Nitric Oxide

Arch Biochem Biophys. 2018 May 9. pii: S0003-9861(18)30073-0. doi: 10.1016/ [Epub ahead of print]

Wavelength-dependence of vasodilation and NO release from S-nitrosothiols and dinitrosyl iron complexes by far red/near infrared light.

Keszler A1, Lindemer B2, Hogg N3, Weihrauch D4, Lohr NL5.

Author information

Department of Medicine- Division of Cardiovascular Medicine, United States. Electronic address:
Department of Medicine- Division of Cardiovascular Medicine, United States. Electronic address:
Department of Biophysics, United States; Department of Redox Biology Program, United States. Electronic address:
Department of Anesthesiology, United States. Electronic address:
Department of Medicine- Division of Cardiovascular Medicine, United States; Department of Cardiovascular Center, Medical College of Wisconsin, United States; Clement J Zablocki VA Medical Center, United States. Electronic address:


Far red/near infrared (R/NIR) energy is a novel therapy, but its mechanism of action is poorly characterized. Cytochrome c oxidase (Cco) of the mitochondrial electron transport chain is considered the primary photoacceptor for R/NIR to photolyze a putative heme nitrosyl in Cco to liberate free nitric oxide (NO). We previously observed R/NIR light directly liberates NO from nitrosylated hemoglobin and myoglobin, and recently suggested S-nitrosothiols (RSNO) and dinitrosyl iron complexes (DNIC) may be primary sources of R/NIR-mediated NO. Here we indicate R/NIR light exposure induces wavelength dependent dilation of murine facial artery, with longer wavelengths (740, and 830?nm) exhibiting reduced potency when compared to 670?nm. R/NIR also stimulated NO release from pure solutions of low molecular weight RSNO (GSNO and SNAP) and glutathione dinitrosyl iron complex (GSH-DNIC) in a power- and wavelength-dependent manner, with the greatest effect at 670?nm. NO release from SNAP using 670 was nearly ten-fold more than GSNO or GSH-DNIC, with no substantial difference in NO production at 740?nm and 830?nm. Thermal effects of irradiation on vasodilation or NO release from S-nitrosothiols and DNIC was minimal. Our results suggest 670?nm is the optimal wavelength for R/NIR treatment of certain vascular-related diseases.


DNIC; Far red/near infrared light; Low level light therapy; Nitric oxide; Photobiomodulation; S-Nitrosothiol; Vasodilation

Free Radic Biol Med. 2017 Dec;113:157-164. doi: 10.1016/j.freeradbiomed.2017.09.012. Epub 2017 Sep 19.

Red/near infrared light stimulates release of an endothelium dependent vasodilator and rescues vascular dysfunction in a diabetes model.

Keszler A1, Lindemer B2, Weihrauch D3, Jones D4, Hogg N5, Lohr NL6.

Author information

Departments of Medicine, Division of Cardiovascular Medicine, Medical College of Wisconsin, United States; Departments of Anesthesiology, Medical College of Wisconsin, United States.
Departments of Medicine, Division of Cardiovascular Medicine, Medical College of Wisconsin, United States.
Departments of Anesthesiology, Medical College of Wisconsin, United States.
Departments of Pediatric Surgery, Medical College of Wisconsin, United States.
Departments of Biophysics, Medical College of Wisconsin, United States.
Departments of Medicine, Division of Cardiovascular Medicine, Medical College of Wisconsin, United States; Departments of Cardiovascular Center, Medical College of Wisconsin, United States; Departments of Clement J Zablocki VA Medical Center, United States. Electronic address:


Peripheral artery disease (PAD) is a morbid condition whereby ischemic peripheral muscle causes pain and tissue breakdown. Interestingly, PAD risk factors, e.g. diabetes mellitus, cause endothelial dysfunction secondary to decreased nitric oxide (NO) levels, which could explain treatment failures. Previously, we demonstrated 670nm light (R/NIR) increased NO from nitrosyl-heme stores, therefore we hypothesized R/NIR can stimulate vasodilation in healthy and diabetic blood vessels. Vasodilation was tested by ex vivo pressure myography in wild type C57Bl/6, endothelial nitric oxide synthase (eNOS) knockout, and db/db mice (10mW/cm2 for 5min with 10min dark period). NOS inhibition with N-Nitroarginine methyl ester (L-NAME) or the NO scavenger Carboxy-PTIO (c-PTIO) tested the specificity of NO production. 4,5-Diaminofluorescein diacetate (DAF-2) measured NO in human dermal microvascular endothelial cells (HMVEC-d). R/NIR significantly increased vasodilation in wild type and NOS inhibited groups, however R/NIR dilation was totally abolished with c-PTIO and blood vessel denudation. Interestingly, the bath solution from intact R/NIR stimulated vessels could dilate light naïve vessels in a NO dependent manner. Characterization of the bath identified a NO generating substance suggestive of S-nitrosothiols or non heme iron nitrosyl complexes. Consistent with the finding of an endothelial source of NO, intracellular NO increased with R/NIR in HMVEC-d treated with and without L-NAME (1mM), yet c-PTIO (100µm) reduced NO production. R/NIR significantly dilated db/db blood vessels. In conclusion, R/NIR stimulates vasodilation by release of NO bound substances from the endothelium. In a diabetes model of endothelial dysfunction, R/NIR restores vasodilation, which lends the potential for new treatments for diabetic vascular disease.


Diabetes mellitus; Electromagnetic energy; Endothelial dysfunction; Microcirculation; Nitric oxide; Photobiomodulation

Lasers Med Sci. 2014 May 24. [Epub ahead of print]

Effects of low-level laser therapy on mitochondrial respiration and nitrosyl complex content.

Buravlev EA1, Zhidkova TV, Vladimirov YA, Osipov AN.

Author information

  • 1Department of General and Medical Biophysics, NI Pirogov Russian National Research Medical University, 1 Ostrovityanov str., 117997, Moscow, Russia,


Among the photochemical reactions responsible for therapeutic effects of low-power laser radiation, the photolysis of nitrosyl iron complexes of iron-containing proteins is of primary importance. The purpose of the present study was to compare the effects of blue laser radiation on the respiration rate and photolysis of nitrosyl complexes of iron-sulfur clusters (NO-FeS) in mitochondria, subjected to NO as well as the possibility of NO transfer from NO-FeS to hemoglobin. It was shown that mitochondrial respiration in State 3 (V3) and State 4 (V4), according to Chance, dramatically decreased in the presence of 3 mM NO, but laser radiation (??=?442 nm, 30 J/cm2) restored the respiration rates virtually to the initial level. At the same time, electron paramagnetic resonance (EPR) spectra showed that laser irradiation decomposed nitrosyl complexes produced by the addition of NO to mitochondria. EPR signal of nitrosyl complexes of FeS-clusters, formed in the presence of 3 mM NO, was maximal in hypoxic mitochondria, and disappeared in a dose-dependent manner, almost completely at the irradiation dose 120 J/cm2. EPR measurements showed that the addition of lysed erythrocytes to mitochondria decreased the amount of nitrosyl complexes in iron-sulfur clusters and produced the accumulation of NO-hemoglobin. On the other hand, the addition of lysed erythrocytes to mitochondria, preincubated with nitric oxide, restored mitochondrial respiration rates V3 and V4 to initial levels. We may conclude that there are two possible ways to destroy FeS nitrosyl complexes in mitochondria and recover mitochondrial respiration inhibited by NO: laser irradiation and ample supply of the compounds with high affinity to nitric oxide, including hemoglobin.

J Mol Cell Cardiol 2013 May 20

Far red/near infrared light treatment promotes femoral artery collateralization in the ischemic hindlimb.

Lohr NL, Ninomiya JT, Warltier DC, Weihrauch D

Department of Medicine, Division of Cardiovascular Medicine, Medical College of Wisconsin, USA; Department of Cardiovascular Center, Medical College of Wisconsin, USA; Clement J Zablocki VA Medical Center, USA. Electronic address:

Nitric oxide (NO) is a crucial mediator of hindlimb collateralization and angiogenesis. Within tissues there are nitrosyl-heme proteins which have the potential to generate NO under conditions of hypoxia or low pH. Low level irradiation of blood and muscle with light in the far red/near infrared spectrum (670nm, R/NIR) facilitates NO release. Therefore, we assessed the impact of red light exposure on the stimulation of femoral artery collateralization. Rabbits and mice underwent unilateral resection of the femoral artery and chronic R/NIR treatment. The direct NO scavenger carboxy-PTIO and the nitric oxide synthase (NOS) inhibitor L-NAME were also administered in the presence of R/NIR. DAF fluorescence assessed R/NIR changes in NO levels within endothelial cells. In vitro measures of R/NIR induced angiogenesis were assessed by endothelial cell proliferation and migration. R/NIR significantly increased collateral vessel number which could not be attenuated with L-NAME. R/NIR induced collateralization was abolished with c-PTIO. In vitro, NO production increased in endothelial cells with R/NIR exposure, and this finding was independent of NOS inhibition. Similarly R/NIR induced proliferation and tube formation in a NO dependent manner. Finally, nitrite supplementation accelerated R/NIR collateralization in wild type C57Bl/6 mice. In an eNOS deficient transgenic mouse model, R/NIR restores collateral development. In conclusion, R/NIR increases NO levels independent of NOS activity, and leads to the observed enhancement of hindlimb collateralization.

Cent Nerv Syst Agents Med Chem.  2012 Apr 25. [Epub ahead of print]

Nitrergic Response to Clostridium perfringens Infection in the Rat Brain Regions. Effect of Red Light Irradiation.

Movsesyan HA, Alchujyan NK, Movsesyan NH, Guevorkian AG, Hairapetyan HL, Barsegyan KA, Kevorkian GA.


Sevak St., Building 5/1. H. Buniatian Institute of Biochemistry, National Academy of Sciences, Republic of Armenia, Yerevan.


A single intraperitoneal injection of a gram-positive pathogen Clostridium perfringens (Cp) causes a remarkable down-regulation the constitutive nitric oxide synthase (cNOS) with a simultaneous increase in the activity of inducible NOS (iNOS) and the level of reactive nitrogen species in the rat brain major regions (cortex, striatum, hippocampus and hypothalamus) at 48 h post-administration of Cp. Treatment by both a semiconductor laser (SCL) and/or a light-emitting diode (LED) with same wavelength, energy density and time exposure (continuous wave, ?=654 nm, fluence=1.27 J/cm2, time exposure=600 s) could modulate brain nitrergic response following Cp-infection. Besides, unlike the LED, the SCL-irradiation prevents the cNOS inhibition in all the studied brain regions and might be useful in restoring its function in neurotransmission and cerebral blood flow, along with providing a protective effect against nitrosative stress-induced iNOS-mediated injury in the brain regions.

Lasers Surg Med. 2010 Aug;42(6):577-83.

Evaluation of inflammatory biomarkers associated with oxidative stress and histological assessment of low-level laser therapy in experimental myopathy.

Servetto N, Cremonezzi D, Simes JC, Moya M, Soriano F, Palma JA, Campana VR.

Cátedra de Física Biomédica, Facultad de Ciencias Médicas, Universidad Nacional de Córdoba, Santa Rosa 1085, (5000) Córdoba, Argentina.


The objective of the present work was to study the effect of helium-neon (He-Ne) and gallium-arsenide (Ga-As) laser upon inflammatory biomarkers associated with oxidative stress: fibrinogen, nitric oxide (NO), L-citrulline, and superoxide dismutase (SOD). These were evaluated through histological assessment, in rats with experimental myopathy.

MATERIALS AND METHODS: The groups studied were: (A) control, (B) injured, (C) injured and treated with He-Ne laser, (D) injured and treated with Ga-As laser, (E) irradiated with He-Ne; and (F) irradiated with Ga-As laser. Myopathy was induced by injecting 0.05 mg/rat/day of adrenaline in the left posterior limb muscle at the same point on 5 consecutive days, in groups B, C, and D. Low-level laser therapy (LLLT) was applied with 9.5 J/cm(2) daily for 7 consecutive days with each laser. The determination of the biomarkers was made by spectrophotometry. The muscles (5/8, single blinded) were stained with Gomori Trichrome and examined by optic microscopy. The quantitative variables were statistically analyzed by the Fisher’s test and categorical data by the Axionvision 4.8 program. Pearson’s chi-squared test was applied, setting significant difference at P < 0.05 for all cases.

RESULTS: In group B, the biomarkers were significantly increased compared to the other groups (P < 0.001), except for NO which in group B decreased significantly (P < 0.001). In group B, there was a higher inflammatory infiltration level (80.67%) in relation to destroyed fibers.

CONCLUSIONS: LLLT caused significant changes in inflammatory biomarkers and oxidative stress: decreased levels of fibrinogen, L-citrulline and SOD as opposed to the increase of NO in rats with experimental myopathies and significant muscle recovery.

Photomed Laser Surg. 2009 Feb;27(1):79-84.

Inflammatory and oxidative stress markers in experimental crystalopathy: their modification by photostimulation.

Rubio CR, Simes JC, Moya M, Soriano F, Palma JA, Campana V.

Cátedra de Física Biomédica, Facultad de Ciencias Médicas, Universidad Nacional de Córdoba, Córdoba, Argentina.


Crystalopathies are inflammatory pathologies caused by cellular reactions to the deposition of crystals in the joints. The anti-inflammatory effect of the helium-neon (He-Ne) laser and that of the nonsteroidal anti-inflammatory drugs (NSAIDs) diclofenac, meloxicam, celecoxib, and rofecoxib was studied in acute and chronic arthritis produced by hydroxyapatite and calcium pyrophosphate in rats. The presence of the markers fibrinogen, L-citrulline, nitric oxide, and nitrotyrosine was determined. Crystals were injected into the posterior limb joints of the rats. A dose of 8 J/cm(2) of energy from an He-Ne laser was applied for 3 d in some groups and for 5 d in other groups. The levels of some of the biomarkers were determined by spectrophotometry, and that of nitrotyrosine was determined by ELISA. For statistical analysis, Fisher’s exact test was used, and p +/- 0.05 was considered significant. In arthritic rats, the fibrinogen, L-citrulline, nitric oxide, and nitrotyrosine levels increased in comparison to controls and to the laser-treated arthritic groups (p +/- 0.001), (p +/- 0.001), (p +/- 0.02), and (p +/- 0.01), respectively. When comparing fibrinogen from arthritic rats with disease induced by hydroxyapatite with undiseased and arthritic rats treated with NSAIDs, the He-Ne laser decreased levels to values similar to those seen in controls (p +/- 0.01). Inflammatory and oxidative stress markers in experimental crystalopathy are positively modified by photobiostimulation.

J Mol Cell Cardiol. 2009 Jan;46(1):4-14. Epub 2008 Sep 30.

Near infrared light protects cardiomyocytes from hypoxia and reoxygenation injury by a nitric oxide dependent mechanism.

Zhang R, Mio Y, Pratt PF, Lohr N, Warltier DC, Whelan HT, Zhu D, Jacobs ER, Medhora M, Bienengraeber M.

Department of Medicine, Medical College of Wisconsin, Milwaukee, WI 53326, USA.

Photobiomodulation with near infrared light (NIR) provides cellular protection in various disease models. Previously, infrared light emitted by a low-energy laser has been shown to significantly improve recovery from ischemic injury of the canine heart. The goal of this investigation was to test the hypothesis that NIR (670 nm) from light emitting diodes produces cellular protection against hypoxia and reoxygenation-induced cardiomyocyte injury. Additionally, nitric oxide (NO) was investigated as a potential cellular mediator of NIR. Our results demonstrate that exposure to NIR at the time of reoxygenation protects neonatal rat cardiomyocytes and HL-1 cells from injury, as assessed by lactate dehydrogenase release and MTT assay. Similarly, indices of apoptosis, including caspase 3 activity, annexin binding and the release of cytochrome c from mitochondria into the cytosol, were decreased after NIR treatment. NIR increased NO in cardiomyocytes, and the protective effect of NIR was completely reversed by the NO scavengers carboxy-PTIO and oxyhemoglobin, but only partially blocked by the NO synthase (NOS) inhibitor L-NMMA. Mitochondrial metabolism, measured by ATP synthase activity, was increased by NIR, and NO-induced inhibition of oxygen consumption with substrates for complex I or complex IV was reversed by exposure to NIR. Taken together these data provide evidence for protection against hypoxia and reoxygenation injury in cardiomyocytes by NIR in a manner that is dependent upon NO derived from NOS and non-NOS sources.

Klin Med (Mosk). 2009;87(6):22-5.

[Effect of intravenous laser blood irradiation on endothelial dysfunction in patients with hypertensive disease]

[Article in Russian]

Burduli NM, Aleksandrova OM.


The aim of this work was to study effect of intravenous laser blood irradiation (ILBI) on endothelial dysfunction in 120 patients (mean age 53.4 +/- 1.3 yr) with grade I-II hypertensive disease (HD) allocated to 2 groups. Traditional drug therapy given to patients of control group was supplemented by ILBI using a Mulat laser therapy device in the study group. Endothelial function was evaluated from the total plasma concentration of stable NOx metabolites, nitrates (NO3-), nitrites (NO2-), and Willebrand’s factor. HD patients were found to have elevated activity of the Willebrand factor and show 3 types of response of the NO generating system: (1) decreased NO synthesis, (2) lack of its changes, and (3) increased NO synthesis. NO production in HD patients negatively correlated with systolic (r = -0.59) and diastolic (r = – 0.64) arterial pressure (AP) which suggests the relationship between decreased NO production and elevated AP. Inclusion of ILBl in the therapy of HD resulted in a significant decrease of Willebrand’s factor activity and normalization of the NO level regardless of its initial value.

Klin Med (Mosk). 2009;87(1):49-52.

[Effect of low-intensity laser radiation on the function of vascular endothelium in patients with chronic viral hepatitis]

[Article in Russian]

Burduli NM, Krifaridi AS.


The aim of the study was to measure plasma levels of stable metabolites of nitric oxide, nitrates, and nitrites (NOx) in patients with chronic viral hepatitis and evaluate the possibility of their correction by low-power laser irradiation. NO metabolites (total nitrites and nitrates) were measured colorimetrically from the development of colour in the reaction of nitrite with sulfanilamide diazotization in Griess reagent. Colour intensity was determined with Victor2 enzyme immunoanalyzer, Perkin Elmaer (Finland). The patients were divided into three groups. In group 1 (control, n = 30) they received combined medicamentous therapy, in group 2 (n = 45) medicamentous therapy and a course of intravenous laser therapy, in goup 3 (n = 45) medicamentous therapy and skin laserotherapy. The results indicate that medicamentous treatment of patients with chronic hepatitis does not bring any beneficial changes in plasma NOx whose levels are significantly improved in case of simultaneous laser therapy. It is concluded that different laserotherapeutic modalities have beneficial effect on NO-producing function of endothelium and thereby improve its functional state. Compensation of NO deficit by laser therapy ensures overall protection of the organism against free radicals and decreases severity of oxidative stress.

Photomed Laser Surg. 2008 Oct;26(5):443-9.

Role of nitric oxide in the visible light-induced rapid increase of human skin microcirculation at the local and systemic levels: II. healthy volunteers.

Samoilova KA, Zhevago NA, Petrishchev NN, Zimin AA.

Institute of Cytology, Russian Academy of Sciences, St. Petersburg, Russia.


OBJECTIVE: The aim of this study is to evaluate the skin microcirculation increase seen in healthy volunteers after a single exposure to polychromatic visible (pVIS) light, and to prove the role of nitric oxide (NO) in the development of this effect.

BACKGROUND DATA: Improvement of microcirculation is one of the most important effects of laser and pVIS light therapy; however, its mechanism of action remains unknown. A main role in the regulation of vascular tone is known to be played by NO. It is produced by NO-synthase (NOS) located in membranes of many cells, including endothelial and blood cells. NOS, a biopteroflavohemoprotein, absorbs pVIS light, resulting in its activation.

MATERIALS AND METHODS: The central area of the dorsal side of the right hand (24 cm2) of 42 volunteers was irradiated for 5 min with pVIS light from a Q-light (385-750 nm, 95% polarization, 40 mW/cm2, 12 J/cm2). Then for 90 min, the blood flow rate (Qas) was measured eight times, both in the area of the irradiation (local effect) and in the non-irradiated left hand (systemic effect) by using a high-frequency ultrasound Doppler device, recording Qas in human skin to a depth up to 5 mm. In the central area of the right hand of 14 volunteers an NOS inhibitor, N-monomethyl-L-arginine (L-NMMA, 0.1% solution), was iontophoretically administered prior to exposure, whereas in 10 other subjects it was administered to the left hand with subsequent exposure of the right hand.

RESULTS: As soon as 2 min after exposure, Qas in the irradiated area rose on average by 32%, and in 20 min by 45%; it then decreased and in 90 min returned to the initial level. A statistically significant Qas increase in the non-irradiated hand was recorded in 5 min (+9%), and in 20 min it reached a maximum level (+39%), and 90 min later it decreased to the initial values. The presence of L-NMMA in the light-exposed area completely blocked the photoinduced rise of microcirculation, both in the irradiated and in non-irradiated hand; however, its administration to the non-irradiated hand did not prevent these effects.

CONCLUSION: The increase in skin microcirculation produced by pVIS light at the local and systemic levels is due to activation of NO synthesis in the irradiated area.

Lasers Surg Med. 2008 Jul;40(5):371-8.

Irradiation with 780 nm diode laser attenuates inflammatory cytokines but upregulates nitric oxide in lipopolysaccharide-stimulated macrophages: implications for the prevention of aneurysm progression.

Gavish L, Perez LS, Reissman P, Gertz SD.

Department of Anatomy and Cell Biology, The Hebrew University, Hadassah Medical School, Jerusalem 91120, Israel.


BACKGROUND AND OBJECTIVES: Low level laser irradiation (LLLI) has been shown to reduce inflammation in a variety of clinical situations. We have shown that LLLI (780 nm) increases aortic smooth muscle cell proliferation and matrix protein secretion and modulates activity and expression of matrix metalloproteinases. Inflammation is a major component of arteriosclerotic diseases including aneurysm. Macrophage recruitment and secretion of pro-inflammatory cytokines and the vasodilator, nitric oxide (NO), are central to most immune responses in the arterial wall. The present study was designed to determine the effect of LLLI on cytokine gene expression and secretion as well as gene expression of inducible nitric oxide synthase (iNOS) and NO production in lipopolysaccharide (LPS)-stimulated macrophages.

STUDY DESIGN/MATERIALS AND METHODS: Murine monocyte/macrophages (RAW 264.7) were irradiated with a 780 nm diode laser (2 mW/cm(2), 2.2 J/cm(2)) during stimulation with LPS (0, 0.1, and 1 microg/ml). Gene expression of chemokines, cytokines, and iNOS were assessed by RT-PCR. Secretion of interleukin (IL)-1beta and monocyte chemotactic protein (MCP)-1 and NO were assessed by ELISA and the Griess reaction, respectively.

RESULTS: LLLI reduced gene expression of MCP-1, IL-1alpha, IL-10 (P<0.01), IL-1beta, and IL-6 (P<0.05) when cells were stimulated by 1 microg/ml LPS. LLLI reduced LPS-induced secretion of MCP-1 over non-irradiated cells by 17+/-5% and 13+/-5% at 12 hours (0.1 and 1 microg/ml LPS; P<0.01 and P=0.05, respectively), and reduced IL-1beta by 22+/-5% and 25+/-9% at 24 hours (0.1 and 1 microg/ml LPS, P=0.01 and P=0.06, respectively). However, LLLI increased NO secretion after 12 hours (LLLI vs. Control: without LPS, 1.72+/-0.37 vs. 0.95+/-0.4 microM, P<0.05; 0.1 microg/ml LPS, 7.46+/-1.62 vs. 4.44+/-1.73 microM, P=0.06; 1 microg/ml LPS, 10.91+/-3.53 vs. 6.88+/-1.52 microM, P<0.05).

CONCLUSIONS: These properties of LLLI, with its effects on smooth muscle cells reported previously, may be of profound therapeutic relevance for arterial diseases such as aneurysm where inflammatory processes and weakening of the matrix structure of the arterial wall are major pathologic components.

Lasers Surg Med. 2008 Jan;40(1):46-54

Low-energy laser irradiation increases endothelial cell proliferation, migration, and eNOS gene expression possibly via PI3K signal pathway.

Chen CH, Hung HS, Hsu SH.

Institute of Biomedical Science, National Chung Hsing University, Taichung, Taiwan, Republic of China.

BACKGROUND AND OBJECTIVES: The purpose of this study, therefore, was to determine the mechanisms by which low-energy laser irradiation (LELI) may exert some of its angiogenic effects via the PI3 kinase/eNOS signaling pathway and induce endothelial cell migration and neovascularization, an important and necessary part of wound healing. STUDY DESIGN/MATERIALS AND METHODS: The possible molecular mechanism of helium-neon (He-Ne) laser irradiation on endothelial cells was proposed. He-Ne laser at 632.5 nm was used to stimulate human umbilical vein endothelial cell (HUVEC), and its effect on cell proliferation, nitric oxide secretion, and cell migration was determined. RESULTS: Irradiation enhanced endothelial nitric oxidase synthase (eNOS) protein expression, and irradiation of less than 0.26 J/cm(2) enhanced eNOS gene expression in HUVEC. The cell migration ability was promoted for HUVEC irradiated with 0.26 J/cm(2). This agreed with the vinculin protein expression induced by irradiation. In addition, the angiogenesis was promoted. The induced eNOS expression was inhibited by LY294002, indicating that the effect of laser on EC could be attributed to the up-regulation of eNOS expression through PI3K pathway at the cellular and molecular levels as a result of the He-Ne laser. CONCLUSIONS: The study has shown that LELI increased endothelial cell proliferation, migration, NO secretion, and identified that activation of PI3K/Akt pathway was a critical step for the elevated for eNOS expression upon LELI.

Lasers Med Sci. 2007 Mar;22(1):30-6. Epub 2006 Nov 21.

Irradiation at 634 nm releases nitric oxide from human monocytes.

Lindgård A, Hultén LM, Svensson L, Soussi B.

Wallenberg Laboratory, Sahlgrenska University Hospital, Göteborg University, Gothenburg, 413 45, Sweden.


Previous studies have shown that irradiation at 634 nm decreases the release of extracellular reactive oxygen species (ROS) without affecting viability in human monocytes. Here, we examined the effect of irradiation at 634 nm on the release of nitric oxide (NO), activation of inducible nitric oxide synthase (iNOS) and endothelial NOS (eNOS), and release of intracellular ROS. Chemiluminescence assays were used to measure NO release, intracellular ROS, and adenosine triphosphate levels (to assess cell viability). Levels of iNOS and eNOS mRNA were analyzed using PCR. Irradiation resulted in elevated levels of NO but had no effect on iNOS or eNOS. Irradiation also caused a decrease in levels of intracellular ROS and had no effect on cell viability. Our studies indicate that irradiation at 634 nm releases NO, possibly from a preformed store, and reduces the production of intracellular ROS without affecting cell viability. Irradiation at 634 nm may have a wide range of clinical applications, including a reduction in oxidative stress-mediated injury in the vasculature.

Mol Med. 2007 Jan-Feb;13(1-2):22-9.

Blue laser light increases perfusion of a skin flap via release of nitric oxide from hemoglobin.


Mittermayr R, Osipov A, Piskernik C, Haindl S, Dungel P, Weber C, Vladimirov YA, Redl H, Kozlov AV.

Ludwig Boltzmann Institute for Experimental and Clinical Traumatology in the AUVA Research Center, Vienna, Austria.

It has recently been shown that nitrosyl complexes of hemoglobin (NO-Hb) are sensitive to low-level blue laser irradiation, suggesting that laser irradiation can facilitate the release of biologically active nitric oxide (NO), which can affect tissue perfusion. The aim of this study was to evaluate the therapeutic value of blue laser irradiation for local tissue perfusion after surgical intervention. Blood was withdrawn from a rat, exposed to NO and infused back to the same rat or used for in vitro experiments. In vitro, an increase of NO-Hb levels (electron paramagnetic resonance spectroscopy) up to 15 microM in rat blood did not result in the release of detectable amounts of NO (NO selective electrode). Blue laser irradiation of NO-Hb in blood caused decomposition of NO-Hb complexes and release of free NO. Systemic infusion of NO-Hb in rats affected neither systemic circulation (mean arterial pressure) nor local tissue perfusion (Doppler blood flow imaging system). In contrast, a clear enhancement of local tissue perfusion was observed in epigastric flap when elevated NO-Hb levels in blood were combined with local He-Cd laser irradiation focused on the left epigastric artery. The enhancement of regional tissue perfusion was not accompanied by any detectable changes in systemic circulation. This study demonstrates that blue laser irradiation improves local tissue perfusion in a controlled manner stimulating NO release from NO-Hb complexes.

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.


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.

Lasers Surg Med. 2006 Aug;38(7):682-8.

Modulations of VEGF and iNOS in the rat heart by low level laser therapy are associated with cardioprotection and enhanced angiogenesis.

Tuby H, Maltz L, Oron U.

Department of Zoology, The George S. Wise Faculty of Life Sciences, Tel-Aviv University, Tel-Aviv, 69978, Israel.


BACKGROUND AND OBJECTIVES: It has been shown previously that low-level laser therapy (LLLT) significantly reduces infarct size following induction of myocardial infarction in rats and dogs. The aim of the present study was to investigate the effect of LLLT on the expression of vascular endothelial growth factor (VEGF) and inducible nitric oxide synthase (iNOS). STUDY DESIGN AND

MATERIAL AND METHODS: Myocardial infarction was induced by occlusion of the left descending artery in 87 rats. LLLT was applied to intact and post-infarction. VEGF, iNOS, and angiogenesis were determined.

RESULTS: Both the laser-irradiated rat hearts post-infarction and intact hearts demonstrated a significant increase in VEGF and iNOS expression compared to non-laser-irradiated hearts. LLLT also caused a significant elevation in angiogenesis.

CONCLUSIONS: It is concluded that VEGF and iNOS expression in the infarcted rat heart is markedly upregulated by LLLT and is associated with enhanced angiogenesis and cardioprotection.

Biofizika. 2006 May-Jun;51(3):509-18.

Effect of low-intensity laser radiation (632.8 nm) on immune cells isolated from mice.

[Article in Russian]

Novoselova EG, Cherenkov DA, Glushkova OV, Novoselova TV, Chudnovski? VM, Iusupov VI, Fesenko EE.


The effect of in vitro exposure to low-power laser light with a power density of 0.2 mW/cm2 and a wavelength of 632.8 nm induced by helium-neon laser on the functional activity of macrophages and splenic lymphocytes was studied. If the exposure period did not exceed 60 sec, the stimulation in interleukin-2 (IL-2) and nitric oxide (NO) production, as well as an increase in the activity of natural killer cells were observed. The increase of irradiation dose by prolongation of the exposure duration up to 180 s induced a significant decrease in NO production and natural killer cell activity, but IL-2 production was not different from the control level. A remarkable decrease in interferon-gamma (IFN-gamma) production was observed following laser light exposure of cells for 60 or 180 s, whereas under lower doses (exposure for 5 or 30 sec) IFN-gamma production increased. Irradiation of isolated macrophages induced a significant stimulation of cellular tumor necrosis factor-alpha (TNF- alpha) production at all dboes used, and, what is more important, an enhancement in both TNF-a phaand interleukin-6 (IL-6) production was revealed as early as after a 5-s exposure. In this case, more prolonged exposure periods, 60 and 180 s, either did not induce changes in IL-6 production (in macrophages) or decreased IL-6 production (in lymphocytes). Thus, upon in vitro exposure of cells to extremely low-power laser light, a basic tendency was observed: short-term irradiation predominantly induced stimulation in secretory activity of cells, whereas prolongation of exposure mainly induced immunosuppression. The only exception to the rule was a change in interleukin-3 (IL-3) production, which decreased after short-time exposure, but, on the opposite, increased when the cells were exposed for 180 s. In addition, a high sensitivity to extremely low-power laser light was supported by expression of the inducible heat shock protein, Hsp70, the effect being observed at all doses used, including the exposure for 5 s. At the same time, expression of another heat shock protein, Hsp90, was somewhat reduced after irradiation of cells with laser light.

Biofizika. 2006 Jan-Feb;51(1):116-22.

[A comparison of the effects of laser and light-emitting diodes on superoxide dismutase activity and nitric oxide production in rat wound fluid]

[Article in Russian]

Klebanov GI, Shuraeva NIu, Chichuk TV, Osipov AN, Vladimirov IuA.


The action of laser and light-emitting diode radiation in the visible region on the content of reactive nitrogen species and activity of superoxide dismutase in rat wound fluid was studied, and efficiency of action of coherent laser and incoherent light emitting diode radiations in the red region of the spectrum on the parameters under study was compared. A model of incised aseptic wounds in rats proposed by L.I. Slutskiy was used. A He-Ne laser (632 nm) and a Y-332B light emitting diode served as radiation sources. It was shown that (1) exposure of wounds to the visible light of both laser and light-emitting diodes causes dose-dependent changes in superoxide dismutase activity and production of nitrites and (2) the radiation coherence does not play any significant role in the changes of superoxide dismutase activity or nitrogen oxide formation by wound fluid phagocytes.

Lasers Surg Med. 2005 Apr;36(4):307-14.

Cellular effects of low power laser therapy can be mediated by nitric oxide.

Karu TI, Pyatibrat LV, Afanasyeva NI.

Institute of Laser and Information Technologies of the Russian Academy of Sciences, 142190 Troitsk, Moscow, Russia.


BACKGROUND AND OBJECTIVES: The objective of this study was to investigate the possibility of involvement of nitric oxide (NO) into the irradiation-induced increase of cell attachment. These experiments were performed with a view to exploring the cellular mechanisms of low-power laser therapy.

STUDY DESIGN/MATERIALS AND METHODS: A suspension of HeLa cells was irradiated with a monochromatic visible-to-near infrared radiation (600-860 nm, 52 J/m2) or with a diode laser (820 nm, 8-120 J/m2) and the number of cells attached to a glass matrix was counted after 30 minute incubation at 37 degrees C. The NO donors sodium nitroprusside (SNP), glyceryl trinitrate (GTN), or sodium nitrite (NaNO2) in the concentration range 5 x 10(-9)-5 x 10(-4)M were added to the cellular suspension before or after irradiation. The action spectra and the concentration and fluence dependencies obtained were compared and analyzed.

RESULTS: The well-structured action spectrum for the increase of the adhesion of the cells, with maxima at 619, 657, 675, 740, 760, and 820 nm, points to the existence of a photoacceptor responsible for the enhancement of this property (supposedly cytochrome c oxidase, the terminal respiratory chain enzyme), as well as signaling pathways between the cell mitochondria, plasma membrane, and nucleus. Treating the cellular suspension with SNP (5 x 10(-5)M) before irradiation significantly modifies the action spectrum for the enhancement of the cell attachment property (band maxima at 642, 685, 700, 742, 842, and 856 nm). The action of SNP, GTN, and NaNO2 added before or after irradiation depends on their concentration and radiation fluence.

CONCLUSIONS: The NO donors added to the cellular suspension before irradiation eliminate the radiation-induced increase in the number of cells attached to the glass matrix, supposedly by way of binding NO to cytochrome c oxidase. NO added to the suspension after irradiation can also inhibit the light-induced signal downstream. Both effects of NO depend on the concentration of the NO donors added. These results indicate that NO can control the irradiation-activated reactions that increase the attachment of cells.