (Karu et al., 2005). Thus, for red-to-near-infrared light, the primary molecular photoacceptor of photon energy is cytochrome oxidase (also called cytochrome c oxidase or cytochrome a-a3) (Pastore et al., 2000).

Therefore, photon energy absorption by cytochrome oxidase is well-established as the primary neurochemical mechanism of action of LLLT in neurons (Wong-Riley et al., 2005). The more the enzymatic activity of cytochrome oxidase increases, the more metabolic energy that is produced via mitochondrial oxidative phosphorylation. LLLT supplies the brain with metabolic energy in a way analogous to the conversion of nutrients into metabolic energy, but with light instead of nutrients providing the source for ATP-based metabolic energy (Mochizuki-Oda et al., 2002). If an effective near-infrared light energy dose is supplied, it stimulates brain ATP production (Lapchak and De Taboada, 2010) and blood flow (Uozumi et al., 2010), thereby fueling ATP-dependent membrane ion pumps, leading to greater membrane stability and resistance to depolarization, which has been shown to transiently reduce neuronal excitability (Konstantinovic et al., 2013). On the other hand, electromagnetic stimulation directly changes the electrical excitability of neurons.

A long-lasting effect is achieved by LLLT’s up-regulating the amount of cytochrome oxidase, which enhances neuronal capacity for metabolic energy production that may be used to support cognitive brain functions. In mice and rats, memory has been improved by LLLT (Michalikova et al., 2008; Rojas et al., 2012a) and by methylene blue, a drug that at low doses donates electrons to cytochrome oxidase (Rojas et al., 2012b). Near-infrared light stimulates mitochondrial respiration by donating photons to cytochrome oxidase, because cytochrome oxidase is the main acceptor of photons from red-to-near-infrared light in neurons. By persistently stimulating cytochrome oxidase activity, transcranial LLLT induces post-stimulation up-regulation of the amount of cytochrome oxidase in brain mitochondria (Rojas et al., 2012a). Therefore, LLLT may lead to the conversion of luminous energy into metabolic energy (during light exposure) and to the up-regulation of the mitochondrial enzymatic machinery to produce more energy (after light exposure).

Bioavailability and hormetic dose-response

The most abundant metalloprotein in nerve tissue is cytochrome oxidase, and its absorption wavelengths are well correlated with its enzymatic activity and ATP production (Wong-Riley et al., 2005). High LLLT bioavailability to the brain in vivo has been shown by inducing brain cytochrome oxidase activity transcranially, leading to enhanced extinction memory retention in normal rats (Rojas et al., 2012a) and improved visual discrimination in rats with impaired retinal mitochondrial function (Rojas et al., 2008). Our LLLT studies utilized varied wavelengths (633–1064 nm), daily doses (1–60 J/cm2), fractionation sessions (1–6), and power densities (2–250 mW/cm2) that identified effective LLLT parameters for rats and humans.

For example, we tested in rats the effects of different LLLT doses in vivo on brain cytochrome oxidase activity, at either 10.9, 21.6, 32.9 J/cm2, or no LLLT. Treatments were delivered for 20, 40, and 60 min via four 660-nm LED arrays with a power density of 9 mW/cm2. One day after the LLLT session, brains were extracted, frozen, sectioned, and processed for cytochrome oxidase histochemistry. A 10.9 J/cm2 dose increased cytochrome oxidase activity by 13.6%. A 21.6 J/cm2 dose produced a 10.3% increase. A non-significant cytochrome oxidase increase of 3% was found after the highest 32.9 J/cm2 dose. Responses of brain cytochrome oxidase to LLLT in vivo were characterized by hormesis, with a low dose being stimulatory, while higher doses were less effective.

The first demonstration that LLLT increased oxygen consumption in the rat prefrontal cortex in vivo was provided by Rojas et al. (2012a). Oxygen concentration in the cortex of rats was measured using fluorescence-quenching during LLLT at 9 mW/cm2 and 660 nm. LLLT induced a dose-dependent increase in oxygen consumption of 5% after 1 J/cm2 and 16% after 5 J/cm2. Since oxygen is used to form water within mitochondria in a reaction catalyzed by cytochrome oxidase, more cytochrome oxidase activity should lead to more oxygen consumption.

LLLT may offer some advantages over other types of stimulation, because LLLT non-invasively targets cytochrome oxidase, a key enzyme for energy production, with induced expression linked to energy demand. Hence LLLT is mechanistically specific and non-invasive, while transcranial magnetic stimulation may be non-specific, prolonged forehead electrical stimulation may produce muscle spasms, and deep brain or vagus nerve stimulations are invasive.

Cognitive and emotional functions

LLLT via commercial low-power sources (such as FDA-cleared laser diodes and LEDs) is a highly promising, affordable, non-pharmacological alternative for improving cognitive function. LLLT delivers safe doses of light energy that are sufficiently high to modulate neuronal functions, but low enough to not result in any damage (Wong-Riley et al., 2005). In 2002, the FDA approved LLLT for pain relief in cases of head and neck pain, arthritis and carpal tunnel syndrome (Fulop et al., 2010). LLLT has been used non-invasively in humans after ischemic stroke to improve neurological outcome (Lampl et al., 2007). It also led to improved recovery and reduced fatigue after exercise (Leal Junior et al., 2010). One LLLT stimulation session to the forehead, as reported by Schiffer et al. (2009), produced a significant antidepressant effect in depressed patients. No adverse side effects were found either immediately or at 2 or 4 weeks after LLLT. Thus, these beneficial LLLT treatments have been found to be safe in humans. Even though LLLT has been regarded as safe and received FDA approval for pain treatment, the use of transcranial lasers for cognitive augmentation should be restricted to research until further controlled studies support this application for clinical use.

We used transcranial laser stimulation to the forehead in a placebo-controlled, randomized study, to influence cognitive tasks related to the prefrontal cortex, including a psychomotor vigilance task (PVT) and a delayed match-to-sample (DMS) memory task (Barrett and Gonzalez-Lima, 2013). The PVT assesses sustained attention, with participants remaining vigilant during delay intervals, and pushing a button when a visual stimulus appears on a monitor. Our laser stimulation targeted prefrontal areas which are implicated in the sustained attentional processes of the PVT (Drummond et al., 2005). Similarly, the DMS task engages the prefrontal cortex as part of a network of frontal and parietal brain regions (Nieder and Miller, 2004).

Healthy volunteers received continuous wave near-infrared light intersecting cytochrome oxidase’s absorption spectrum, delivered to the forehead using a 1064 nm low-power laser diode (also known as “cold laser”), which maximizes tissue penetration due to its long wavelength, and has been used in humans for other indications. The power density (or irradiance), 250 mW/cm2, as well as the cumulative energy density (or fluence), 60 J/cm2, were the same that showed beneficial psychological effects in Schiffer et al. (2009). This laser exposure produces negligible heat and no physical damage at the low power level used. This laser apparatus is used safely in a clinical setting by the supplier of the laser (Cell Gen Therapeutics, HD Laser Center, Dallas, TX). Reaction time in the PVT was improved by the laser treatment, as shown by a significant pre-post reaction time change relative to the placebo group. The DMS memory task also revealed significant enhancements in measures of memory retrieval latency and number of correct trials, when comparing the LLLT-treated with the placebo group (Figure (Figure1).1). Self-reported positive and negative affective (emotional) states were also measured using the PANAS-X questionnaire before and 2 weeks after laser treatment. As compared to the placebo, treated subjects reported significantly improved affective states. We suggest that this kind of transcranial laser stimulation may serve as a non-invasive and efficacious method to augment cognitive brain functions related to attention, memory, and emotional functions.

Figure 1

Cognitive performance in the delayed match-to-sample (DMS) memory task was improved after transcranial infrared stimulation to the right forehead. The DMS task involves presentation of a visual stimulus (grid pattern) on a screen. Then the stimulus disappears,

LLLT’s bioenergetics mechanisms leading to cognitive augmentation may also be at play in its neuroprotective effects (Gonzalez-Lima et al., 2013). LLLT’s stimulation of mitochondrial respiration should improve cellular function due to increased metabolic energy, as well as cellular survival after injury, due to the antioxidant effects of increases in cytochrome oxidase and superoxide dismutase (Rojas et al., 2008).

Laser transmittance of the 1064-nm wavelength at the forehead LLLT site was estimated in a post-mortem human specimen, which showed that approximately 2% of the light passed through the frontal bone. This yielded an absorption coefficient of a = 0.24, similar to the reported a = 0.22 transmittance through cranial bone for this wavelength (Bashkatov and Genina, 2006). Thus, we estimated that about 1.2 J/cm2 of the 60 J/cm2 LLLT dose applied reached the surface of the prefrontal cortex. This value is similar to 1 J/cm2, the peak effective LLLT dose in neuron cultures for increasing cytochrome oxidase activity (Rojas and Gonzalez-Lima, 2011).

Conclusions

Transcranial absorption of photon energy by cytochrome oxidase, the terminal enzyme in mitochondrial respiration, is proposed as the bioenergetic mechanism of action of LLLT in the brain. Transcranial LLLT up-regulates cortical cytochrome oxidase and enhances oxidative phosphorylation. LLLT improves prefrontal cortex-related cognitive functions, such as sustained attention, extinction memory, working memory, and affective state. Transcranial infrared stimulation may be used efficaciously to support neuronal mitochondrial respiration as a new non-invasive, cognition-improving intervention in animals and humans. This fascinating new approach should also be able to influence other brain functions depending on the neuroanatomical site stimulated and the stimulation parameters used.

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Alzheimers Res Ther. 2014; 6(1): 2.
Published online Jan 3, 2014. doi:  10.1186/alzrt232

Photobiomodulation with near infrared light mitigates Alzheimer’s disease- related pathology in cerebral cortex – evidence from two transgenic mouse models.

Sivaraman Purushothuman,1,2 Daniel M Johnstone,corresponding author1,2 Charith Nandasena,1,2 John Mitrofanis,1,3 and Jonathan Stone1,2

Alzheimer’s disease (AD) is a chronic, debilitating neurodegenerative disease with limited therapeutic options; at present there are no treatments that prevent the physical deterioration of the brain and the consequent cognitive deficits. Histopathologically, AD is characterised by neurofibrillary tangles (NFTs) of hyperphosphorylated tau protein and amyloid-beta (A?) plaques [1,2]. The extent of these histopathological features is considered to vary with and to determine clinical disease severity [2]. While the initiating pathogenic events underlying AD are still debated, there is strong evidence to suggest that oxidative stress and mitochondrial dysfunction have important roles in the neurodegenerative cascade [35]. Therefore, it has been proposed that targeting mitochondrial dysfunction could prove valuable for AD therapeutics [6].

One safe, simple yet effective approach to the repair of damaged mitochondria is photobiomodulation with near-infrared light (NIr). This treatment, which involves the irradiation of tissue with low intensity light in the red to near-infrared wavelength range (600 to 1000 nm), was originally pioneered for the healing of superficial wounds [7] but has been recently shown to have efficacy in protecting the central nervous system. While the mechanism of action remains to be elucidated, there is evidence that NIr preserves and restores cellular function by reversing dysfunctional mitochondrial cytochrome c oxidase (COX) activity, thereby mitigating the production of reactive oxygen species and restoring ATP production to normal levels [8,9].

To date, NIr treatment has yielded neuroprotective outcomes in animal models of retinal damage [9,10], traumatic brain injury [11,12], Parkinson’s disease [1315] and AD [16,17]. Furthermore, NIr therapy has yielded beneficial outcomes in clinical trials of human patients with mild to moderate stroke [18] and depression [19]. This treatment represents a promising alternative to drug therapy because it is safe, easy to apply and has no known side-effects at levels even higher than optimal doses [20].

The aim of this study was to assess the efficacy of NIr in mitigating the brain pathology and associated cellular damage that characterise AD. We utilised two mouse models, each manifesting distinct AD-related pathologies: the K3 tau transgenic model, which develops NFTs [21,22]; and the APP/PS1 transgenic model, which develops amyloid plaques [23]. Here, we present histochemical evidence that NIr treatment over a period of 1 month reduces the severity of AD-related pathology and oxidative stress and restores mitochondrial function in brain regions susceptible to neurodegeneration in AD, specifically the neocortex and hippocampus. The findings extend our previous NIr work in models of acute neurodegeneration [13,14] to demonstrate that NIr is also effective in protecting the brain against chronic insults due to AD-related genetic aberrations, a pathogenic mechanism that is likely to more closely model the human neurodegenerative condition.:

Methods

Mouse models

The K3 transgenic mouse model, originally generated as a model of frontotemporal dementia [21,22], harbours a human tau gene with the pathogenic K369I mutation; expression is driven by the neuron-specific mThy1.2 promoter. This model manifests high levels of hyperphosphorylated tau and NFTs by 2 to 3 months of age and cognitive deficits by about 4 months of age [21,22]. We commenced our experiments on K3 mice and matched C57BL/6 wildtype (WT) controls at 5 months of age, when significant neuropathology is already present.

The APPswe/PSEN1dE9 (APP/PS1) transgenic mouse model, obtained from the Jackson Laboratory (Stock number 004462; Bar Harbor, ME, USA), harbours two human transgenes: the amyloid beta precursor protein gene (APP) containing the Swedish mutation; and the presenilin-1 gene (PS1) containing a deletion of exon 9 [23]. The APP/PS1 mice exhibit increased A? and amyloid plaques by 4 months of age [24] and cognitive deficits by 6 months of age [25]. We commenced our experiments on APP/PS1 mice and matched C57BL/6 × C3H WT controls at 7 months of age, when numerous amyloid plaques and associated cognitive deficits are present.

Genotyping of mice was achieved by extracting DNA from tail tips through a modified version of the Hot Shot preparation method [26] and amplifying the transgene sequence by polymerase chain reaction. As reported previously, K3 mice were identified using the primers 5-GGGTGTCTCCAATGCCTGCTTCTTCAG-3 (forward) and 5-AAGTCACCCAGCAGGGAGGTGCTCAG-3 (reverse) [21,22] and APP/PS1 mice were genotyped using primers 5-AGGACTGACCACTCGACCAG-3 (forward) and 5-CGGGGGTCTAGTTCTGCAT-3 (reverse) [23].

Experimental design

For each series of experiments on K3 mice (aged 5 months) or APP/PS1 mice (aged 7 months) there were three experimental groups: untreated WT mice, untreated transgenic mice and NIr-treated transgenic mice (n = 5 mice per experimental group for the K3 series, 15 mice in total; n = 6 mice per experimental group for the APP/PS1 series, 18 mice in total). Our design did not include a WT control group exposed to NIr because NIr has no detectable impact on the survival and function of cells in normal healthy brain [1315]. Given the consistency of the previous results, use of animals for this extra control group did not seem justified [27].

Mice in the NIr-treated groups were exposed to one 90-second cycle of NIr (670 nm) from a light-emitting device (LED) (WARP 10; Quantum Devices, Barneveld, WI, USA) for 5 days per week over 4 consecutive weeks. Light energy emitted from the LED during each 90-second treatment equates to 4 Joule/cm2; a total of 80 Joule/cm2 was delivered to the skull over the 4 weeks. Our measurements of NIr penetration across the fur and skull of a C57BL/6 mouse indicate that ~2.5% of transmitted light reaches the cortex.

For each treatment, the mouse was restrained by hand and the LED was held 1 to 2 cm above the head. The LED light generated no heat and reliable delivery of the radiation was achieved [1315]. For the sham-treated WT, K3 and APP/PS1 groups, animals were restrained in the same way and the device was held over the head, but the light was not switched on. This treatment regime is similar to that used in previous studies where beneficial changes to neuropathology and behavioural signs were reported [1315].

Experimental animals were housed two or more to a cage and kept in a 12-hour light (<5 lux)/dark cycle at 22°C; food pellets and water were available ad libitum. All protocols were approved by the Animal Ethics Committee of the University of Sydney.

Histology and immunohistochemistry

At the end of the experimental period, mice were anaesthetised by intraperitoneal injection of sodium pentobarbital (60 mg/kg) and perfused transcardially with 4% buffered paraformaldehyde. Brains were post fixed for 3 hours, washed with phosphate-buffered saline and cryoprotected in 30% sucrose/phosphate-buffered saline. Tissue was embedded in OCT compound (ProSciTech, Thuringowa, QLD, Australia) and coronal sections of the neocortex and the hippocampus (between bregma ?1.8 and ?2.1) were cut at 20 ?m thickness on a Leica cryostat (Nussloch, Germany).

Immunohistochemistry

For most antibodies, antigen retrieval was achieved using sodium citrate buffer with 0.1% Triton. Sections were blocked in 10% normal goat serum and then incubated overnight at 4°C with a mouse monoclonal antibody – paired helical filaments-tau AT8, 1:500 (Innogenetics, Ghent, Belgium); 4-hydroxynonenal (4-HNE), 1:200 (JaICA, Fukuroi, Shizuoka, Japan); 8-hydroxy-2?-deoxyguanosine (8-OHDG), 1:200 (JaICA); COX, 1:200 (MitoSciences, Eugene, OR, USA) – and/or a rabbit polyclonal antibody (200 kDa neurofilament, 1:500; Sigma, St. Louis, MO, USA). Sections were then incubated for 3 hours at room temperature in Alexa Fluor-488 (green) and/or Alexa Fluor-594 (red) tagged secondary antibodies specific to host species of the primary antibodies (1:1,000; Molecular Probes, Carlsbad, CA, USA). Sections were then counterstained for nuclear DNA with bisbenzimide (Sigma).

Two different but complementary antibodies were used to label A? peptide: 6E10, which recognises residues 1 to 16; and 4G8, which recognises residues 17 to 24. We have previously used these two antibodies in combination to validate A? labelling, demonstrating identical labelling patterns in the rat neocortex and hippocampus [28]. For double labelling using 6E10 antibodies (1:500; Covance, Princeton, NJ, USA) and anti-glial fibrillary acidic protein antibodies (1:1,000; DAKO, Glostrup, Denmark), antigen retrieval was achieved by incubation in 90% formic acid for 10 minutes, and primary antibody incubation was carried out overnight at room temperature. For labelling using the 4G8 (1:500; Covance) antibody, slides were treated with 3% H2O2 in 50% methanol, incubated in 90% formic acid and then washed several times in dH2O before the blocking step, as described previously [28]. After blocking, sections were incubated overnight at room temperature with 4G8 antibody. Sections were then incubated in biotinylated goat anti-mouse IgG for 1 hour followed by ExtrAvidin peroxidase for 2.5 hours. The sections were then washed and developed with 3,3?-Diaminobenzidine.

Negative control sections were processed in the same fashion as described above except that primary antibodies were omitted. These control sections were immunonegative. Fluorescent images were taken using a Zeiss Apotome 2, Carl Zeiss, Oberkochen, Germany. Brightfield images were taken using a Nikon Eclipse E800, Nikon Instruments, Melville, NY, USA.

Histology

NFTs were assessed using the Bielschowsky silver staining method, as described previously [21,22]. Briefly, sections were placed in prewarmed 10% silver nitrate solution for 15 minutes, washed and then placed in ammonium silver nitrate solution at 40°C for a further 30 minutes. Sections were subsequently developed for 1 minute and then transferred to 1% ammonium hydroxide solution for 1 minute to stop the reaction. Sections were then washed in dH2O, placed in 5% sodium thiosulphate solution for 5 minutes, washed, cleared and mounted in dibutyl phathalate xylene.

As described previously [28], A? plaques were studied by staining with Congo red, a histological dye that binds preferentially to compacted amyloid with a ?-sheet secondary structure [29]. Briefly, sections were treated with 2.9 M sodium chloride in 0.01 M NaOH for 20 minutes and were subsequently stained in filtered alkaline 0.2% Congo red solution for 1 hour.

Morphological analysis

Staining intensity and area measurements

To quantify the average intensity and area of antibody labelling within the neocortex and hippocampal regions, an integrated morphology analysis was undertaken using MetaMorph software. For each section, the level of nonspecific staining (using an adjacent region of unstained midbrain) was adjusted to a set level to ensure a standard background across different groups. Next, outlines of retrosplenial cortex area 29 and hippocampal CA1 region were traced and the average intensity and area of immunostaining were calculated by the program. Measurements were conducted on ?4 representative sections per animal and ?3 animals per experimental group. Statistical analyses were performed in Prism 5.0 (Graphpad, La Jolla, CA, USA) using one-way analysis of variance with Tukey’s multiple comparison post test. All values are given as mean ± standard error of mean.

Amyloid-beta plaque measurements

Digital brightfield images of 4G8 staining in the neocortical and hippocampal regions (between bregma ?1.8 and ?2.1) were taken at 4× magnification and analysed with Metamorph, Molecular Devices LLC, Sunnyvale, CA, USA. The software was programmed to measure the number of plaques and the average size of plaques after thresholding for colour. The percentage of area covered by plaques (plaque burden) was calculated by multiplying the number of plaques by the average size of plaques, divided by the area of interest, as described previously [30]. The average number of Congo red-positive plaques in the APP/PS1 brain regions was estimated using the optical fractionator method (StereoInvestigator; MBF Science, Williston, VT, USA), as outlined previously [14]. Briefly, systematic random sampling of sites was undertaken using an unbiased counting frame (100 ?m × 100 ?m). All plaques that came into focus within the frame were counted. Measurements were conducted on ?4 representative sections per animal and ?3 animals per experimental group. Plaque numbers and size were analysed using a two-tailed unpaired t test (when variances were equal) or Welch’s t test (when variances were unequal). All values are given as mean ± standard error of mean. For all analyses, investigators were blinded to the experimental groups.

Results

Evidence of NIr-induced neuroprotection is presented from the neocortex (retrosplenial area) and the hippocampus (CA1 and subiculum), two cortical regions affected in the early stages of human AD [2].

Near-infrared light mitigates the tau pathology of K3 cortex

Hyperphosphorylation of the neuronal microtubule stabilising protein tau and the resulting NFTs are much studied features of dementia pathology [2,31]. The K3 mouse model manifests hyperphosphorylated tau and NFTs by 2 to 3 months of age and cognitive deficits by about 4 months of age [21,22]. We observe strong labelling for hyperphosphorylated tau in the neocortex and the hippocampus at 6 months of age; expression appears to plateau after this age, with similar labelling observed in 12-month-old mice (Figure 1A,B,C,D,E,F).

Figure 1

Time course of the natural development of cortical pathology in K3 and APP/PS1 mice. (A), (B), (C), (D), (E), (F) Micrographs of hyperphosphorylated tau labelling (red), using the AT8 antibody, in the neocortex (A to C) and hippocampus (D toF) of untreated

In the retrosplenial area of the neocortex there was a significant overall difference in AT8 immunolabelling for tau between the experimental groups, both when considering average intensity of labelling (P < 0.01 by analysis of variance; Figure 2A) and labelled area (P < 0.01; Figure 2B). Tukeypost hoc testing revealed significant differences between the untreated K3 group and the other two groups; labelling was much stronger and more widespread in K3 mice than WT controls (17-fold higher intensity, P < 0.01), and this labelling was reduced by over 70% in NIr-treated mice (P < 0.05). Interestingly, there was no significant difference between the WT and K3-NIr groups, suggesting that NIr treatment had reduced hyperphosphorylated tau to control levels in K3 mice. A similar trend was observed when considering the NFT pathology (Figure 2C,D,E). In contrast to WT brain, which showed no NFT-like lesions (Figure 2C), the K3 brain contained many ovoid shaped NFT-like lesions (that is, spheroids; Figure 2D). Such structures were less frequent in the K3-NIr brain (Figure 2E).

Figure 2

Effect of near-infrared light treatment on hyperphosphorylated tau and neurofibrillary tangles in the neocortex of K3 mice. (A), (B) Quantification of tau AT8 immunolabelling, based on average labelling intensity (A) and labelled area (B). All error bars

Similar effects were observed in the hippocampus (Figure 3). There was a significant overall difference between the experimental groups in AT8 immunolabelling of the CA1 pyramidal cells (P < 0.01). As for the neocortex, K3 mice showed far greater labelling than WT mice (17-fold higher intensity, P < 0.01) and this was reduced over 65% by NIr treatment (P < 0.01). Again, there were no significant differences between the WT and K3-NIr groups (P > 0.05). Bielschowsky silver staining of the subiculum (Figure 3C,D,E) revealed axonal swellings and spheroids in the hippocampal region of K3 mice (Figure 3D), which were less pronounced in mice from the K3-NIr group (Figure 3E). No pathology was observed in the hippocampus of WT mice (Figure 3C).

Figure 3

Effect of near-infrared light treatment on hyperphosphorylated tau and neurofibrillary tangles in the hippocampus of K3 mice. (A), (B) Quantification of tau AT8 immunolabelling, based on average labelling intensity (A) and labelled area (B). All error

One should note that the large white matter pathways associated with the hippocampus were labelled intensely by silver staining in all three groups (Figure 3C,D,E). This labelling has been described previously and is not associated with any neuropathology [32].

Near-infrared light reduces oxidative stress in K3 cortex

Oxidative stress and damage are common features of neurodegenerative diseases such as AD, and may be a precursor to neuronal death [35]. We assessed two common markers of oxidative stress: 4-HNE, a toxic end-product of lipid peroxidation that may bind to proteins that then trigger mitochondrial dysfunction and cellular apoptosis in AD [33]; and 8-OHDG, a marker for nuclear and mitochondrial DNA oxidation, which is elevated in AD brains [34].

Overall, 4-HNE immunoreactivity in the neocortex was significantly different between the experimental groups (Figure 4), by both average labelling intensity (P < 0.01) and labelled area (P < 0.001). As with AT8 labelling above, the K3 group showed a much higher average 4-HNE labelling intensity and area than the WT group (fivefold and 20-fold, respectively) and this labelling was significantly reduced (by 50% and 80%, respectively) in the K3-NIr group. Again, these measures showed no significant differences between the WT and K3-NIr groups (P > 0.05).

Figure 4

Effect of near-infrared light treatment on oxidative stress markers in the neocortex of K3 mice. (A), (B), (F), (G)Quantification of immunolabelling of two oxidative stress markers, 4-hydroxynonenal (4-HNE; A, B) and 8-hydroxy-2?-deoxyguanosine

Similar patterns were observed for 8-OHDG immunoreactivity. Overall, there was a significant difference between the groups for 8-OHDG immunolabelling, by both average intensity (P < 0.0001) and labelled area (P < 0.0001). Again the K3 group showed significantly higher 8-OHDG labelling intensity and area than the WT group (sixfold and 17-fold, respectively), and the 8-OHDG labelling intensity and area were significantly reduced in the K3-NIr group relative to untreated K3 (65% and 85% reduction, respectively). The intensity and area of 8-OHDG labelling did not differ significantly between the WT and the K3-NIr groups (P > 0.05), suggesting that NIr treatment reduces markers of oxidative stress to control levels. The representative photomicrographs of 8-OHDG immunoreactivity in the retrosplenial area (Figure 4H,I,J) reflect the quantitative data, with many 8-OHDG+ structures in the K3 group (Figure 4I) but not in the WT and K3-NIr groups (Figure 4H,J).

Near-infrared light mitigates mitochondrial dysfunction in K3 cortex

We assessed expression patterns of the mitochondrial enzyme COX in the neocortex and the hippocampus as a marker of mitochondrial function. Overall, there were statistically significant differences in the patterns of COX immunoreactivity between the different experimental groups, both in the neocortex and the hippocampus (both P < 0.0001; Figure 5). Relative to WT mice, the COX labelling intensity and area were reduced in K3 mice in both the neocortex and the hippocampus (>70% and >75% reductions, respectively). The K3-NIr mice showed a significant recovery of COX immunoreactivity relative to untreated K3 mice in both the neocortex (>1.7-fold increase, P < 0.05) and the hippocampus (>3.4-fold increase, P < 0.001). However, recovery was not complete, with K3-NIr mice having significantly lower COX immunoreactivity than WT mice in the neocortex (~50%, P < 0.001) and significantly lower COX labelling intensity (~20%, P < 0.05) in the hippocampus. These two groups did not differ significantly in COX labelling area in the hippocampus (P > 0.05).

Figure 5

Effect of near-infrared light treatment on cytochrome coxidase labelling in the neocortex and hippocampus of K3 mice. (A), (B), (F), (G) Quantification of immunolabelling of the mitochondrial marker cytochrome c oxidase (COX) in the neocortex retrosplenial

Near-infrared mitigates amyloid pathology in APP/PS1 cortex

Along with NFTs, A? plaques are considered a primary pathological hallmark of AD and A? load is often used as a marker of AD severity [1,35]. We assessed the distribution of A? plaques and more immature forms of the A? peptide in the neocortex and hippocampus of APP/PS1 mice aged 7 months; this age is after the first signs of intracellular A? within cells (at 3 months; Figure 1G) and extracellular A? plaques (at 4.5 and 12 months; Figure 1H and ?and1I,1I, respectively).

Three quantitative measures of plaque pathology were used: percentage plaque burden, average plaque size and number of plaques. Immunohistochemical labelling with the anti-A? antibody 4G8 revealed a significant reduction in percentage plaque burden (Figure 6A,D), average plaque size (Figure 6B,E) and number of plaques (Figure 6C,F) in both the neocortex and the hippocampus of NIr-treated APP/PS1 mice relative to untreated APP/PS1 controls. Percentage plaque burden was reduced by over 40% in the neocortex (Figure 6A; P < 0.001) and over 70% in the hippocampus (Figure 6D; P < 0.01), average plaque size was reduced 25% in the neocortex (Figure 6B) and 30% in the hippocampus (Figure 6E), and the number of plaques was reduced by over 20% in the neocortex (Figure 6C) and by over 55% in the hippocampus (Figure 6F; all P < 0.05).

Figure 6

Effect of near-infrared light on amyloid-beta and plaque pathology in APP/PS1 mice. (A), (B), (C), (D), (E), (F)Quantification of amyloid-beta (A?) 4G8 immunolabelling of amyloid plaques in the neocortex (A, B, C) and hippocampus (D, E, F), based

The photomicrographs of the 4G8 immunoreactivity in Figure 6 reflect the quantitative data described earlier. The WT brain is free of plaques (Figure 6H,K); many 4G8+ plaques (arrows) are present in the neocortex (Figure 6I) and the hippocampus (Figure 6L) of untreated APP/PS1 mice, and fewer plaques are present in NIr-treated APP/PS1 mice (Figure 6J,M). Comparable immunolabelling was achieved using the 6E10 anti-A? antibody (data not shown).

A similar but less pronounced trend was observed when staining with Congo red (Figure 7), which stains only mature plaques. Mean counts of plaques in the neocortex (Figure 7A) and the hippocampus (Figure 7B) of NIr-treated APP/PS1 brains were lower than mean counts in untreated APP/PS1 brains (reductions >30%). However, the differences did not reach statistical significance; given the findings described above with the 4G8 and 6E10 anti-A? antibodies, this suggests that NIr may have greatest effect on recently formed A? deposits. The micrographs in Figure 7 show that mature plaques were absent from the WT brain (Figure 7C,D) but were present in the neocortex (Figure 7E) and hippocampus (Figure 7F) of untreated APP/PS1 brains. There appeared to be fewer plaques in the NIr-treated APP/PS1 brains (Figure 7G,H).

Figure 7

Effect of near-infrared light on Congo red-positive plaque numbers in APP/PS1 mice. (A), (B) Quantification of Congo red-positive plaque counts in the neocortex (A) and hippocampus(B). All error bars indicate standard error of the mean. (C), (D), (E),

Discussion

Using two mouse models with distinct AD-related pathologies (tau pathology in K3, amyloid pathology in APP/PS1), we report evidence that NIr treatment can mitigate the pathology characteristic of AD as well as reduce oxidative stress and restore mitochondrial function in brain regions affected early in the disease. Further, the extent of mitigation – to levels less than at the start of treatment – suggests that NIr can reverse some elements of AD-related pathology.

The present results add to our previous findings of NIr-induced neuroprotection in models of toxin-induced acute neurodegeneration (that is, MPTP-induced parkinsonism). When incorporated into the growing body of evidence that NIr can also protect against CNS damage in models of stroke, traumatic brain injury and retinal degeneration [912,36], the findings provide a basis for trialling NIr treatment as a strategy for protection against neurodegeneration from a range of causes. Present evidence is based on the use of multiple methods, immunohistochemical and histological, to demonstrate pathological features (for example, 4G8 antibody labelling and Congo red staining for amyloid plaques, AT8 antibody labelling and Bielschowsky silver staining for NFTs).

Relationship to previous studies

The present study focused on pathological features considered characteristic of AD, as well as on signs of cellular damage (for example, oxidative stress, mitochondrial dysfunction) that have been demonstrated in AD and in animal models [24]. Our observations in the K3 strain add to previous studies by providing the first evidence in this strain of extensive oxidative damage and mitochondrial dysfunction [27].

Our findings are consistent with previous reports of the effects of red to infrared light on AD pathology in animal models. De Taboada and colleagues assessed the capacity of 808 nm laser-sourced infrared radiation, delivered three times per week over 6 months, to reduce pathology in an APP transgenic model of A? amyloidosis [17]. Treatment led to a reduction in plaque number, amyloid load and inflammatory markers, an increase in ATP levels and mitochondrial function, and mitigation of behavioural deficits. De Taboada and colleagues commenced treatment at 3 months of age, before the expected onset of amyloid pathology and cognitive effects. Similarly, Grillo and colleagues reported that 1,072 nm infrared light, applied 4 days per week for 5 months, reduces AD-related pathology in another APP/PS1 transgenic mouse model (TASTPM) [16]. These investigators also initiated light treatment before the onset of pathology, at 2 months of age. Both studies thus provide evidence that infrared radiation can slow the progression of cerebral degeneration in these models. The present results confirm these observations, in two distinct transgenic strains; they also confirm that the wound-healing and neuroprotective effects of red-infrared length do not vary qualitatively with wavelength, over a wide range.

Evidence of reversal of pathology

Previous reports have described the natural history of the K3 [21,22] and APP/PS1 transgenic models [24,37]. Based on these previous reports and our own baseline data (Figure 1), significant brain pathology and functional deficits are present in both models at the ages when we commenced treatment. Our results therefore suggest that significant reversal of pathology has been induced by the NIr treatment. This has implications for clinical practice, where most patients are not diagnosed until pathogenic mechanisms have already been initiated and resultant neurologic symptoms manifest [15,27].

This evidence that AD-related neuropathology can be transient – appear then disappear – is not novel. Garcia-Alloza and colleagues described evidence of the transient deposition of A?, including the formation of plaque-like structures, in a transgenic model of A? deposition [24]. Reversal of such pathology, by interventions such as NIr treatment, may therefore be possible. However our results suggest that reversal may also be limited to recently formed, immature plaques, as we observed a significant NIr-induced reduction in immunolabelling with the 4G8 and 6E10 antibodies but no significant difference in Congo red staining. Because the 4G8 and 6E10 antibodies recognise various forms of A?, while Congo red stains only mature, compacted plaques, a reasonable deduction is that NIr treatment reduces only the transient, recently formed A? deposits, with no substantial effect on mature plaques. As there is still no consensus as to the pathogenic roles of different forms of A?, it is unclear how this might impact on the therapeutic potential of NIr in a clinical setting.

Mechanisms

The mechanisms underlying the neuroprotective actions of red to infrared light are not completely understood. There is considerable evidence that NIr photobiomodulation enhances mitochondrial function and ATP synthesis by activating photoacceptors such as COX and increasing electron transfer in the respiratory chain, while also reducing harmful reactive oxygen species [3840]. NIr photobiomodulation could also upregulate protective factors such as nerve growth factor and vascular endothelial growth factor [41,42] and mesenchymal stem cells [43] that could target specific areas of degeneration.

The ability of NIr to reduce the expression of hyperphosphorylated tau, which in turn reduces oxidative stress [44], may be key to its neuroprotective effect. Oxidative stress and free radicals increase the severity of cerebrovascular lesions [45,46], mitochondrial dysfunction [4,47], oligomerisation of A? [5,48] and tauopathies and cell death [48,49] in AD. Considering the brain’s high consumption of oxygen and consequent susceptibility to oxidative stress, mitigating such stressors would probably have a pronounced protective effect [50].

Conclusions

Overall, our results in two transgenic mouse models with existing AD-related pathology suggest that low-energy NIr treatment can reduce characteristic pathology, oxidative stress and mitochondrial dysfunction in susceptible regions of the brain. These results, when taken together with those in other models of neurodegeneration, strengthen the notion that NIr is a viable neuroprotective treatment for a range of neurodegenerative conditions. We believe this growing body of work provides the impetus to begin trialling NIr treatment as a broad-based therapy for AD and other neurodegenerations.

Abbreviations

A?: Amyloid-beta; AD: Alzheimer’s disease; APP: Amyloid beta precursor protein gene; COX: Cytochrome c oxidase; 4-HNE: 4-hydroxynonenal; LED: Light-emitting diode; NFT: Neurofibrillary tangle; NIr: Near-infrared light; 8-OHDG: 8-hydroxy-2?-deoxyguanosine; PS1: Presenilin 1; WT: Wildtype.

Competing interests

The authors declare that they have no competing interests.

Authors’ contributions

SP undertook the bulk of the experimental work and analysis and wrote the manuscript. DMJ and JM were involved with the analysis of the data and the writing of the manuscript. CN was involved with genotyping and treating the animals. JS was involved in conceiving and designing the study and the writing of the manuscript. All authors read and approved the final manuscript.

Acknowledgments

The authors thank Tenix Corporation, Sir Zelman Cowen Universities Fund and Bluesand Foundation for funding. They are grateful to Prof. Lars Ittner for providing the breeding litter for K369I mice, and to Dr Louise Cole and the Bosch Advanced Microscopy facility for the help with MetaMorph. Sharon Spana was splendid for her technical help. DMJ is supported by a National Health and Medical Research Council of Australia (NHMRC) Early Career Fellowship.

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J Neuroinflammation.  2012 Sep 18;9(1):219. [Epub ahead of print]

Low-level laser therapy regulates microglial function through Src-mediated signaling pathways: implications for neurodegenerative diseases.

Song S, Zhou F, Chen WR, Xing D.

Abstract

ABSTRACT:

BACKGROUND:

Activated microglial cells are an important pathological component in brains of patients with neurodegenerative diseases. The purpose of this study was to investigate the effect of He-Ne (632.8 nm, 64.6 mW/cm2) low-level laser therapy (LLLT), a non-damaging physical therapy, on activated microglia, and the subsequent signaling events of LLLT-induced neuroprotective effects and phagocytic responses.

METHODS:

To model microglial activation, we treated the microglial BV2 cells with lipopolysaccharide (LPS). For the LLLT-induced neuroprotective study, neuronal cells with activated microglial cells in a Transwell[trade mark sign] cell-culture system were used. For the phagocytosis study, fluorescence-labeled microspheres were added into the treated microglial cells to confirm the role of LLLT.

RESULTS:

Our results showed that LLLT (20 J/cm2) could attenuate toll-like receptor (TLR)-mediated proinflammatory responses in microglia, characterized by down-regulation of proinflammatory cytokine expression and nitric oxide (NO) production. LLLT-triggered TLR signaling inhibition was achieved by activating tyrosine kinases Src and Syk, which led to MyD88 tyrosine phosphorylation, thus impairing MyD88-dependent proinflammatory signaling cascade. In addition, we found that Src activation could enhance Rac1 activity and F-actin accumulation that typify microglial phagocytic activity. We also found that Src/PI3K/Akt inhibitors prevented LLLT-stimulated Akt (Ser473 and Thr308) phosphorylation and blocked Rac1 activity and actin-based microglial phagocytosis, indicating the activation of Src/PI3K/Akt/Rac1 signaling pathway.

CONCLUSIONS:

The present study underlines the importance of Src in suppressing inflammation and enhancing microglial phagocytic function in activated microglia during LLLT stimulation. We have identified a new and important neuroprotective signaling pathway that consists of regulation of microglial phagocytosis and inflammation under LLLT treatment. Our research may provide a feasible therapeutic approach to control the progression of neurodegenerative diseases.

Postepy High Med Dosw (Online).  2011 Feb 17;65:73-92.

The role of biological sciences in understanding the genesis and a new therapeutic approach to Alzheimer’s disease.

Tegowska E, Wosinska A.

Zaklad Toksykologii Zwierz?a, Wydzial Biologii i Nauk o Ziemi, Uniwersytet Mikolaja Kopernika w Toruniu.

Abstract

The paper contrasts the historical view on causal factors in Alzheimer’s disease (AD) with the modern concept of the symptoms’ origin. Biological sciences dealing with cell structure and physiology enabled comprehension of the role of mitochondrial defects in the processes of formation of neurofibrillary tangles and ?-amyloid, which in turn gives hope for developing a new, more effective therapeutic strategy for AD. It has been established that although mitochondria constantly generate free radicals, from which they are protected by their own defensive systems, in some situations these systems become deregulated, which leads to free radical-based mitochondrial defects. This causes an energetic deficit in neurons and a further increase in the free radical pool. As a result, due to compensation processes, formation of tangles and/or acceleration of ?-amyloid production takes place. The nature of these processes is initially a protective one, due to their anti-oxidative action, but as the amount of the formations increases, their beneficial effect wanes. They become a storage place for substances enhancing free radical processes, which makes them toxic themselves. It is such an approach to the primary causal factor for AD which lies at the roots of the new view on AD therapy, suggesting the use of methylene blue-based drugs, laser or intranasally applied insulin. A necessary condition, however, for these methods’ effectiveness is definitely an earlier diagnosis of the disease. Although there are numerous diagnostic methods for AD, their low specificity and high price, often accompanied by a considerable level of patient discomfort, make them unsuitable for early, prodromal screening. In this matter a promising method may be provided using an olfactory test, which is an inexpensive and non-invasive method and thus suitable for screening, although as a test of low specificity, it should be combined with other methods. Introducing new methods of AD treatment does not mean abandoning the traditional ones, based on enhancing cholinergic transmission. They are valuable as long as the therapy starts when abundant brain inclusions disturb the transmissions.<br />

Photomed Laser Surg.  2010 Oct;28(5):663-7.

Long-term safety of single and multiple infrared transcranial laser treatments in Sprague-Dawley rats.

McCarthy TJ, De Taboada L, Hildebrandt PK, Ziemer EL, Richieri SP, Streeter J.

Source

PhotoThera, Inc., 5925 Priestly Drive, Suite 120, Carlsbad, California, USA. tmccarthy@photothera.com

Abstract

BACKGROUND AND OBJECTIVE:

Growing interest exists in the use of near-infrared laser therapies for the treatment of numerous neurologic conditions, including acute ischemic stroke, traumatic brain injury, Parkinson’s disease, and Alzheimer’s disease. In consideration of these trends, the objective of this study was to evaluate the long-term safety of transcranial laser therapy with continuous-wave (CW) near-infrared laser light (wavelength, 808?±?10?nm, 2-mm diameter) with a nominal radiant power of 70?mW; power density, 2,230?mW/cm(2), and energy density, 268?J/cm(2) at the scalp (10?mW/cm(2) and 1.2?J/cm(2) at the cerebral cortical surface) in healthy Sprague-Dawley rats.

MATERIALS AND METHODS:

In this study, 120 anesthetized rats received sequential transcranial laser treatments to the right and left parietal areas of the head on the same day (minimum of 5?min between irradiation of each side), on either Day 1 or on each of Days 1, 3, and 5. Sixty anesthetized rats served as sham controls. Rats were evaluated 1 year after treatment for abnormalities in clinical hematology and brain and pituitary gland histopathology.

RESULTS:

No toxicologically important differences were found in the clinical hematology results between sham-control and laser-treated rats for any hematologic parameters examined. All values fell within historic control reference ranges for aged Sprague-Dawley rats. Similarly, brain and pituitary gland histopathology showed no treatment-related abnormalities or induced neoplasia.

CONCLUSIONS:

Single and multiple applications of transcranial laser therapy with 808-nm CW laser light at a nominal power density of 10?mW/cm(2) at the surface of the cerebral cortex appears to be safe in Sprague-Dawley rats 1 year after treatment.

The Efficacy of 904 nm Laser Therapy for Alzheimer’s Diseases

Kazuyoshi Zenba, Vice president of Kanagawa Acupuncture Massage Association

Prof. Masayuki Inoue, Secretary of JLPLTPA

Preface

Although we had reported about the possible efficacy of low power laser therapy (LPLT) for Senile Dementia(S D) 3 times from 1993 at the annual meetings of Japan Society for Laser Medicine, there was no practically useful treatment found for Alzheimer’s disease(AD) and Parkinson disease and other Senile Dementia even after the start of elderly-care-insurance system in Japan. As we have continued above said laser therapy for SD at home care visit of elderly persons and felt very useful and effective, we would like to report about recent situation of laser therapy for AD patients.

Especially recently, the number of Alzheimer’s disease patients is increasing by the arrival of super-aged world in Japan. However the cause of this disease is not known and there is no effective treatment established at present. As to the mechanism of LPLT, its main mechanism is mostly elucidated by the progress in the field of Molecular biology and widely used for the removal of pain, decrease of swelling and treatment of wound. However its application for the treatment of Brain diseases is hardly practiced.

We have continued the treatment of Senile Dementia patients by LPL considering it as to be one of practical and effective treatment of this disease

LPLT is very useful for the medical treatment of the senile dementia patients at home for the expansion of ADL, pain relief, mitigation of inflammation, prevention of bed sore, the treatment of hemiplegia in a brain blood vessel obstacle and the braking of aggravation of Alzheimer’s disease without any fear of side effects by the irradiation of LPL to the head of patients.

It will be not to exaggerate to say LPLT can be one of main treatments of senior patients at home in near future.

(Object of study)

To study the practical usefulness of LPLT for the treatment of Alzheimer’s disease patients at home in terms of improvement of ADL and QOL and also for the reduction of burden of families of the care of patients.

(Method of treatment)

15 Alzheimer’s disease patients, 5 male and 10 female, received irradiation of LPL for 2 minutes at each points, 2-3 times a week for one year. Laser irradiation points were as follows. Acupuncture points established as effective based on long history of Oriental medicine .

(1) Acupuncture point to improve blood circulation (2) Acupuncture point for the treatment of stroke (3) Acupuncture point for adjustment of blood pressure (4) Acupuncture point for adjustment of balance of autonomous nerve.( the forehead, the right and left temple, occiput)

In addition, the method (based on papers in Russia and Armenia that intravenous LPL irradiation  improved the viscosity of blood) of irradiating LPL to the place which touches the pulse of an artery under collarbone was used as an additional medical treatment point.

(LPL instrument)

[Result]

Among evaluation items, cooperativeness and the lack of composure were observed as useful as an effect, the effect appeared half a year after and continued after one year and later on.

It was suggested that LPLT was useful for the improvement of orientation disturbance, normalization of clothing and the dress. Because, many families and the care workers talked us LPL was very helpful since the present condition could be maintained, without getting worse.

After the start of LPL treatment, It was reported that the coldness of hands and legs of patients vanished and joints and muscular stiffness were also mitigated. Therefore, the joint movable region was also secured comparatively.

Also in excretion care, it became very easy to carry out the care of patients.

It was able to say about all patients that their expression became quiet and came to show understanding to directions of a care worker. It is suggested by this that LPLT as one of practical treatment of patients at home by the improvement of care power at home.

(Discussion)

Since the senile-dementia-of-Alzheimer-type has a feature of advance of condition and it was said that condition became gradually critical, we tried this treatment expecting the maintenance of condition, and examination whether there was any delay effect. It is considered to have been suggested at least there was an effect of maintaining present condition in a certain field.

About the effect over the brain of laser irradiation, it was reported at the annual meeting of Japan Society for Laser Surgery and Medicine meeting in 1991 by Jun-Ichi Nishimura et al., of  Department of Physiology, Yokohama City University School of Medicine. The 780 nm wavelength and 1mW laser irradiation to the inner core of rats made the increase of cerebral blood flows at hippocampus by the amount of about 20% in average (control:15, laser:15). Although after 30 minute it was confirmed having maintained the increase of 10%.

In 1992 at the same medical conference, Takayuki Obata et. al., of the same University reported that laser irradiation of 780nm wavelength10mW to the head surface of rats activated cranial nerves activities (control:16, laser:15).

These reports suggested the possible usefulness of LPL treatment to Senile Dementia and other brain diseases patients. Unfortunately these findings did not much attention of medical world In Japan.

However, recently a possibility that ATP and cellmembrane potential of brain neuron could be controlled specifically by the irradiation of near infrared lasers (830nm wavelength) on the surface of heads of rats was reported by Oda-Mochizuki etc.al.?Ritsumeikan University, Synchrotron Light Life Science Center.

It was suggested by this research center that the condition of Epilepsy could be stabilized by Irradiating infrared laser from out side of heads of patients and decreasing the unusual excitement of cerebral neurons and in case of cerebral infarction, the aggravation of progress of Necrosis and Apotosis of cerebral neurons could be stopped by making stabilize the electric potential of cell membrane of cerebral neurons.

Development of future research in this field is expected as what supports scientifically the medical treatment of LPL and the result of condition improvements, such as Senile Dementia, brain blood vessel obstacles, hemiplegia and Parkinson patients.

Although the wavelength of LPL used for “Examination of the validity of LPL to Senile Dementia Patients” which we announced at the annual meetings of Japan Society for Laser Surgery and Medicine meeting over three years from 1993, was 780nm and out put was10mW, and 1mw.  The LPL used for this examination was of the wavelength of 904nm and the peak value of a pulse was 5W and the average output was 5mW. However, the same medical treatment effect was confirmed. Although it is thought that there was no wavelength dependability of laser to the efficacy over the Alzheimer’s diseases of LPL(780,830,904nm lasers are equally effective for pain removal and wound healing), how is it sure enough? A question remains.

By this examination, at least following effects were confirmed. Namely (1) the advance of condition of Alzheimer’s diseases has been blocked (2) and the expression of patients changed to smiling from disinterestedness, cooperativeness came out , an understanding came to be shown to a partner (3) We received comments from many families that the care of patients became much easier than before. It is considered that the head irradiation of near infrared laser light makes the cerebral blood flow improve, activates nerve activities and have applied brakes to the advance of the apotosis of brain cells as animal experiments are proving. Since the medical treatment efficacy is seldom acknowledged to middle degree class and a serious  patient, although it is hard to call it the fundamental cure for Alzheimer’s disease by the present method, if medical a treatment is started in early stage and continued, it may be possible to call it one of practical cures which can stop subsequent advance of disease.

Based on this experience, collecting newest information overseas, research results in the biology field, we will continue to study the possible LPL method for the dramatic cure of Alzheimer’s diseases by changing the wavelength of laser, the output and the irradiation method and also combination with other therapies

Breast Cancer

BMC Cancer. 2017 Dec 7;17(1):833. doi: 10.1186/s12885-017-3852-x.

Low level laser therapy (Photobiomodulation therapy) for breast cancer-related lymphedema: a systematic review.

Baxter GD1, Liu L2, Petrich S3, Gisselman AS2, Chapple C2, Anders JJ4, Tumilty S2.

Author information

1
Centre for Health, Activity and Rehabilitation Research, School of Physiotherapy, University of Otago, Dunedin, New Zealand. david.baxter@otago.ac.nz.
2
Centre for Health, Activity and Rehabilitation Research, School of Physiotherapy, University of Otago, Dunedin, New Zealand.
3
Department of Surgical Sciences, Southern District Health Board, Dunedin, New Zealand.
4
Department of Anatomy, Physiology and Genetics, Uniformed Services University of the Health Sciences, Maryland, MD, USA.

Abstract

BACKGROUND:

Breast cancer related lymphedema (BCRL) is a prevalent complication secondary to cancer treatments which significantly impacts the physical and psychological health of breast cancer survivors. Previous research shows increasing use of low level laser therapy (LLLT), now commonly referred to as photobiomodulation (PBM) therapy, for BCRL. This systematic review evaluated the effectiveness of LLLT (PBM) in the management of BCRL.

METHODS:

Clinical trials were searched in PubMed, AMED, Web of Science, and China National Knowledge Infrastructure up to November 2016. Two reviewers independently assessed the methodological quality and adequacy of LLLT (PBM) in these clinical trials. Primary outcome measures were limb circumference/volume, and secondary outcomes included pain intensity and range of motion. Because data were clinically heterogeneous, best evidence synthesis was performed.

RESULTS:

Eleven clinical trials were identified, of which seven randomized controlled trials (RCTs) were chosen for analysis. Overall, the methodological quality of included RCTs was high, whereas the reporting of treatment parameters was poor. Results indicated that there is strong evidence (three high quality trials) showing LLLT (PBM) was more effective than sham treatment for limb circumference/volume reduction at a short-term follow-up. There is moderate evidence (one high quality trial) indicating that LLLT (PBM) was more effective than sham laser for short-term pain relief, and limited evidence (one low quality trial) that LLLT (PBM) was more effective than no treatment for decreasing limb swelling at short-term follow-up.

CONCLUSIONS:

Based upon the current systematic review, LLLT (PBM) may be considered an effective treatment approach for women with BCRL. Due to the limited numbers of published trials available, there is a clear need for well-designed high-quality trials in this area. The optimal treatment parameters for clinical application have yet to be elucidated.

Lasers Med Sci. 2016 Dec;31(9):1775-1782. Epub 2016 Aug 12.

The effects of low-level laser irradiation on breast tumor in mice and the expression of Let-7a, miR-155, miR-21, miR125, and miR376b.

Khori V1, Alizadeh AM2, Gheisary Z3, Farsinejad S3, Najafi F4, Khalighfard S3, Ghafari F3, Hadji M3, Khodayari H3.

Author information

  • 1Ischemic Disorders Research Center, Golestan University of Medical Sciences, Gorgan, Iran.
  • 2Cancer Research Center, Tehran University of Medical Sciences, Tehran, Iran, Zip Code: 1419733141. aalizadeh@sina.tums.ac.ir.
  • 3Cancer Research Center, Tehran University of Medical Sciences, Tehran, Iran, Zip Code: 1419733141.
  • 4Medical Engineering, Faculty of Biomedical Engineering, Amir Kabir University, Tehran, Iran.

Abstract

Low-level laser therapy (LLLT) is a form of photon therapy which can be a non-invasive therapeutic procedure in cancer therapy using low-intensity light in the range of 450-800 nm. One of the main functional features of laser therapy is the photobiostimulation effects of low-level lasers on various biological systems including altering DNA synthesis and modifying gene expression, and stopping cellular proliferation. This study investigated the effects of LLLT on mice mammary tumor and the expression of Let-7a, miR155, miR21, miR125, and miR376b in the plasma and tumor samples. Sixteen mice were equally divided into four groups including control, and blue, green, and red lasers at wavelengths of 405, 532, and 632 nm, respectively. Weber Medical Applied Laser irradiation was carried out with a low power of 1-3 mW and a series of 10 treatments at three times a week after tumor establishment. Tumor volume was weekly measured by a digital vernier caliper, and qRT-PCR assays were performed to accomplish the study. Depending on the number of groups and the p value of the Kolmogorov-Smirnov test of normality, a t test, a one-way ANOVA, or a non-parametric test was used for data analyses, and p?<?0.05 was considered to be statistically significant. The average tumor volume was significantly less in the treated blue group than the control group on at days 21, 28, and 35 after cancerous cell injection. Our data also showed an increase of Let-7a and miR125a expression and a decrease of miR155, miR21, and miR376b expression after LLLT with the blue laser both the plasma and tumor samples compared to other groups. It seems that the non-invasive nature of laser bio-stimulation can make LLLT an attractive alternative therapeutic tool.

Lasers Med Sci. 2016 Aug 19. [Epub ahead of print]

The use of low-level light therapy in supportive care for patients with breast cancer: review of the literature.

Robijns J1,2, Censabella S3, Bulens P4,3, Maes A4,3, Mebis J5,4,3.

Author information

  • 1Faculty of Medicine & Life Sciences, Hasselt University, Martelarenlaan 42, 3500, Hasselt, Belgium. jolien.robijns@uhasselt.be.
  • 2Limburg Oncology Center, Stadsomvaart 11, 3500 Hasselt, Belgium. jolien.robijns@uhasselt.be.
  • 3Division of Medical Oncology, Jessa Hospital, Campus Virga Jesse, Stadsomvaart 11, 3500 Hasselt, Belgium.
  • 4Limburg Oncology Center, Stadsomvaart 11, 3500 Hasselt, Belgium.
  • 5Faculty of Medicine & Life Sciences, Hasselt University, Martelarenlaan 42, 3500, Hasselt, Belgium.

Abstract

Breast cancer is the most common cancer in women worldwide, with an incidence of 1.7 million in 2012. Breast cancer and its treatments can bring along serious side effects such as fatigue, skin toxicity, lymphedema, pain, nausea, etc. These can substantially affect the patients’ quality of life. Therefore, supportive care for breast cancer patients is an essential mainstay in the treatment. Low-level light therapy (LLLT) also named photobiomodulation therapy (PBMT) has proven its efficiency in general medicine for already more than 40 years. It is a noninvasive treatment option used to stimulate wound healing and reduce inflammation, edema, and pain. LLLT is used in different medical settings ranging from dermatology, physiotherapy, and neurology to dentistry. Since the last twenty years, LLLT is becoming a new treatment modality in supportive care for breast cancer. For this review, all existing literature concerning the use of LLLT for breast cancer was used to provide evidence in the following domains: oral mucositis (OM), radiodermatitis (RD), lymphedema, chemotherapy-induced peripheral neuropathy (CIPN), and osteonecrosis of the jaw (ONJ). The findings of this review suggest that LLLT is a promising option for the management of breast cancer treatment-related side effects. However, it still remains important to define appropriate treatment and irradiation parameters for each condition in order to ensure the effectiveness of LLLT.

Antioxid Redox Signal. 2015 Sep 28. [Epub ahead of print]

Phototherapy-induced antitumor immunity: long-term tumor supression effects via photoinactivation of respiratory chain oxidase-triggered superoxide anion burst.

Lu C1,2, Zhou F3, Wu S4,5,6, Liu L7, Xing D8.
Author information
1Guangzhou, China.
2MOE Key Laboratory of Laser Life Science & Institute of Laser Life Science, College of Biophotonics, South China Normal University , No. 55 Zhongshan Avenue West, Tianhe District,Guangzhou , guangzhou, China , 510631 ; lucx@scnu.edu.cn.
3MOE Key Laboratory of Laser Life Science & Institute of Laser Life Science, College of Biophotonics, South China Normal University , No. 55 Zhongshan Avenue West, Tianhe District,Guangzhou , guangzhou, China , 510631 ; zhouff@scnu.edu.cn.
4South China Normal UniversityGuang Zhou, China , 510631.
5China.
6MOE Key Laboratory of Laser Life Science & Institute of Laser Life Science, College of Biophotonics, South China Normal University , No. 55 Zhongshan Avenue West, Tianhe District,Guangzhou , guangzhou, China , 510631 ; wushn@scnu.edu.cn.
7MOE Key Laboratory of Laser Life Science & Institute of Laser Life Science, College of Biophotonics, South China Normal University , No. 55 Zhongshan Avenue West, Tianhe District,Guangzhou , guangzhou, China , 510631 ; liulei@scnu.edu.cn.
8South China Normal University , No. 55 Zhongshan west road, Tianhe district , guangzhou, China , 510631 ; xingda@scnu.edu.cn.

Abstract
AIMS:
Our previous studies have demonstrated that as a mitochondria-targeting cancer phototherapy, high-fluence low-power laser irradiation (HF-LPLI) results in oxidative damage that induces tumor cell apoptosis. In this study, we focused on the immunological effects of HF-LPLI phototherapy and explored its antitumor immune regulatory mechanism.
RESULTS:
We found not only that HF-LPLI treatment induced tumor cell apoptosis but also that HF-LPLI-treated apoptotic tumor cells activated macrophages. Due to mitochondrial superoxide anion burst after HF-LPLI treatment, tumor cells displayed a high level of phosphatidylserine oxidation, which mediated the recognition and uptake by macrophages with the subsequent secretion of cytokines and generation of cytotoxic T lymphocytes. In addition, in vivo results showed that HF-LPLI treatment caused leukocyte infiltration into the tumor and efficaciously inhibited tumor growth in an EMT6 tumor model. These phenomena were absent in the respiration-deficient EMT6 tumor model, implying that the HF-LPLI-elicited immunological effects were dependent on the mitochondrial superoxide anion burst.
INNOVATION:
Here, for the first time, we show that HF-LPLI mediates tumor-killing effects via targeting photoinactivation respiratory chain oxidase to trigger a superoxide anion burst, leading to a high level of oxidatively modified moieties, which contributes to the phenotypic changes in macrophages and mediates the antitumor immune response.
CONCLUSION:
Our results suggest that HF-LPLI may be an effective cancer treatment modality that both eradicates the treated primary tumors and induces an antitumor immune response via photoinactivation of respiratory chain oxidase to trigger superoxide anion burst.
Discov Med. 2015 Apr;19(105):293-301.

Advances in strategies and methodologies in cancer immunotherapy.

Lam SS1, Zhou F2, Hode T1, Nordquist RE1, Alleruzzo L1, Raker J1, Chen WR3.

Author information

  • 1Immunophotonics Inc., 4320 Forest Park Ave. #303, St. Louis, MO 63108, USA.
  • 2Biophotonics Research Laboratory, Center for Interdisciplinary Biomedical Education and Research, University of Central Oklahoma, Edmond, OK 73034, USA.
  • 3Biophotonics Research Laboratory, Center for Interdisciplinary Biomedical Education and Research, University of Central Oklahoma, Edmond, OK 73034, USA and Immunophotonics Inc., 4320 Forest Park Ave. #303, St. Louis, MO 63108, USA.

Abstract

Since the invention of Coley’s toxin by William Coley in early 1900s, the path for cancer immunotherapy has been a convoluted one. Although still not considered standard of care, with the FDA approval of trastuzumab, Provenge and ipilimumab, the medical and scientific community has started to embrace the possibility that immunotherapy could be a new hope for cancer patients with otherwise untreatable metastatic diseases. This review aims to summarize the development of some major strategies in cancer immunotherapy, from the earliest peptide vaccine and transfer of tumor specific antibodies/T cells to the more recent dendritic cell (DC) vaccines, whole cell tumor vaccines, and checkpoint blockade therapy. Discussion of some major milestones and obstacles in the shaping of the field and the future perspectives is included. Photoimmunotherapy is also reviewed as an example of emerging new therapies combining phototherapy and immunotherapy.

 J Biomed Opt.  2012 Oct;17(10):101516. doi: 10.1117/1.JBO.17.10.101516.

Low-level laser therapy on MCF-7 cells: a micro-Fourier transform infrared spectroscopy study.

Magrini TD, dos Santos NV, Milazzotto MP, Cerchiaro G, da Silva Martinho H.

Source

Centro de Ciências Naturais e Humanas, Universidade Federal do ABC, Rua Santa Adélia 166, Bangu, Santo André, SP 09210-170, Brazil.

Abstract

Low-level laser therapy (LLLT) is an emerging therapeutic approach for several clinical conditions. The clinical effects induced by LLLT presumably scale from photobiostimulation/photobioinhibition at the cellular level to the molecular level. The detailed mechanism underlying this effect remains unknown. This study quantifies some relevant aspects of LLLT related to molecular and cellular variations. Malignant breast cells (MCF-7) were exposed to spatially filtered light from a He-Ne laser (633 nm) with fluences of 5, 28.8, and 1000 mJ/cm². The cell viability was evaluated by optical microscopy using the Trypan Blue viability test. The micro-Fourier transform infrared technique was employed to obtain the vibrational spectra of each experimental group (control and irradiated) and identify the relevant biochemical alterations that occurred due to the process. It was observed that the red light influenced the RNA, phosphate, and serine/threonine/tyrosine bands. We found that light can influence cell metabolism depending on the laser fluence. For 5 mJ/cm², MCF-7 cells suffer bioinhibition with decreased metabolic rates. In contrast, for the 1 J/cm² laser fluence, cells present biostimulation accompanied by a metabolic rate elevation. Surprisingly, at the intermediate fluence, 28.8 mJ/cm², the metabolic rate is increased despite the absence of proliferative results. The data were interpreted within the retrograde signaling pathway mechanism activated with light irradiation.

Photomed Laser Surg.  2012 Sep;30(9):551-8. doi: 10.1089/pho.2011.3186. Epub 2012 Aug 1.

A preliminary study of the safety of red light phototherapy of tissues harboring cancer.

Myakishev-Rempel M, Stadler I, Brondon P, Axe DR, Friedman M, Nardia FB, Lanzafame R.

Source

Department of Dermatology, University of Rochester, Rochester, New York, USA. max.rempel@gmail.com

Abstract

OBJECTIVE:

Red light phototherapy is known to stimulate cell proliferation in wound healing. This study investigated whether low-level light therapy (LLLT) would promote tumor growth when pre-existing malignancy is present.

BACKGROUND DATA:

LLLT has been increasingly used for numerous conditions, but its use in cancer patients, including the treatment of lymphedema or various unrelated comorbidities, has been withheld by practitioners because of the fear that LLLT might result in initiation or promotion of metastatic lesions or new primary tumors. There has been little scientific study of oncologic outcomes after use of LLLT in cancer patients.

METHODS:

A standard SKH mouse nonmelanoma UV-induced skin cancer model was used after visible squamous cell carcinomas were present, to study the effects of LLLT on tumor growth. The red light group (n=8) received automated full body 670 nm LLLT delivered twice a day at 5 J/cm(2) using an LED source. The control group (n=8) was handled similarly, but did not receive LLLT. Measurements on 330 tumors were conducted for 37 consecutive days, while the animals received daily LLLT.

RESULTS:

Daily tumor measurements demonstrated no measurable effect of LLLT on tumor growth.

CONCLUSIONS:

This experiment suggests that LLLT at these parameters may be safe even when malignant lesions are present. Further studies on the effects of photoirradiation on neoplasms are warranted.

J Biomed Opt. 2012 Oct 25;17(10):101516-1. doi: 10.1117/1.JBO.17.10.101516.

Low-level laser therapy on MCF-7 cells: a micro-Fourier transform infrared spectroscopy study.

Magrini TD, Dos Santos NV, Milazzotto MP, Cerchiaro G, da Silva Martinho H.

Abstract

ABSTRACT. Low-level laser therapy (LLLT) is an emerging therapeutic approach for several clinical conditions. The clinical effects induced by LLLT presumably scale from photobiostimulation/photobioinhibition at the cellular level to the molecular level. The detailed mechanism underlying this effect remains unknown. This study quantifies some relevant aspects of LLLT related to molecular and cellular variations. Malignant breast cells (MCF-7) were exposed to spatially filtered light from a He-Ne laser (633 nm) with fluences of 5, 28.8, and 1000 mJ/cm2. The cell viability was evaluated by optical microscopy using the Trypan Blue viability test. The micro-Fourier transform infrared technique was employed to obtain the vibrational spectra of each experimental group (control and irradiated) and identify the relevant biochemical alterations that occurred due to the process. It was observed that the red light influenced the RNA, phosphate, and serine/threonine/tyrosine bands. We found that light can influence cell metabolism depending on the laser fluence. For 5 mJ/cm2, MCF-7 cells suffer bioinhibition with decreased metabolic rates. In contrast, for the 1 J/cm2 laser fluence, cells present biostimulation accompanied by a metabolic rate elevation. Surprisingly, at the intermediate fluence, 28.8 mJ/cm2, the metabolic rate is increased despite the absence of proliferative results. The data were interpreted within the retrograde signaling pathway mechanism activated with light irradiation.

Photomed Laser Surg.  2012 Aug 1. [Epub ahead of print]

A Preliminary Study of the Safety of Red Light Phototherapy of Tissues Harboring Cancer.

Myakishev-Rempel M, Stadler I, Brondon P, Axe DR, Friedman M, Nardia FB, Lanzafame R.

Source

1 Department of Dermatology, University of Rochester , Rochester, New York.

Abstract

Abstract Objective: Red light phototherapy is known to stimulate cell proliferation in wound healing. This study investigated whether low-level light therapy (LLLT) would promote tumor growth when pre-existing malignancy is present.

Background data: LLLT has been increasingly used for numerous conditions, but its use in cancer patients, including the treatment of lymphedema or various unrelated comorbidities, has been withheld by practitioners because of the fear that LLLT might result in initiation or promotion of metastatic lesions or new primary tumors. There has been little scientific study of oncologic outcomes after use of LLLT in cancer patients.

Methods: A standard SKH mouse nonmelanoma UV-induced skin cancer model was used after visible squamous cell carcinomas were present, to study the effects of LLLT on tumor growth. The red light group (n=8) received automated full body 670 nm LLLT delivered twice a day at 5 J/cm(2) using an LED source. The control group (n=8) was handled similarly, but did not receive LLLT.

Measurements on 330 tumors were conducted for 37 consecutive days, while the animals received daily LLLT. Results: Daily tumor measurements demonstrated no measurable effect of LLLT on tumor growth.

Conclusions: This experiment suggests that LLLT at these parameters may be safe even when malignant lesions are present. Further studies on the effects of photoirradiation on neoplasms are warranted.

Vopr Kurortol Fizioter Lech Fiz Kult.  2012 Jul-Aug;(4):23-32.

The efficacy of polychromatic visible and infrared radiation used for the postoperative immunological rehabilitation of patients with breast cancer.

[Article in Russian]
[No authors listed]

Abstract

The immunological rehabilitation of the patients with oncological problems after the completion of standard anti-tumour therapy remains a topical problem in modern medicine. The up-to-date phototherapeutic methods find the increasingly wider application for the treatment of such patients including the use of monochromatic visible (VIS) and near infrared (nIR) radiation emitted from lasers and photodiodes. The objective of the present study was to substantiate the expediency of postoperative immune rehabilitation of the patients with breast cancer (BC) by means of irradiation of the body surface with polychromatic visible (pVIS) in combination with polychromatic infrared (pIR) light similar to the natural solar radiation without its minor UV component. The study included 19 patients with stage I–II BC at the mean age of 54.0 +/- 4.28 years having the infiltrative-ductal form of the tumour who had undergone mastectomy. These patients were randomly allocated to two groups, one given the standard course of postoperative rehabilitation (control), the other (study group) additionally treated with pVIS + pIR radiation applied to the lumbar-sacral region from days 1 to 7 after surgery. A Bioptron-2 phototherapeutic device, Switzerland, was used for the purpose (480-3400 nm, 40 mW/cm2, 12 J/cm2, with the light spot diameter of 15 cm). The modern standard immunological methods were employed. It was found that mastectomy induced changes of many characteristics of cellular and humoral immunity; many of them in different patients were oppositely directed. These changes were apparent within the first 7 days postoperatively. The course of phototherapy (PT) was shown to prevent the postoperative decrease in the counts of monocytes and natural killer (NK) cells, the total amount of CD3+ -T-lymphocytes (LPC), CD4+ -T-helpers, activated T-lymphocytes (CD3+ HLA-DR+ cells) and IgA levels as well as intracellular digestion rate of neutrophil-phagocyted bacteria. Moreover PT promoted faster normalization of postoperative leukocytosis and activation of cytotoxic CD8+ -T-LPC, reduced the elevated concentration of immune complexes in blood. Among the six tested cytokines, viz. IL-1beta, TNF-alpha, IL-6, IL-10, IFN-alpha, and IFN-gamma, only the latter two underwent significant elevation of their blood concentrations (IL-6 within 1 day) and IFN-gamma (within 7 days after mastectomy). The course of PT resulted in the decrease of their levels to the initial values. The follow-up of the treated patients during 4 years revealed neither recurrence of the disease nor the appearance of metastases.

Photomed Laser Surg. 2010 Feb;28(1):115-23.

The effect of laser irradiation on proliferation of human breast carcinoma, melanoma, and immortalized mammary epithelial cells.

Powell K, Low P, McDonnell PA, Laakso EL, Ralph SJ.

School of Medical Science, Griffith University, Gold Coast, Queensland, Australia.

Abstract

OBJECTIVE: This study compared the effects of different doses (J/cm(2)) of laser phototherapy at wavelengths of either 780, 830, or 904 nm on human breast carcinoma, melanoma, and immortalized human mammary epithelial cell lines in vitro. In addition, we examined whether laser irradiation would malignantly transform the murine fibroblast NIH3T3 cell line.

BACKGROUND: Laser phototherapy is used in the clinical treatment of breast cancer-related lymphoedema, despite limited safety information. This study contributes to systematically developing guidelines for the safe use of laser in breast cancer-related lymphoedema. METHODS: Human breast adenocarcinoma (MCF-7), human breast ductal carcinoma with melanomic genotypic traits (MDA-MB-435S), and immortalized human mammary epithelial (SVCT and Bre80hTERT) cell lines were irradiated with a single exposure of laser. MCF-7 cells were further irradiated with two and three exposures of each laser wavelength. Cell proliferation was assessed 24 h after irradiation.

RESULTS: Although certain doses of laser increased MCF-7 cell proliferation, multiple exposures had either no effect or showed negative dose response relationships. No sign of malignant transformation of cells by laser phototherapy was detected under the conditions applied here.

CONCLUSION: Before a definitive conclusion can be made regarding the safety of laser for breast cancer-related lymphoedema, further in vivo research is required.

Vopr Kurortol Fizioter Lech Fiz Kult. 2009 Nov-Dec;(6):49-52.

Application of low-power visible and near infrared radiation in clinical oncology.

[Article in Russian]

Zimin AA, Zhevago NA, Buniakova AI, Samoilova KA.

Although low-power visible (VIS) and near infrared (nIR) radiation emitted from lasers, photodiodes, and other sources does not cause neoplastic transformation of the tissue, these phototherapeutic techniques are looked at with a great deal of caution for fear of their stimulatory effect on tumour growth. This apprehension arises in the first place from the reports on the possibility that the proliferative activity of tumour cells may increase after their in vitro exposure to light. Much less is known that these phototherapeutic modalities have been successfully used for the prevention and management of complications developing after surgery, chemo- and radiotherapy. The objective of the present review is to summarize the results of applications of low-power visible and near infrared radiation for the treatment of patients with oncological diseases during the last 20-25 years. It should be emphasized that 2-4 year-long follow-up observations have not revealed any increase in the frequency of tumour recurrence and metastasis.

Photomed Laser Surg. 2009 Oct;27(5):763-9.

Managing postmastectomy lymphedema with low-level laser therapy.

Lau RW, Cheing GL.

Department of Rehabilitation Sciences, The Hong Kong Polytechnic University, Hong Kong SAR, China.

OBJECTIVE: We aimed to investigate the effects of low-level laser therapy (LLLT) in managing postmastectomy lymphedema. BACKGROUND DATA: Postmastectomy lymphedema (PML) is a common complication of breast cancer treatment that causes various symptoms, functional impairment, or even psychosocial morbidity. A prospective, single-blinded, controlled clinical trial was conducted to examine the effectiveness of LLLT on managing PML.

METHODS: Twenty-one women suffering from unilateral PML were randomly allocated to receive either 12 sessions of LLLT in 4 wk (the laser group) or no laser irradiation (the control group). Volumetry and tonometry were used to monitor arm volume and tissue resistance; the Disabilities of Arm, Shoulder, and Hand (DASH) questionnaire was used for measuring subjective symptoms. Outcome measures were assessed before and after the treatment period and at the 4 wk follow-up.

RESULTS: Reduction in arm volume and increase in tissue softening was found in the laser group only. At the follow-up session, significant between-group differences (all p < 0.05) were found in arm volume and tissue resistance at the anterior torso and forearm region. The laser group had a 16% reduction in the arm volume at the end of the treatment period, that dropped to 28% in the follow-up. Moreover, the laser group demonstrated a cumulative increase from 15% to 33% in the tonometry readings over the forearm and anterior torso. The DASH score of the laser group showed progressive improvement over time.

CONCLUSION: LLLT was effective in the management of PML, and the effects were maintained to the 4 wk follow-up.

Clin Rehabil. 2009 Feb;23(2):117-24

Efficacy of pneumatic compression and low-level laser therapy in the treatment of postmastectomy lymphoedema: a randomized controlled trial.

Kozanoglu E, Basaran S, Paydas S, Sarpel T.

Department of Physical Medicine and Rehabilitation, Faculty of Medicine, Cukurova University, Adana, Turkey.

Objective: To compare the long-term efficacy of pneumatic compression and low-level laser therapies in the management of postmastectomy lymphoedema.

Design: Randomized controlled trial.Setting: Department of Physical Medicine and Rehabilitation of Cukurova University, Turkey.

Subjects: Forty-seven patients with postmastectomy lymphoedema were enrolled in the study.Interventions: Patients were randomly allocated to pneumatic compression (group I, n=24) and low-level laser (group II, n=23) groups. Group I received 2 hours of compression therapy and group II received 20 minutes of laser therapy for four weeks. All patients were advised to perform daily limb exercises.Main measures: Demographic features, difference between sum of the circumferences of affected and unaffected limbs (triangle upC), pain with visual analogue scale and grip strength were recorded.

Results: Mean age of the patients was 48.3 (10.4) years. triangle upC decreased significantly at one, three and six months within both groups, and the decrease was still significant at month 12 only in group II (P = 0.004). Improvement of group II was greater than that of group I post treatment (P = 0.04) and at month 12 after 12 months (P = 0.02). Pain was significantly reduced in group I only at posttreatment evaluation, whereas in group II it was significant post treatment and at follow-up visits. No significant difference was detected in pain scores between the two groups. Grip strength was improved in both groups, but the differences between groups were not significant.

Conclusions: Patients in both groups improved after the interventions. Group II had better long-term results than group I. Low-level laser might be a useful modality in the treatment of postmastectomy lymphoedema.

Photomed Laser Surg. 2008 Aug;26(4):393-400.

Low-level laser therapy in the prevention and treatment of chemotherapy-induced oral mucositis in young patients.

Abramoff MM, Lopes NN, Lopes LA, Dib LL, Guilherme A, Caran EM, Barreto AD, Lee ML, Petrilli AS.

Private practice, São Paulo, Brazil.

Abstract Objective: A pilot clinical study was conducted to evaluate the efficacy and feasibility of low-level laser therapy (LLLT) in the prevention and treatment of chemotherapy (CT)-induced oral mucositis (OM) in young patients. Background Data: Besides compromising the patient’s nutrition and well-being, oral mucositis represents a portal of entry into the body for microorganisms present in the mouth, which may lead to sepsis if there is hematological involvement. Oncologic treatment tolerance decreases and systemic complications may arise that interfere with the success of cancer treatment. LLLT appears to be an interesting alternative to other approaches to treating OM, due to its trophic, anti-inflammatory, and analgesic properties. Materials and Methods: Patients undergoing chemotherapy (22 cycles) without mucositis were randomized into a group receiving prophylactic laser-irradiation (group 1), and a group receiving placebo light treatment (group 2). Patients who had already presented with mucositis were placed in a group receiving irradiation for therapeutic purposes (group 3, with 10 cycles of CT). Serum granulocyte levels were taken and compared to the progression of mucositis. Results: In group 1, most patients (73%) presented with mucositis of grade 0 (p = 0.03 when compared with the placebo group), and 18% presented with grade 1. In group 2, 27% had no OM and did not require therapy. In group 3, the patients had marked pain relief (as assessed by a visual analogue scale), and a decrease in the severity of OM, even when they had severe granulocytopenia. Conclusion: The ease of use of LLLT, high patient acceptance, and the positive results achieved, make this therapy feasible for the prevention and treatment of OM in young patients.

Ann Oncol. 2007 Apr;18(4):639-46. Epub 2006 Oct 3

A systematic review of common conservative therapies for arm lymphoedema secondary to breast cancer treatment.

Moseley AL, Carati CJ, Piller NB.

School of Nursing & Midwifery, University of South Australia, Adelaide, Australia. amanda.moseley@yahoo.com.au

Secondary arm lymphoedema is a chronic and distressing condition which affects a significant number of women who undergo breast cancer treatment. A number of health professional and patient instigated conservative therapies have been developed to help with this condition, but their comparative benefits are not clearly known. This systematic review undertook a broad investigation of commonly instigated conservative therapies for secondary arm lymphoedema including; complex physical therapy, manual lymphatic drainage, pneumatic pumps, oral pharmaceuticals, low level laser therapy, compression bandaging and garments, limb exercises and limb elevation. It was found that the more intensive and health professional based therapies, such as complex physical therapy, manual lymphatic drainage, pneumatic pump and laser therapy generally yielded the greater volume reductions, whilst self instigated therapies such as compression garment wear, exercises and limb elevation yielded smaller reductions. All conservative therapies produced improvements in subjective arm symptoms and quality of life issues, where these were measured. Despite the identified benefits, there is still the need for large scale, high level clinical trials in this area.

Lasers Med Sci. 2006 Jul;21(2):90-4. Epub 2006 May 4.

Low-level laser therapy in management of postmastectomy lymphedema.

Kaviani A, Fateh M, Yousefi Nooraie R, Alinagi-zadeh MR, Ataie-Fashtami L.

Tehran University of Medical Sciences and Iranian Center for Medical Laser Research, Tehran, Iran. akaviani@sina.tims.ac.ir

The aim of this paper was to study the effects of low-level laser therapy (LLLT) in the treatment of postmastectomy lymphedema. Eleven women with unilateral postmastectomy lymphedema were enrolled in a double-blind controlled trial. Patients were randomly assigned to laser and sham groups and received laser or placebo irradiation (Ga-As laser device with a wavelength of 890 nm and fluence of 1.5 J/cm2) over the arm and axillary areas. Changes in patients’ limb circumference, pain score, range of motion, heaviness of the affected limb, and desire to continue the treatment were measured before the treatment and at follow-up sessions (weeks 3, 9, 12, 18, and 22) and were compared to pretreatment values. Results showed that of the 11 enrolled patients, eight completed the treatment sessions. Reduction in limb circumference was detected in both groups, although it was more pronounced in the laser group up to the end of 22nd week. Desire to continue treatment at each session and baseline score in the laser group was greater than in the sham group in all sessions. Pain reduction in the laser group was more than in the sham group except for the weeks 3 and 9. No substantial differences were seen in other two parameters between the two treatment groups. In conclusion, despite our encouraging results, further studies of the effects of LLLT in management of postmastectomy lymphedema should be undertaken to determine the optimal physiological and physical parameters to obtain the most effective clinical response.

J Photochem Photobiol B. 2000 Dec;59(1-3):1-8.

Magnetic resonance imaging (MRI) controlled outcome of side effects caused by ionizing radiation, treated with 780 nm-diode laser –preliminary results.

Schaffer M, Bonel H, Sroka R, Schaffer PM, Busch M, Sittek H, Reiser M, Duhmke E.
Department of Radiation Therapy, University of Munich, Germany.

sroka@life.med.uni-muenchen.de

BACKGROUND and OBJECTIVE: Ionizing radiation therapy by way of various beams such as electron, photon and neutron is an established method in tumor treatment. The side effects caused by this treatment such as ulcer, painful mastitis and delay of wound healing are well known, too. Biomodulation by low level laser therapy (LLLT) has become popular as a therapeutic modality for the acceleration of wound healing and the treatment of inflammation. Evidence for this kind of application, however, is not fully understood yet. This study intends to demonstrate the response of biomodulative laser treatment on the side effects caused by ionizing radiation by means of magnetic resonance imaging (MRI). STUDY

DESIGN/PATIENTS and METHODS: Six female patients suffering from painful mastitis after breast ionizing irradiation and one man suffering from radiogenic ulcer were treated with lambda=780 nm diode laser irradiation at a fluence rate of 5 J/cm2. LLLT was performed for a period of 4-6 weeks (mean sessions: 25 per patient, range 19-35). The tissue response was determined by means of MRI after laser treatment in comparison to MRI prior to the beginning of the LLLT.

RESULTS: All patients showed complete clinical remission. The time-dependent contrast enhancement curve obtained by the evaluation of MR images demonstrated a significant decrease of enhancement features typical for inflammation in the affected area.

CONCLUSION: Biomodulation by LLLT seems to be a promising treatment modality for side effects induced by ionizing radiation.