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Effect of NASA light-emitting diode irradiation on wound healing

Whelan HT1, Smits RL Jr, Buchman EV, Whelan NT, Turner SG, Margolis DA, Cevenini V, Stinson H, Ignatius R, Martin T, Cwiklinski J, Philippi AF, Graf WR, Hodgson B, Gould L, Kane M, Chen G, Caviness J. - J Clin Laser Med Surg. 2001 Dec;19(6):305-14. (Publication)
Study showed increases in growth of 155-171% of normal human epithelial cells and an improvment of greater than 40% in musculoskeletal training injuries in Navy SEAL
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OBJECTIVE:

The purpose of this study was to assess the effects of hyperbaric oxygen (HBO) and near-infrared light therapy on wound healing.

BACKGROUND DATA:

Light-emitting diodes (LED), originally developed for NASA plant growth experiments in space show promise for delivering light deep into tissues of the body to promote wound healing and human tissue growth. In this paper, we review and present our new data of LED treatment on cells grown in culture, on ischemic and diabetic wounds in rat models, and on acute and chronic wounds in humans.

MATERIALS AND METHODS:

In vitro and in vivo (animal and human) studies utilized a variety of LED wavelength, power intensity, and energy density parameters to begin to identify conditions for each biological tissue that are optimal for biostimulation.

RESULTS:

LED produced in vitro increases of cell growth of 140-200% in mouse-derived fibroblasts, rat-derived osteoblasts, and rat-derived skeletal muscle cells, and increases in growth of 155-171% of normal human epithelial cells. Wound size decreased up to 36% in conjunction with HBO in ischemic rat models. LED produced improvement of greater than 40% in musculoskeletal training injuries in Navy SEAL team members, and decreased wound healing time in crew members aboard a U.S. Naval submarine. LED produced a 47% reduction in pain of children suffering from oral mucositis.

CONCLUSION:

We believe that the use of NASA LED for light therapy alone, and in conjunction with hyperbaric oxygen, will greatly enhance the natural wound healing process, and more quickly return the patient to a preinjury/illness level of activity. This work is supported and managed through the NASA Marshall Space Flight Center-SBIR Program.

Read more at: https://pdfs.semanticscholar.org/1f5b/0a4ce02a9c58dfd8531552fd2d2e2f3e701e.pdf

Original Source: https://www.ncbi.nlm.nih.gov/pubmed/11776448

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

- Photomedicine and Laser Surgery (Publication)
This study anaylizes the effect of whole-body LLLT on tissues harboring cancer and concluded that suggests that LLLT at these parameters may be safe even when malignant lesions are present.
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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/cm2 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.

Introduction

Low-level light therapy (LLLT) is being used increasingly for the treatment of a variety of conditions including trauma, wound healing, arthritis, musculoskeletal disorders, and dental and cosmetic applications.14 The current therapeutic approach is to be cautious of potential harmful effects from the use of LLLT in patients with cancer. Its use for the management of lymphedema and other complications in cancer patients has been withheld because of the fear that LLLT might promote metastasis.5,6. This approach is summarized by the review of Hawkins et al., which stated that “LLLT should be avoided or given with special caution in…patients with cancer if there is any doubt of a recurrence of metastases.…Although LLLT has not induced cancer in any of the reported studies, the precise reactions of existing tumors to LLLT are unknown.”6 There is little scientific evidence available as regards oncologic outcomes and local responses to LLLT in cancer patients. Although it is unlikely that LLLT would induce de novo cancer development as there is no evidence that LLLT causes DNA damage, its effects on cellular proliferation have been the empiric basis for withholding treatment in cancer patients.

Red light is known to have a mitogenic effect based on its ability to activate cell division at certain spectral and dose ranges in vitro.79 We are aware only of two studies on the effects of LLLT on cancer.10,11 Revazova demonstrated the acceleration of tumor growth by 633 nm laser irradiation at 3.5 J/cm2 three times per week for 2 weeks in a model of human gastric adenocarcinoma transplanted into immunodeficient athymic nude mice.11 This suggests that LLLT is indeed capable of activating tumor growth under conditions that exclude immune resistance. In another study, the irradiation of squamous cell carcinomas (SCC) in the hamster cheek pouch with 660 nm light at 56 J/cm2 and a 3 mm spot caused significant progression of the severity of SCC as judged by histology.10 The bulk of literature on the topic of LLLT and cancer does not address the question of LLLT effects on tumor growth.

The present study investigated the potential promotion of tumor growth by LLLT cause by the stimulation of cellular proliferation in cancerous cells. A standard nonmelanoma mouse skin cancer model was used to test the effect of automated full body photoirradiation twice a day at 670 nm and at an energy density 2.5 J/cm2 on tumor growth in already developed lesions.

We hypothesized that the systemic effects of phototherapy with red light might offset activation of cell division observed in vitro.

Discussion

The use of phototherapy in the treatment of cancer patients has been controversial. Current recommendations suggest that therapy should be carefully considered and used cautiously in patients with cancer, and that treatment in areas bearing tumors should be avoided. This empiric advice is based on our current knowledge of the experimental acceleration of cellular proliferation and stimulation of wound healing and tissue repair as demonstrated in both animal models and clinical scenarios.1618

There have been few studies that have investigated the influence of LLLT on tumors and tumor growth. The hamster cheek pouch DMBA-induced oral SCC has been recently investigated by Monteiro et al.10 The authors treated the oral cavity with 660 nm LLLT after induction of tumors. Histological evaluation demonstrated an increase in the progression and severity of SCC.10

Liebow et al. had also demonstrated an apparent stimulation of tumor induction and growth after CO2 laser incisions were created in cheek pouch tissue that had been transformed as a result of DMBA painting.19,20 Both the Montiero and Liebow investigations involved manipulations of tissue that had been manipulated into a transformed field as a result of DMBA induction. This process inevitably results in tumor formation and it is well known that scalpel incisions and other perturbations of the epithelium can stimulate tumor induction. It is also well known that these tumors are dependent upon epidermal growth factor (EGF) for growth.21 Saliva contains significant concentrations of EGF and other growth factors and cytokines. Inflammation results in consumption and degradation of these growth factors, and processes that reduce or modulate the inflammatory response similarly affect tumor development in these tissues. CO2 laser use results in a reduction and delay in the inflammatory response.2225 This particular laser is capable of inducing heat shock proteins by a mechanism similar to that observed in modification of wound healing and scar formation in laser-assisted-scar-healing (LASH) in humans.26,27 Similarly, phototherapy at 660 nm is known to reduce inflammation.28

Both of these studies demonstrate that the local milieu is important in the induction and proliferation of malignant lesions. However, it would not be appropriate to make generalizations about all types of cancers based on this very specific model and tumor system.

The model

We chose a model that can produce a large number of malignant cutaneous lesions economically and automatically (Fig. 2), provides a way to irradiate them with red light automatically (Fig. 3), and allows us to monitor the growth of these tumors daily. This experimental model (Fig. 1) induces spontaneous and genetically heterogeneous nonmelanoma skin cancers on the backs of hairless mice after UV damage. The induction of cancer by UV exposure is a random process and involves a combination of randomly induced mutations in multiple genes per tumor. The tumors produced by this model are heterogeneous, which is more representative of a wider range of clinically observed cancers as contrasted to models that use genetically homogeneous cancer cell lines. Although nonmelanoma skin cancer is not as deadly as other cancers in humans, it is a true cancer genetically and functionally and therefore with the effects of red light, LLLT in the presence of these neoplasms is relevant to the potential effects of red light therapy on other types of cancer.

The advantage of SKH-1 mouse cancer model is that the cells producing cancer in the overwhelming majority are epidermal keratinocytes, that is the fast-dividing keratinocytes of the lowest layer of epidermis, which is very thin in mice, less than 0.05 mm. Therefore, the tumors grow on the surface of the skin and a very minor part of each tumor is below the surface.2933 Early investigations using the SKH-1 model documented the high degree of histologic similarity in the numerous cutaneous malignant lesions produced in this model.2933 In addition, the high throughput method of periodic photographing the tumors and measuring their diameter on the photographs, a well-established method of measurement, fosters the analysis of hundreds of tumors longitudinally, which is not possible with other end-point methods, such as histology. The majority of the research studies utilizing this SKH-1 cancer model use the size of the visible tumor as a function of time as a measure of tumor proliferation.12,3441

The sensitivity of the model to detect small therapeutic effects is limited by the fact that the tumors in treatment and control groups are by their nature different genetically, as each tumor is a result of random mutagenic events. Although this difference is of no significance, because of large numbers of medium-sized tumors in both groups, the individual random mutations resulting in the induction of small numbers of large fast growing tumors potentially affected the overall statistical results. This limitation can be overcome in the future by increasing the number of mice treated or by measuring baseline growth rates for each tumor before the beginning of red light therapy, and then comparing the growth rate of each tumor before and after beginning the therapy.

Automation and human interventions

We have developed a new method that uses a well-characterized animal model for the study of the effects of LLLT on cancer. The advantage of this model is that the setup minimizes the human factor, both in influencing mouse behavior and in data analysis. The mice are irradiated automatically and the tumors are sized across time using image morphometry blinded to treatment, thus excluding human bias.

Evidence supporting the safety of red light

This study aimed to maximize the healing and activation effects while avoiding the inhibitory effects of red light. We selected the red light dose and fluence very conservatively based on our prior studies on wound healing.13 Treated mice received two irradiations per day at 8 mW/cm2 fluence for 312 sec per session, resulting in a total dose density of 2.5 J/cm2 per session (5 J/cm2 per day). This regimen is in general agreement with the one used by Erdle et al.14 Erdle et al. used the same red light source and mouse strain (SKH-1), measured incisional wound healing, and demonstrated the high efficiency of chronic daily treatment at a dose of 3.6 J/cm2 (either 450 sec at 8 mW/cm2 or 37 min at 1.6 mW/cm2).

This study documented the absence of strong positive or negative effects of LLLT on tumor growth in this model and red light treatment parameters. Prior studies using the same red LLLT system demonstrated that these parameters stimulate wound healing.13 The present study provides some evidence that phototherapy at these parameters should not be empirically contraindicated in the treatment of patients with cancer. Our qualitative observations of improvement in skin quality at early time points, and relief of sickness behavior at later stages of the investigation, are also suggestive of the fact that the light was capable of producing beneficial effects for the whole animal despite the presence of tumors. It should be recognized, however, that the present study delivers, essentially, whole body therapy to the affected individual, rather than treating a specific area.

The small but statistically significant decrease in tumor area observed on days 16–23 demonstrates the ability of our model to detect small changes in tumor volume because of the low degree of random histotype variability in the model and the high number of examined tumors and time points. An additional explanation as to why red light was beneficial at days 16–23 may be the stimulation of antitumor immune activity or, perhaps, a local photodynamic effect as a result of red light activation of endogenous porphyrins present in tumors in and around areas of spontaneous hemorrhage and necrosis. Red light treatment was qualitatively observed to relieve sickness behavior, which suggests that there was an improved host response and increased antitumor immunity; at least until the tumor burden overwhelmed these effects. Future studies directed at studying these immune effects would be helpful in determining the biological basis for these observations.

Targets of the red light

Important factors to consider are: what tissues were reached by the red light during whole body phototherapy as was the case in this study, and which chromophores are absorbing the light. Because the mice have hairless fair skin, the light was not shielded by hair or melanin. The necrotic tissue covering some of the tumors might have shielded some tumor cells from the red light and/or may have generated local photodynamic effects caused by interaction with endogenous porphyrins. Much of the light likely did penetrate deeper in the mouse, potentially stimulating lymphatic vessels, lymph nodes, internal organs such as the spleen, and, possibly, even the bone marrow. It is likely that both actively dividing tumor cells and immune cells including white blood cells; immune cells infiltrating the skin such as mast cells, dendritic cells, neutrophils, and other, lymphatic vessels and nodes; bone marrow; and, possibly, spleen were absorbing and being activated by the light treatments.

As this study suggests that the outcome of red light therapy depends upon competition between possible activation of tumor growth on the one hand, and improvement of systemic antitumor immune response on the other, future studies should address the issue of local versus systemic red light therapy. Treatment was systemic in this case because of whole-body photoirradiation. Specific studies would be helpful, particularly if treatment can be isolated and directed solely to healthy tissues, both tumor-bearing and healthy tissue, or tumors alone.

Conclusions

The present study failed to demonstrate a harmful effect of whole-body red LLLT on tumor growth in an experimental model of UV-induced SCC. There was a transient and small reduction in relative tumor area in the treatment group compared with controls. This study suggests that LLLT should not be withheld from cancer patients on an empiric basis. Further investigations designed to build upon these observations and determine the mechanism for the host–tumor responses noted during the early treatment phase are warranted.


Original Source: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3423866/

When is the best moment to apply photobiomodulation therapy (PBMT) when associated to a treatmill endurance-training program? A randomized, triple-blinded, placebo-controlled clinical trial.

Eduardo Foschini MirandaShaiane Silva TomazoniPaulo Roberto Vicente de PaivaHenrique Dantas PintoDenis SmithLarissa Aline SantosPaulo de Tarso Camillo de CarvalhoErnesto Cesar Pinto Leal-Junior - Lasers in Medical Science May 2018 (Publication)
A studying showing the benefits of using LEDT before and after a cardio workout.
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Abstract

Photobiomodulation therapy (PBMT) employing low-level laser therapy (LLLT) and/or light emitting diode therapy (LEDT) has emerged as an electrophysical intervention that could be associated with aerobic training to enhance beneficial effects of aerobic exercise. However, the best moment to perform irradiation with PBMT in aerobic training has not been elucidated. The aim of this study was to assess the effects of PBMT applied before and/or after each training session and to evaluate outcomes of the endurance-training program associated with PBMT. Seventy-seven healthy volunteers completed the treadmill-training protocol performed for 12 weeks, with 3 sessions per week. PBMT was performed before and/or after each training session (17 sites on each lower limb, using a cluster of 12 diodes: 4 × 905 nm super-pulsed laser diodes, 4 × 875 nm infrared LEDs, and 4 × 640 nm red LEDs, dose of 30 J per site). Volunteers were randomized in four groups according to the treatment they would receive before and after each training session: PBMT before + PBMT after, PBMT before + placebo after, placebo before + PBMT after, and placebo before + placebo after. Assessments were performed before the start of the protocol and after 4, 8, and 12 weeks of training. Primary outcome was time until exhaustion; secondary outcome measures were oxygen uptake and body fat. PBMT applied before and after aerobic exercise training sessions (PBMT before + PBMT after group) significantly increased (p < 0.05) the percentage of change of time until exhaustion and oxygen uptake compared to the group treated with placebo before and after aerobic exercise training sessions (placebo before + placebo after group) at 4th, 8th, and 12th week. PBMT applied before and after aerobic exercise training sessions (PBMT before + PBMT after group) also significantly improved (p < 0.05) the percentage of change of body fat compared to the group treated with placebo before and after aerobic exercise training sessions (placebo before + placebo after group) at 8th and 12th week. PBMT applied before and after sessions of aerobic training during 12 weeks can increase the time-to-exhaustion and oxygen uptake and also decrease the body fat in healthy volunteers when compared to placebo irradiation before and after exercise sessions. Our outcomes show that PBMT applied before and after endurance-training exercise sessions lead to improvement of endurance three times faster than exercise only.

Introduction

Physical activity is recommended and beneficial for both asymptomatic persons and individuals with chronic diseases [1, 2]. Aerobic endurance is considered a useful tool for the assessment of physical fitness and the detection of changes in aerobic fitness resulting from systematic training [3].

Regular aerobic exercise has various beneficial metabolic, vascular, and cardiorespiratory effects [4]. Additionally, it decreases body fat and increases muscle mass, muscle strength, and bone density [5]. Moreover, it improves self-esteem and physical and mental health and reduces the incidence of anxiety and depression [4, 6].

Various ergogenic agents, such as whey protein [7], caffeine [8], creatine [9], and neuromuscular electrical stimulation [10], are currently used to increase the benefits of aerobic training. Photobiomodulation therapy (PBMT) has emerged as an electrophysical intervention that could be associated with aerobic training to enhance beneficial effects of aerobic exercise, since several studies used PBMT to improve physical performance when associated with different kinds of exercise [11, 12, 13, 14].

Several studies have recently used PBMT to improve muscle performance during aerobic activities in healthy adults [15, 16, 17, 18] and postmenopausal women [19, 20]. However, to the best of our knowledge, the best moment to perform irradiation with PBMT in aerobic training has not been yet elucidated.

For instance, the current literature shows that the application of PBMT before progressive aerobic exercise has ergogenic effects and acutely increases the time until exhaustion, covered distance, and pulmonary ventilation and decreases the score of dyspnea during progressive cardiopulmonary test [15]. In addition, PBMT irradiation performed prior to aerobic exercises improves the exercise performance by decreasing the exercise-induced oxidative stress and muscle damage [18] and increasing the oxygen extraction by peripheral muscles [16]. When performed during aerobic training sessions, PBMT improves the quadriceps power and reduces the peripheral fatigue in postmenopausal women [19, 20]. Additionally, when applied after the sessions of endurance-training program, PBMT leads to a greater fatigue reduction than endurance training without PBMT irradiation [17].

Therefore, the optimal moment to perform PBMT in aerobic training is still open to discussion. With this perspective in mind, we aimed to assess the effects of PBMT applied at different time points (before and/or after) of each training session and its potential effects on the outcomes of an endurance-training program (aerobic exercise).

Materials and methods

Study design and protocol

We performed a triple-blind (assessors, therapists, and volunteers), placebo-controlled, randomized clinical trial. The study was conducted in the Laboratory of Phototherapy in Sports and Exercise.

Ethical aspects

All participants signed informed consent prior to enrollment and the study was approved by the research ethics committee of Nove de Julho University (process 553.831) and registered at Clinical Trials.gov (NCT02874976).

Sample

The sample size was calculated assuming a type I error of 0.05 and a type II error of 0.2, based on previous study [21], and the primary established outcome was the time until exhaustion.

Inclusion and exclusion criteria

We recruited 96 healthy volunteers (48 men and 48 women) between 18 and 35 years of age and without training or involvement in a regular exercise program (i.e., exercise more than once per week) [22, 23]. Volunteers were excluded if they had any skeletal muscle injury, used any nutritional supplement or pharmacologic agent, presented with signs or symptoms of any disease (i.e., neurologic, inflammatory, pulmonary, metabolic, oncologic), or had a history of cardiac arrest that might limit performance of high-intensity exercises. Volunteers that were unable to attend a minimum rate of 80% of the training sessions and volunteers with immune diseases that require continuous use of anti-inflammatory drugs were also excluded.

Randomization and blinding procedures

Volunteers were distributed in four experimental groups (24 volunteers in each group) through a simple drawing of lots (A, B, C, or D) that determined the moment they would receive active and/or placebo PBMT treatment:
  • PBMT + PBMT: volunteers were treated with active PBMT before and after each training session.

  • PBMT + placebo: volunteers were treated with active PBMT before and placebo PBMT after each training session.

  • Placebo + PBMT: volunteers were treated with placebo PBMT before and active PBMT after each training session.

  • Placebo + placebo: volunteers were treated with placebo PBMT before and after each training session.

Randomization labels were created by using a randomization table at a central office where a series of sealed, opaque, and numbered envelopes ensured confidentiality. The researcher who programmed the PBMT device (manufactured by Multi Radiance Medical™, Solon, OH, USA) based on the randomization results was not involved in any other procedure of the study. He was instructed not to inform the participants or other researchers of the PBMT program (active or placebo). None of the researchers involved in aerobic endurance-training assessments and data collection knew which program corresponded to active or placebo PBMT.

Identical PBMT devices were used in both programs (active or placebo) by a researcher who was not involved in any phase of the projected data collection to ensure the study blinding. All displays and sounds emitted were identical regardless of the selected program. The active PBMT treatment did not demonstrate discernable amounts of heat [24].

Therefore, volunteers were unable to differentiate between active or placebo treatments. All volunteers were required to wear opaque goggles during treatments to safety and to maintain the triple-blind design.

Procedures

The study included three sessions of aerobic endurance training per week performed over 12 weeks, and each session lasted 30 min; the load for each exercise session (treadmill speed) progressed constantly in order to keep subjects’ heart rate between 70 and 80% from maximum heart rate. The assessments were conducted before the start of the training protocol and after 4, 8, and 12 weeks of training. A summary of the study design is presented in Fig. 1.
Fig. 1

CONSORT flowchart

Cardiopulmonary exercise test

Participants performed a standardized progressive cardiopulmonary exercise test on a treadmill with a fixed inclination of 1% until exhaustion. They began the test with a 3-min warm-up at a velocity of 3 km/h. Next, the treadmill velocity was increased by 1 km/h at 1-min intervals until the velocity of 16 km/h was reached. Participants were instructed to use hand signals to request termination of the test at any time. A 3-min recovery phase at a velocity of 6 km/h was allowed after each test [18]. During testing, we monitored the rates of oxygen uptake (VO2), carbon dioxide production measured with a VO 2000 gas analyzer (Inbrasport, Indústria Brasileira de Equipamentos Médico-Desportivos LTDA, Porto Alegre, RS, Brazil), total time until exhaustion, and heart rate measured with a digital electrocardiograph (Medical Graphs Ergomet, São Paulo, SP, Brazil).

These data were used to evaluate the performance of participants during progressive cardiopulmonary exercise testing, because this test is currently the most widely used in the literature for this purpose [25]. The entire test was monitored by electrocardiogram and blood pressure measurement. If any abnormal heart rate or blood pressure changes were observed or if the test was terminated prematurely on request, the test was stopped, and the volunteer’s data were deleted.

Body composition assessment

Body composition was assessed by the same technician (blinded to volunteer’s allocation in different experimental groups) using the procedures established by ISAK [26]. Measurements of height, body mass, and skinfolds were used to establish the percentage of fat [26].

Aerobic training protocol

Aerobic treadmill training, associated or not with PBMT, was performed three times a week for 12 weeks, each session lasting 30 min, with training intensity kept between 70 and 80% of maximum heart rate [27]; changes in running speed (training load) were constantly performed to achieve the 70–80% heart rate.

Training was interrupted based on the criteria established by the guidelines of the American Heart Association. Training intensity was monitored by a heart rate monitor manufactured by Polar®.

Photobiomodulation therapy

PBMT was applied employing MR4 Laser Therapy Systems outfitted with LaserShower 50 4D emitters (both manufactured by Multi Radiance Medical, Solon, OH, USA). The cluster style emitter contains 12 diodes composing of four super-pulsed laser diodes (905 nm, 0.3125 mW average power, and 12.5 W peak power for each diode), four red LED diodes (640 nm, 15 mW average power for each diode), and four infrared LEDs diodes (875 nm, 17.5 mW average power for each diode).

The cluster probe was selected due to the available coverage area and to reduce the number of sites needing treatment. Treatment was applied in direct contact with the skin with a slight applied overpressure to nine sites on extensor muscles of the knee (Fig. 2a), six sites on knee flexors of the knee, and two sites on the calf (Fig. 2b) of both lower limbs [15, 28]. To ensure blinding, the device emitted the same sounds and regardless of the programmed mode (active or placebo). The researcher, who was blinded to randomization and the programming of PBMT device, performed the PBMT.
Fig. 2

a Treatment sites at knee extensor muscles. b Treatment sites at knee flexor and ankle plantar flexor muscles

PBMT parameters and irradiation sites were selected based upon previous positive outcomes demonstrated with the same family of device [13, 15, 28, 29]. Table 1 provides a full description of the PBMT parameters. The volunteers received PBMT or placebo from 5 to 10 min before and/or after aerobic training sessions.

 

Statistical analysis

The obtained results were tested for their normality through the Shapiro-Wilk test. Since the data showed a normal distribution, two-way ANOVA test with Bonferroni post hoc analysis was applied. The data were described as mean values with the respective standard deviations and both absolute and percentage values were analyzed. Graphical data are described as mean and standard errors of mean (SEM). The level of statistical significance was p < 0.05.

Results

After data collection, we analyzed the results of 77 volunteers of both genders (PBMT + PBMT: 18 volunteers; PBMT + placebo: 21 volunteers; placebo + PBMT: 18 volunteers; and placebo + placebo: 20 volunteers) that had completed the aerobic training protocol after 12 weeks (Fig. 1). None of the recruited volunteers were excluded due abnormal heart rate or blood pressure during the execution of procedures of this study. The characteristics of the volunteers are summarized in Table 2.

 
 

As shown in Table 2, no statistically significant differences (p > 0.05) were found for anthropometric variables and baseline data among the different experimental study groups.

Table 3 shows all results of cardiopulmonary progressive test in absolute values for different variables analyzed in all experimental groups of this study. We observed a statistically significant improvement in oxygen uptake when PBMT was performed before and after training sessions (PBMT + PBMT group), comparing baseline values vs 4-, 8-, and 12-week values (p < 0.001). The same was observed for pulmonary ventilation, comparing baseline values vs 8- and 12-week values (p = 0.0018 and p = 0.003, respectively), and for time until exhaustion, comparing baseline values vs 4-, 8-, and 12-week values (p < 0.001).
Table 3

Progressive endurance test variables

   

Baseline

4 weeks

8 weeks

12 weeks

VO2 (mL/kg/min)

PBMT + PBMT

35.8 ± 9.5

40.2 ± 10.2*

41.5 ± 10.4*

42.5 ± 11.2*

PBMT + Placebo

34.8 ± 7.0

37.6 ± 7.0

38.6 ± 8.0

38.2 ± 7.0

Placebo + PBMT

35.2 ± 8.9

36.6 ± 8.1

38.6 ± 8.3

38.5 ± 8.3

Placebo + placebo

36.2 ± 7.7

36.8 ± 8.0

37.6 ± 7.5

38.4 ± 10.1

VCO2 (mL/kg/min)

PBMT + PBMT

38.7 ± 7.0

40.4 ± 8.6

41.3 ± 7.8

41.4 ± 8.7

PBMT + placebo

38.,5 ± 7.8

39.5 ± 6.6

41.7 ± 7.9

41.9 ± 6.8

Placebo + PBMT

38.5 ± 9.5

38.2 ± 9.5

41.5 ± 8.4

40.7 ± 9.6

Placebo + placebo

38.8 ± 10.6

40.7 ± 9.4

43.1 ± 13.4

40.9 ± 10.5

VE (mL/kg/min)

PBMT + PBMT

73.6 ± 22.8

77.9 ± 21.5

83.5 ± 24.5*

85.3 ± 22.5*

PBMT + Placebo

70.6 ± 20.3

71.0 ± 23.1

78.1 ± 23.0

77.2 ± 22.1

Placebo + PBMT

66.2 ± 25.3

70.6 ± 24.2

73.9 ± 20.6

73.4 ± 20.7

Placebo + placebo

69.9 ± 17.9

70.8 ± 18.8

70.3 ± 22.4

77.1 ± 18.3

Time until exhaustion (s)

PBMT + PBMT

681.5 ± 111.9

752.1 ± 111.7*

787.7 ± 114.2*

808.5 ± 124.5*

PBMT + placebo

698.7 ± 131.1

739.3 ± 142.2

773.4 ± 165.9

792.1 ± 186.9

Placebo + PBMT

693.1 ± 106.9

738.4 ± 116.6

766.1 ± 121.0

797.0 ± 139.0

Placebo + placebo

699.5 ± 137.3

720.2 ± 150.0

741.3 ± 154.3*

766.1 ± 159.8*

Data is expressed in average and standard deviation (±)

VO 2 oxygen uptake, VCO 2 carbon dioxide production, VE pulmonary ventilation

*Statistically significant difference compared to baseline (p < 0.05)

Furthermore, PBMT applied before and after each aerobic exercise training session (PBMT + PBMT group) significantly increased (p < 0.05) the percentage change of oxygen consumption and time-to-exhaustion compared to the group treated with placebo before and after each aerobic exercise training session (placebo + placebo group) from 4th to 12th week. Similarly, PBMT applied before and after each aerobic exercise training session (PBMT + PBMT group) significantly improved (p < 0.05) the percentage change of body fat compared to group treated with placebo before and after each aerobic exercise training session (placebo + placebo group). The outcomes are summarized in Figs. 3, 4, and 5, respectively.

Fig. 3

Percentage of change in time-to-exhaustion. The data are presented in mean and SEM. Letter a indicates statistical significance between PBMT + PBMT and placebo + placebo (p < 0.05)

Fig. 4

Percentage of change in maximum oxygen uptake. The data are presented in mean and SEM. Letter a indicates statistical significance between PBMT + PBMT and placebo + placebo (p < 0.05)

Fig. 5

Percentage of change in body fat. The data are presented in mean and SEM. Letter a indicates statistical significance between PBMT + PBMT and placebo + placebo (p < 0.05)

Discussion

To the best of our knowledge, this is the first study aiming to test the optimal moment to perform PBMT in an aerobic training protocol (before, after, or before and after training). Few studies have assessed chronic effects of PBMT [17, 20, 21]; however, PBMT has been applied at different moments (before, after, or during exercise) of the aerobic training program. Briefly, we observed that the combination of super-pulsed lasers and LEDs applied before and after exercise sessions increased the oxygen uptake, time-to-exhaustion, and reduced body fat in healthy sedentary volunteers after 12 weeks of aerobic training.

Paolillo et al. [20] investigated the effects of PBMT applied during the sessions of aerobic training on the treadmill in 20 postmenopausal women. The training was performed twice a week for 3 months, with an intensity of 85–90% of maximum heart rate. The volunteers received LED therapy with 850 nm, 31 mW/cm2, 30 min irradiation, and 14,400 J applied bilaterally to the tight regions. PBMT increased the exercise tolerance time when compared to the control group. These data corroborate with the results of our study, however, we used different light sources and wavelengths simultaneously (4 × 905 nm super-pulsed lasers, 4 × 875 nm infrared LEDs, and 4 × 640 nm red LEDs) to irradiate the volunteers and we found an increase in exercise tolerance of 13.4%. The magnitude of the difference in outcomes between studies might be related to the used irradiation protocol (in our study, the volunteers were irradiated before and after the aerobic training sessions, while Paolillo et al. [20] irradiated volunteers during the training sessions).

The same authors [21] also investigated the effects of PBMT (infrared LEDs—850 nm) when applied during treadmill training in 45 postmenopausal women. The training was performed twice a week for 6 months, and each training session lasted 45 min. The authors found a significant increase in exercise tolerance, and metabolic equivalents, and a longer duration of Bruce test. In our study, the association of PBMT before and after sessions of the aerobic training program was able to increase the oxygen consumption (with 18.7%) and time-to-exhaustion (with 13.4%) and improve the percentage of change of body fat (with 13.9%) after only 12 weeks of aerobic training.

Duarte et al. [30] evaluated the effects of PBMT (808 nm) associated with aerobic and resistance training performed three times a week for 16 weeks in obese women. The authors found a significant decrease in the percentage of fat and in neck and waist circumference. It is important to highlight that in our study, we observed statistically significant improvement in the percentage of change of body fat (13.9%) after only 12 weeks of aerobic training when associated with PBMT before and after the training sessions. We believe that the association of PBMT before and after training was able to enhance the performance and the tolerance of the volunteers during the aerobic training protocol, favoring the reduction of the body fat at the end of the 12 weeks of training.

It is interesting how outcomes in the fourth week for PBMT + PBMT group were similar to those of placebo + placebo group (or exercise alone) in the 12th week. This means that PBMT with optimal irradiation protocol (before and after exercise training sessions) can increase the endurance capacity of volunteers three times faster than exercise alone.

Regarding the mechanisms of the observed effects, we strongly believe that mitochondrial activity modulation is the key mechanism, despite the fact that our study only focused on clinical and functional aspects and not on mechanisms. Hayworth et al. [31] demonstrated that the activity of cytochrome c oxidase is enhanced by PBMT with a single wavelength in skeletal muscle fibers of rats. More recently, Albuquerque-Pontes et al. [32] showed that PBMT with different wavelengths (660, 830, or 905 nm) was able to increase the expression of cytochrome c oxidase in the intact skeletal muscle tissue in different time windows (5 min to 24 h after irradiation), which means that the muscle metabolism can be improved through the action of PBMT. These findings help us to explain the increase in performance observed by the use of PBMT associated with an aerobic training protocol and provide the rationale for the concurrent use of different wavelengths at the same time, which can represent a therapeutic advantage in various clinical situations.

In fact, different studies have shown that the concurrent use of different light sources and wavelengths enhances muscular performance [13, 14, 15, 28, 29, 33] decreases pain [34


Original Source: https://link-springer-com.colorado.idm.oclc.org/article/10.1007%2Fs10103-017-2396-2

Light-emitting diode therapy in exercise-trained mice increases muscle performance, cytochrome c oxidase activity, ATP and cell proliferation

Cleber Ferraresi, Nivaldo Antonio Parizotto, Marcelo Victor Pires de Sousa, Beatriz Kaippert, Ying?Ying Huang, Tomoharu Koiso, Vanderlei Salvador Bagnato, Michael R. Hamblin - Wiley Online Library/ 09-01-2015 (Publication)
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Abstract

Light-emitting diode therapy (LEDT) applied over the leg, gluteus and lower-back muscles of mice using a LED cluster (630 nm and 850 nm, 80 mW/cm2, 7.2 J/cm2) increased muscle performance (repetitive climbing of a ladder carrying a water-filled tube attached to the tail), ATP and mitochondrial metabolism; oxidative stress and proliferative myocyte markers in mice subjected to acute and progressive strength training. Six bi-daily training sessions LEDT-After and LEDT-Before-After regimens more than doubled muscle performance and increased ATP more than tenfold. The effectiveness of LEDT on improving muscle performance and recovery suggest applicability for high performance sports and in training programs.

 

Positioning of the mice and light-emitting diode therapy (LEDT) applied on mouse legs, gluteus and lower-back muscles without contact.

Introduction

Low-level laser (light) therapy has several applications in medicine such as treatment of pain 1, 2, tendinopathies 3 and acceleration of tissue repair 2, 4. Since the 1960s when the first laser (Light Amplification by Stimulated Emission of Radiation) devices were constructed, many applications of this therapy and its mechanisms of action have been investigated around the world 5.

Light therapy can be delivered by different light sources such as diode lasers or light emitting diodes (LEDs). These light sources differ in monochromaticity and coherence, since diode lasers are coherent with a tiny spectral bandwidth and less divergence of the light beams compared to the light emitted by LEDs 5. The spectral regions generally used for light therapy range between red (600 nm) to near infrared (1,000 nm) with total power in range of 1 mW–500 mW and power density (irradiance) in the range of range 1 mW–5 W/cm2 5. These lasers and LEDs are considered to produce equivalent effects on the tissue if the dose of light delivered/applied is in accordance with the possible biphasic dose?response previously reported 5-7. The light?tissue interaction depends on light absorption by specific structures in the cells that are known as chromophores 8-11.

Recently light therapy using lasers and LEDs has been used to increase muscle performance in exercises involving strength 12 or fatigue resistance 13-15; and light therapy may have a role to play in preparing athletes competing in high performance sports. Recent reviews have reported positive effects of light therapy on muscle performance, highlighting protection from exercise?induced muscle damage 16; an increased number of repetitions in maximum exertion tests 17; increased workload, torque and muscle fatigue resistance in training programs; as well as an overview of the main possible mechanisms of action of the light therapy on muscle tissue 18.

Several biological factors govern success or optimum performance in sports that involve high?intensity exercise, or alternatively involve endurance exercise, that both require muscle adaptation during pre?competition training programs. Among these factors are the depletion of the energy supply for muscle contraction which comprises adenosine triphosphate (ATP) and glycogen; accumulation of possibly deleterious metabolites from energy metabolism such as lactate, adenosine diphosphate (ADP), adenosine monophosphate (AMP), ions Ca2+ and H+; production of reactive oxygen species (ROS) 19-22; and the recovery process from microlesions or muscle damage 23. Light therapy seems to be able to benefit all these ”limitations” since its mechanism of action involves the improvement of mitochondrial metabolism and increased ATP synthesis 24, 25 owing to increased activity of cytochrome c oxidase (COX) in the electron transport chain (ETC) 9, 25, 26; reduction of reactive oxygen species (ROS) or improvement of oxidative stress defense 27, 28; and can stimulate faster muscle repair due to an increased proliferation and differentiation of muscle cells 29.

Experimental and clinical trials with different methodologies have reported the benefits of light therapy on muscle performance when applied before 15, 30, 31 or after exercise 12, 13, 32. However there is no consensus about the best time regimen for use of light therapy 18. The best wavelength (red or infrared) to stimulate muscle cells and increase muscle performance is also unclear.

In the current study we used an experimental model of mice exercising on a ladder similar to that reported in a previous study 33, in order to simulate a clinical strength training program that would allow us to identify which light therapy regimen would be better to increase muscle performance. Four different regimens of light therapy were applied to the mouse leg, gluteus and lower?back muscles during a training program: sham; before; before?after; and after each training session. Light therapy was delivered from LEDs (LEDT) with two simultaneous wavelengths (red and infrared). Assessment of muscle performance (load, number of repetitions, muscle work and power), markers of cellular energy and metabolism (ATP, glycogen and COX), oxidative stress markers (protein carbonyls, glutathione, catalase activity, lipid peroxidation, protein thiols) and muscle cell proliferation (BrdU – 5?bromo?2′?deoxyuridine) and adult myonuclei (DAPI – 4′,6?diamidino?2?phenylindole) were carried out.

Materials and methods

Animals

This study was performed with 8 week?old male Balb/c mice, weighing on average 22.22 g (SEM 0.24), housed at five mice per cage and kept on a 12 hour light 12 hour dark cycle. The 22 animals were provided by Charles River Inc and were provided with water and fed ad libitum at the animal facility of Massachusetts General Hospital. All procedures were approved by the IACUC of Massachusetts General Hospital (protocol #2014N000055) and met the guidelines of the National Institutes of Health.

Experimental groups

Twenty?two animals were randomly allocated into 4 exercise groups with 5 animals in each group, and 2 animals were allocated into an ”absolute” control group:

  • LEDT?Sham group: animals were treated with sham LEDT (LEDT device in placebo mode) over both legs, gluteus and lower?back muscles 5 minutes before each training session on ladder.

  • LEDT?Before: animals were treated with real LEDT over both legs, gluteus and lower?back muscles 5 minutes before each training session on ladder.

  • LEDT?Before?After: animals were treated with real LEDT over both legs, gluteus and lower?back muscles 5 minutes before and 5 minutes after each training session on ladder.

  • LEDT?After: animals were treated with real LEDT over both legs, gluteus and lower?back muscles 5 minutes after each training session on ladder.

  • Control: animals were not subjected to any LEDT or exercise or muscle performance assessment.

Ladder

An inclined ladder (80°) with dimensions of 100 cm × 9 cm (length and width, respectively) with bars spaced at 0.5 cm intervals was used in this study as reported in a previous study 33 (Figure 1).

Ladder. Inclined ladder (80°) with 100 cm × 9 cm (length and width, respectively) used for the training program and muscle performance assessments. Falcon tube filled with water and attached to the mouse tail.

Load

A Falcon tube (50 ml) was filled with measured volumes of water and weighed using a precise scale. The target load was achieved adding or removing water from the tube and then this tube was attached to the mouse tail using adhesive tape (Figure 1). All loads were calculated in grams.

Procedures

The schedule of the various exercise procedures is described in Table 1.

Table 1. Schedule for exercise procedures

Day

Procedure

# repetitions

Load

Day 1

Familiarization

4 × 10 = 40

zero

Day 2

3RM baseline

3

Starting at 2 × BWa

Day 3

Training 1

5 × 10 = 50

0.8 × 3RMb

Day 5

Training 2

5 × 10 = 50

0.9 × 3RM

Day 7

Training 3

5 × 10 = 50

1.0 × 3RM

Day 9

Training 4

5 × 10 = 50

1.1 × 3RM

Day 11

Training 5

5 × 10 = 50

1.2 × 3RM

Day 13

Training 6

5 × 10 = 50

1.3 × 3RM

Day 14

3RM final

3

Starting at 3 × BW

  • a : body weight
  • b : average load carried during 3RM baseline measurement

Familiarization with ladder?climbing

All experimental groups, except Control group, were familiarized with climbing the ladder one day before the start of muscle performance assessment and training. The familiarization procedure was 4 sets of 10 climbs on the ladder (repetitions) with rest periods of 2 minutes between individual sets. No load was attached to the mouse tail during this procedure.

Three repetitions maximum load (3RM)

This test was the first evaluation of muscle performance and was set as the average of the maximum load carried by each animal during 3 consecutive full climbs of the inclined ladder (3RM). Slight pressure with tweezers was applied on mouse tail if the animal stopped during a climb. The test was stopped when mice were not able to climb or lost their grip on the ladder due to failure of concentric muscle contraction. The first attempt included a load corresponding to 200% of the individual mouse body weight. A maximum of 3 climb attempts was applied. If a mouse finished the climb the load was increased by 10% for the next climb, while if the mouse failed to finish a climb, the load was decreased by 10% for the next climb. The 3RM evaluation was performed twice; the first time was 24 h after familiarization procedure (baseline) and the second time was 24 h after the last training session (final).

Acute strength training protocol

After 24 h from initial 3RM baseline assessment, all experimental groups, except Control, were subjected to 6 training sessions carried out on alternate days (every 48 h). Each training session consisted of 5 sets of 10 repetitions (climbs) on the ladder with a rest period of 2 minutes between each set. If the animal could not complete a set or failed during a climb, the distance climbed (in cm) was measured and the rest period was started immediately. During some repetitions, a slight pressure on the mouse tail was performed with tweezers to stimulate the animal to climb and complete the exercise. If after three applications of gentle pressures the mouse could not resume climbing, and stopped or lost its grip on the ladder, the set of repetitions was stopped and the rest interval was started.

The number of repetitions in each set was measured as well as the time spent to complete the exercise. These data were used to calculate the muscle work and muscle power in each training session. The load of each training session was progressively increased and calculated as percentages of the 3RM (in grams) measured at baseline as follows: first training (80%), second training (90%), third training (100%), fourth training (110%), fifth training (120%) and sixth training (130%).

Light?emitting diode therapy (LEDT)

A non?commercial cluster of 40 LEDs (20 red – 630 ± 10 nm; 20 infrared – 850 ± 20 nm) with diameter of 76 mm was used in this study. A complete description of the LEDT parameters is presented in Table 2. The optical power reaching the surface of the mouse skin was measured with an optical energy meter PM100D Thorlabs® fitted with a sensor S142C (area of 1.13 cm2). All mice (except mice in Control) were shaved and fixed on a plastic plate using adhesive tapes. Afterwards, in accordance with experimental group, these animals were treated with LEDT over both legs, gluteus and lower?back muscles at a distance of 45 mm (without contact) (Figure 2). Irradiation lasted 90 s per session with fixed parameters as described in Table 1. LEDT placebo had no energy (0 J) and no power (0 mW) applied over the targeted muscles. The light dose was based on the possible biphasic dose response reported previously 5, 6. Moreover, dual wavelengths were chosen to function at the same time in this study based on specificities of the chromophores in the cells and therefore optimizing the effects of the light therapy (LEDT) by a double band of absorption 8-11.

LEDT. Positioning of the mice and light?emitting diode therapy (LEDT) applied on mouse legs, gluteus and lower?back muscles without contact.

 

Muscle performance

The 3RM test was the first evaluation for muscle performance. This test measured the maximum load (in grams) carried by each animal during 3 consecutive full climbs on the inclined ladder.

During each training session the load, number of repetitions (rep), distance climbed and time spent to complete each repetition were recorded. These data were used to calculate muscle work and power.

Although the ladder had a total length of 100 cm available the maximum distance available to climb was set at 70 cm in order to avoid the load touching the floor. Thereby the muscle work was calculated as follows:

Work (J) = mgh

where ”m” is mass of the load (grams converted to kilogram) in each training session plus mouse body mass (values converted to kilogram); ”g” is acceleration due to gravity and ”h” is the distance climbed (converted to meters). Results were obtained in Joules (J) and presented as average ± standard error of mean (SEM) for each group at each training session.

Muscle power was calculated from results of muscle work (J) and time spent (s) to perform all repetitions of each set at all training sessions as follows:

Power (mW) = J/s

where ”J” is Joule and represents the muscle work performed and ”s” is time in seconds. Result were obtained in milliwatts (mW) and presented as average ± standard error of mean (SEM) per each group at each training session.

Muscular ATP

The gastrocnemius muscle from one leg of each animal was used for analysis of muscular ATP. Muscle samples were thawed in ice for 5 min, homogenized at a proportion of 3–4 mg of tissue to 500 µl of 10% perchloric acid (HClO4) following procedures previously published 34. Afterwards, an aliquot of 10 µl of the muscle homogenate plus 40 µl of CellTiter Glo Luminescent Cell Viability Assay mix (Promega), totaling 50 µl, were placed in the well microplate (CostarTM 96?Well White Clear?Bottom Plates). Luminescence signals were measured in a SpectraMax M5 Multi?Mode Microplate Reader (Molecular Devices, Sunnyvale, CA) with integration time of 5 s to increase low signals 34. A standard curve was prepared using ATP standard (Sigma) according to manufacturer's guidelines and then ATP concentration was calculated in nanomol (nmol) per milligram (mg) of protein. An aliquot of muscle homogenate was used to quantify the total protein by QuantiProTM BCA Assay kit (Sigma?Aldrich) following manufacturer's guidelines.

Muscular glycogen

Quadriceps femoris muscles were thawed in ice for 30 min and muscular glycogen was measured in 50 mg of quadriceps femoris tissue homogenized with 6 N NaOH at a proportion of 50 mg/ml. A standard curve was prepared using absolute ethanol (100%), K2SO4 (10%), phenol (4.1%) and 1 mM of glucose (2%) according to Dubois et al. 35. Optical density was read at 480 nm in spectrophotometer (EvolutionTM 300 UV?Vis, software VISPRO – Thermo Scientific). Data were normalized per mg of muscle tissue.

Oxidative stress markers

Protein carbonyl: Quadriceps femoris muscles were homogenized in deionized water (dH2O) at a proportion of 10 mg/200 µl. Protein carbonyl content was quantified using Protein Carbonyl Content Assay kit (Biovision) with the colorimetric method and following manufacturer's guidelines. All results were normalized per total protein quantified by QuantiProTM BCA Assay kit (Sigma?Aldrich) following manufacturer's guidelines.

Glutathione: Quadriceps femoris muscles were homogenized in 100 mM ice cold phosphate buffer (pH = 7.4) at a proportion of 10 mg/250 µl. Phosphate buffer was prepared with dibasic (Na2HPO4) and monobasic (NaH2PO4) sodium phosphate at equal proportions. Total and oxidized glutathione analysis was carried out with Glutathione Colorimetric Assay kit (ARBOR Assays) following manufacturer's guidelines. In addition, all results were normalized per total protein of the samples using QuantiProTM BCA Assay kit (Sigma?Aldrich) following manufacturer's guidelines.

Catalase activity: Quadriceps femoris muscles were homogenized in cold assay buffer provided in a Catalase Activity Assay kit (Biovision) at a proportion of 50 mg/100 µl. This analysis used the colorimetric method and followed manufacture's guidelines.

Lipid peroxidation using TBARS (Thiobarbituric Acid Reactive Substances): Quadriceps femoris muscles were homogenized with RIPA Buffer (Sigma?Aldrich) at a proportion of 25 mg/250 µl. Next, TBARS Colorimetric Assay kit (Cayman Chemical) was used following manufacturer's guidelines.

Protein Thiols: Quadriceps femoris muscles were homogenized in ice cold 100 mM phosphate buffer at a proportion of 10 mg/250 µl. Next, a Fluorescent Protein Thiol Detectiont kit (ARBOR Assays) was used following manufacturer's guidelines. In addition, all results were normalized per total protein quantified by QuantiProTM BCA Assay kit (Sigma?Aldrich) following manufacturer's guidelines.

Immunofluorescence analyses

5?bromo?2′?deoxyuridine (BrdU): BrdU reagent (Sigma?Aldrich) was diluted in saline solution (PBS) at a concentration of 10 mg/ml. Next, during the last 8 days of the experiment all animals (including Control group) received a single daily intra peritoneal injection (50 mg/kg) of BrdU. Mice were anesthetized and submitted to surgical procedures described previously. Gastrocnemius muscles were embedded in paraffin, cut in axial slices of 5 µm thickness from the muscle belly region by a microtome and mounted on slides for immunohistochemical procedures. Briefly, slides were deparaffinized with graded ethanol and then passed through antigen retrieval solution in a water bath pre?heated at 98 °C for 30 min. Afterwards slides were washed and incubated for 15 min at room temperature with 0.1% Triton X?100 TBS for cell membrane permeabilization, washed again and incubated for 30 min in protein blocking solution consisting of 3% BSA (Bovine Serum Albumin – Sigma) and 10% goat serum in TBS. Next, slides were immunostained with sheep anti?BrdU (Ab1893 – Abcam, Cambridge, MA) at 1 : 50 working concentration and selected anti?sheep (Alexa Fluor® 647 – Invitrogen) fluorescent secondary antibody matched to the primary antibody to stain at 1 : 200 working concentration. Finally, slides were cover?slipped with mounting media containing DAPI (4′,6?diamidino?2?phenylindole) (Invitrogen). Cells positively stained for BrdU were imaged using confocal microscope (Olympus America Inc. Center Valley, PA, USA) from three random fields. BrdU and DAPI staining were quantified using software Image J (NIH, Bethesda, MD).

Cytochrome c oxidase subunit IV (COX IV): Gastrocnemius muscles were subjected to the same procedures described for BrdU staining. Slides were immunostained with rabbit anti?COX IV (Cell Signaling Technology®) at 1 : 500 working concentration and selected anti?rabbit (Alexa Fluor® 680 – Invitrogen) secondary antibody matched with primary antibody to stain at 1 : 200 working concentration. Cells positively stained for COX IV were imaged using confocal microscopy as above and then the red channel of the exported images was changed to yellow.

Statistical analysis

Shapiro?Wilk's W test verified the normal distribution of the data. All experimental groups subjected to training protocols were compared at each training session for number of repetitions, muscle work and muscle power using one?way analysis of variance (ANOVA) and Tukey HSD post?hoc test. The load of 3RM among these same groups was compared by Two?way ANOVA with repeated measures (baseline versus final) and Tukey HSD post?hoc test. For muscular ATP, glycogen, oxidative stress markers and immunofluorescence stains, all experimental groups were compared by one?way ANOVA and Tukey's HSD post?hoc test. Significance was set at p < 0.05.

 

Results

Muscle performance

3RM: The final load 3RM was significantly higher (p < 0.05) in all experimental groups at the end of the experiment period compared to baseline. The final load of LEDT?After (92.28 g, SEM 0.82) was higher than LEDT?Sham (59.58 g, SEM 5.28; p < 0.001) and LEDT?Before (78.98 g, SEM 1.96; p = 0.020). In addition, LEDT?Sham had a significantly lower final load (p < 0.001) compared to LEDT?Before as well as LEDT?Before/After (83.91 g, SEM 1.49) (Figure 4A).

Muscle performance (n = 5 animals per group). (A) Baseline and Final test of 3 repetitions maximum (3RM) measuring the total load carried by mice during this test. * statistical significance (p < 0.05) comparing the final 3RM load between groups. (B) Number of repetitions or climbs performed by each group treated with different regimens of LEDT during the progressive training program. (C) Muscle power developed by each group treated with different regimens of LEDT during the progressive training program. (D) Muscle work developed by each group treated with different regimens of LEDT during the progressive training program. * statistical significance (p < 0.05) compared to LEDT?Sham. # statistical significance (p < 0.05) compared to LEDT?After. & statistical significance (p < 0.05) compared to LEDT?Before. Abbreviations: LEDT = light?emitting diode therapy; LEDT?Sham (Sham – S) = LEDT placebo (LEDT device in placebo mode) on muscles immediately before (5 minutes) each training session on ladder; LEDT?Before (Before – B) = LEDT applied on muscles immediately before (5 minutes) each training session on ladder; LEDT?Before?After (Before?After – A?B) = LEDT applied on muscles immediately before (5 minutes) and immediately after (5 minutes) each training session on ladder; LEDT?After (After – A) = LEDT applied on muscles immediately after (5 minutes) each training session on ladder. The load of 3RM at baseline versus final was analyzed by Two?way analysis of variance (ANOVA) with repeated measures. Number of repetitions, muscle work and power were analyzed by One?way ANOVA.

Number of repetitions: There were significantly differences (p < 0.05) between all groups in each training session (Figure 4B). At 80% of 3RM (first session): animals in LEDT?Before and LEDT?Before?After groups performed more repetitions compared to animals in LEDT?Sham and LEDT?After (p < 0.01) groups. At 90% of 3RM (second session): animals in LED?Sham group performed fewer repetitions than animals in LEDT?Before, LEDT?Before?After and LEDT?After groups (p < 0.001). At 100% of 3RM (third session): animals in LEDT?Sham group performed fewer repetitions compared to animals in LEDT?Before (p = 0.014), LED?Before?After (p = 0.010) and LEDT?After (p = 0.002) groups. At 110% of 3RM (fourth session): animals in LEDT?Sham group performed fewer repetitions than animals in LEDT?Before?After (p = 0.013) and LEDT?After (p = 0.009) groups. At 120% of 3RM (fifth session): animals in LEDT?After group performed more repetitions than animals in LEDT?Before (p = 0.022) and LEDT?Sham (p < 0.001) groups. In addition, animals in LEDT?Sham performed fewer repetitions than animals in LEDT?Before (p = 0.022), LEDT?Before?After and LEDT?After (p < 0.001) groups. At 130% of 3RM (sixth session): animals in LEDT?Before?After and LEDT?After groups performed more repetitions than animals in LEDT?Sham (p < 0.001) and LEDT?Before (p < 0.01) groups.

Muscle Power: At 80% of 3RM there were no significant differences among all groups (p > 0.05). At 90% of 3RM: animals in LEDT?Sham group had lower muscle power compared to animals in LEDT?Before, LEDT?Before?After and LEDT?After (p < 0.01) groups. At 100% of 3RM: animals in LEDT?Sham group had lower muscle power than animals in LEDT?Before?After (p = 0.025) and LEDT?After (p = 0.007) groups. At 110% of 3RM: animals in LEDT?Before?After group developed more muscle power than animals in LEDT?Sham (p < 0.001) and LEDT?Before (p = 0.013) groups. In addition, animals in LEDT?After group had more muscle power than animals in LEDT?Sham (p = 0.002) group. At 120% of 3RM: animals in LEDT?Before?After and LEDT?After groups developed more muscle power than animals in LEDT?Sham and LEDT?Before (p < 0.001) groups. At 130% of 3RM: animals in LEDT?Before?After group developed more muscle power than animals in LEDT?Sham and LEDT?Before (p < 0.001) as well as LEDT?After (p = 0.001) groups. In addition, animals in LEDT?After group had more muscle power than animals in LEDT?Sham (p < 0.001) and LEDT?Before (p = 0.004) groups. Finally, animals in LEDT?Before group had major muscle power than animals in LEDT?Sham (p = 0.020) group (Figure 4C).

Muscle Work: Similar to results presented in Figure 4B, at 80% of 3RM only animals in LEDT?Before and LEDT?Before?After groups performed more muscle work compared to LEDT?Sham (p < 0.05) group (Figure 4D). At 90% of 3RM: animals in LEDT?Sham group performed less muscle work than animals in LEDT?Before, LEDT?Before?After and LEDT?After (p < 0.001) groups. These results were similar at 100% of 3RM (p < 0.001). At 110% of 3RM: animals in LEDT?Sham group had lower muscle work compared to animals in LEDT?Before?After (p = 0.015) and LEDT?After (p = 0.011) groups. At 120% of 3RM: animals in LEDT?Sham group performed lower muscle work compared to animals in LEDT?Before (p = 0.027) and LEDT?Before?After and LEDT?After (p < 0.001) groups. In addition, animals in LEDT?After group performed more muscle work than animals in LEDT?Before (p = 0.026) group. At 130% of 3RM: animals in LEDT?Before?After and LEDT?After groups performed more muscle work than animals in LEDT?Sham (p < 0.001) and LEDT?Before (p < 0.01) groups (Figure 4D).

Muscle ATP content

Animals in LEDT?After group had significantly (p < 0.001) more ATP concentration (1,367.64 nmol/ mg protein, SEM 105.30) compared to animals in LEDT?Sham (15.85 nmol/mg protein, SEM 5.14), LEDT?Before (81.00 nmol/ mg protein, SEM 10.11), LEDT?Before?After (687.62 nmol/ mg protein, SEM 11.76) and Control (17.53 nmol/mg protein, SEM 7.47) groups. In addition, animals in LEDT?Before?After group had also major contents of ATP compared to animals in LEDT?Before, LEDT?Sham and Control (p < 0.001) groups (Figure 5A).

 

 

Muscular ATP and glycogen contents (n = 5 animals per group). (A) Adenosine triphosphate (ATP) contents in gastrocnemius muscle after the training program. (B) Glycogen contents in quadriceps femoris muscles after the training program. * statistical significance (p < 0.05). Abbreviations: LEDT = light?emitting diode therapy; LEDT?Sham (Sham – S) = LEDT placebo (LEDT device in placebo mode) on muscles immediately before (5 minutes) each training session on ladder; LEDT?Before (Before – B) = LEDT applied on muscles immediately before (5 minutes) each training session on ladder; LEDT?Before?After (Before?After – A?B) = LEDT applied on muscles immediately before (5 minutes) and immediately after (5 minutes) each training session on ladder; LEDT?After (After – A) = LEDT applied on muscles immediately after (5 minutes) each training session on ladder. Control (C) = no exercise or muscle performance assessment. Comparisons among all groups were conducted using One?way analysis of variance (ANOVA).

 

 

Muscle glycogen content

Animals in LEDT?After (137.76 nmol/mg tissue, SEM 11.40) and LEDT?Before?After (144.44 nmol/ mg tissue, SEM 16.23) groups had significantly higher concentrations of glycogen in quadriceps femoris muscles (p < 0.001) compared to animals in LEDT?Sham (31.36 nmol/mg tissue, SEM 7.45), LEDT?Before (52.76 nmol/mg tissue, SEM 6.53) and Control (58.78 nmol/ mg tissue, SEM 7.17) groups (Figure 5B).

Oxidative stress markers

Total glutathione: Animals in Control group (1.33 µM/µg protein, SEM 0.11) had a significantly higher concentration of total glutathione compared to animals in LEDT?Sham (0.097 µM/µg protein, SEM 0.046; p = 0.005) and LEDT?Before (1.00 µM/µg protein, SEM 0.02; p = 0.010) groups (Figure 6A).

Oxidative stress markers (n = 5 animals per group) in quadriceps femoris muscles. (A) Total Glutathione (reduced glutathione – GSH). (B) Oxidized Glutathione (GSSG). (C) Protein Carbonyl. (D) Catalase activity. (E) Lipid peroxidation using TBARS (Thiobarbituric Acid Reactive Substances). (F) Protein Thiol. * statistical significance (p < 0.05). Abbreviations: LEDT = light?emitting diode therapy; LEDT?Sham (Sham – S) = LEDT placebo (LEDT device in placebo mode) on muscles immediately before (5 minutes) each training session on ladder; LEDT?Before (Before – B) = LEDT applied on muscles immediately before (5 minutes) each training session on ladder; LEDT?Before?After (Before?After – A?B) = LEDT applied on muscles immediately before (5 minutes) and immediately after (5 minutes) each training session on ladder; LEDT?After (After – A) = LEDT applied on muscles immediately after (5 minutes) each training session on ladder. Control (C) = no exercise or muscle performance assessment. Comparisons among all groups were conducted using One?way analysis of variance (ANOVA).

Oxidized glutathione: Animals in LEDT?Sham group (0.005 µM/µg protein, SEM 0.001) had significantly minor concentration of glutathione oxidized compared to animals in LEDT?Before (0.20 µM/µg protein, SEM 0.002; p = 0.015), LEDT?Before?After (0.035 µM/µg protein, SEM 0.003; p < 0.001), LEDT?After (0.041 µM/µg protein, SEM 0.003; p < 0.001) and Control (0.027 µM/µg protein, SEM 0.007; p = 0.006) groups. In addition, animals in LEDT?Before group had significantly minor concentration of oxidized glutathione compared to animals in LEDT?After (p < 0.001) and LEDT?Before?After (p = 0.024) groups (Figure 6B).

Protein carbonyl: Animals in LEDT?After group (1.40 nmol/µg protein, SEM 0.15) had significantly lower concentrations of protein carbonyls compared to animals in LEDT?Sham (6.31 nmol/µg protein, SEM 1.09; p = 0.030), LEDT?Before (6.81 nmol/µg protein, SEM 1.21; p = 0.040) and LEDT?Before?After (8.27 nmol/µg protein, SEM 2.35; p = 0.008) groups (Figure 6C).

Catalase activity: Animals in LEDT?Sham group (2.11 nmol/min/ml, SEM 0.10) had significantly lower catalase activity (p < 0.01) compared to animals in LEDT?Before?After (4.33 nmol/min/ml, SEM 0.62), LEDT?After (4.22 nmol/min/ml, SEM 0.37) and Control (4.47 nmol/min/ml, SEM 0.52) groups (Figure 6D).

Lipid peroxidation using TBARS: There were no significant differences between any of the groups (p > 0.05) assessed. Animals in Control group had a concentration of 21.29 µM (SEM 1.13); animals in LEDT?Sham had 21.12 µM (SEM 2.86); animals in LEDT?Before had 23.87 µM (SEM 1.13); animals in LEDT?Before?After had 19.19 µM (SEM 1.01) and animals in LEDT?After had 19.55 µM (SEM 1.24) (Figure 6E).

Protein
Original Source: https://onlinelibrary-wiley-com.colorado.idm.oclc.org/doi/full/10.1002/jbio.201400087


Effect of near-infrared light-emitting diodes on nerve regeneration.

Ishiguro M, Ikeda K, Tomita K - J Orthop Sci. 2010 Mar (Publication)
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Background: Photobiomodulation by red to near-infrared light-emitting diodes (LEDs) has been reported to accelerate wound healing, attenuate degeneration of an injured optic nerve, and promote tissue growth. The purpose of this study was to investigate the effect of LEDs on nerve regeneration. A histological study as well as a measurement of antioxidation levels in the nerve regeneration chamber fluid was performed.

Methods: For the histological study, the bilateral sciatic nerves were transected, and the left proximal stump and the right distal stump were inserted into the opposite ends of a silicone chamber, leaving a 10-mm gap. Light from an LED device (660 nm, 7.5 mW/cm2) was irradiated for 1 hr per day. At 3 weeks after surgery, regenerated tissue was fixed and examined by light microscopy. For the antioxidation assay of chamber fluid, the left sciatic nerve and a 2-mm piece of nerve from the proximal stump were transected and inserted into opposite sides of a silicone chamber leaving a 10-mm gap. LEDs were irradiated using the same parameters as those described in the histological study. At 1, 3, and 7 days after surgery, antioxidation of the chamber fluid was measured using an OXY absorbent test.

Results: Nerve regeneration was promoted in the LED group. Antioxidation of the chamber fluid significantly decreased from 3 days to 7 days in the control group. In the LED group, antioxidation levels did not decrease until 7 days.

Conclusions: Chamber fluid is produced from nerve stumps after nerve injury. This fluid contains neurotrophic factors that may accelerate axonal growth. Red to near-infrared LEDs have been shown to promote mitochondrial oxidative metabolism. In this study, LED irradiation improved nerve regeneration and increased antioxidation levels in the chamber fluid. Therefore, we propose that antioxidation induced by LEDs may be conducive to nerve regeneration.

Original Source: https://www.ncbi.nlm.nih.gov/pubmed/20358337

A NASA discovery has current applications in orthopaedics

Howard B. Cotler, MD, FACS, FAAOS - Curr Orthop Pract. 2015 Jan; 26(1): 72–74 (Publication)
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Low-level laser therapy (LLLT) has been actively used for nearly 40 yr, during which time it has been known to reduce pain, inflammation, and edema. It also has the ability to promote healing of wounds, including deep tissues and nerves, and prevent tissue damage through cell death. Much of the landmark research was done by the National Aeronautics and Space Administration (NASA), and these studies provided a springboard for many additional basic science studies. Few current clinical studies in orthopaedics have been performed, yet only in the past few years have basic science studies outlined the mechanisms of the effect of LLLT on the cell and subsequently the organism. This article reviews the basic science of LLLT, gives a historical perspective, and explains how it works, exposes the controversies and complications, and shows the new immediately applicable information in orthopaedics.

Key Words: Laser, LED, NASA, orthopaedic, injury

BACKGROUND

The pursuit of space travel has opened new areas for study and knowledge. Space medicine has had applications in various subspecialties. Although some think there is little application in orthopaedics, it may be that there has been much discovered but little appreciated. The National Aeronautics and Space Administration (NASA) was established by the United States government in 1958 as a civilian space program for aeronautics and aerospace research. In 1959 the Astronaut Corps was founded. The insertion of humans into space presented many challenges from a biologic standpoint. Astronauts in space perform physically demanding work in a challenging environment that includes among other hazards, microgravity, which is known to have an adverse effect on bone and muscle to the extent that it places an increased risk for musculoskeletal injury. There is a threefold higher injury rate during mission periods than outside of mission periods for astronauts, and it has been observed that wounds heal more slowly in orbit.

In 1993, Quantum Devices (Barneveld, WI) developed a light-emitting diode (LED) for NASA to use in their plant growth experiments. The experiments demonstrated that red LED wavelengths could boost plant growth, but coincidentally the scientist’s skin lesions began to heal faster as well. NASA subsequently began to study the use of LED to increase the metabolism of human cells and stem the loss of bone and muscle in astronauts.

Dr. Harry T. Whelan, a professor of pediatric neurology at the University of Wisconsin, began the study of LEDs and lasers, receiving grants from NASA and and the National Institutes of Health. He determined that astronauts get four problems: immune deficiency, pituitary insufficiency, delayed wound healing, and muscle and bone atrophy. He observed these results in the laboratory.

MECHANISMS OF ACTION

From a historical perspective we now know that light has a biologic effect, but what we need to know is how energy from lasers and LEDs work on a cellular level and what the optimal light parameters are for different uses.

The power plant of cells is located in the mitochondria that are able to produce cellular energy or adenosine triphosphate (ATP) from pyruvate and oxygen. When tissues are stressed or ischemic, mitochondria make their own mitochondrial nitric acid (MtNO), which competes with oxygen. The MtNO bind to cytochrome C oxidase (CcO) that displaces oxygen. This subsequently reduces ATP synthesis and increased oxidative stress, which leads to inflammation. Hypoxic or stressed tissues are affected by LLLT in four stages: (1) light energy is absorbed by cytochrome C oxidase, triggering several downstream effects; (2) nitric oxide is released; (3) ATP is increased; and (4) oxidative stress is reduced. These biochemical intermediates affect components in the cytosol, cell membrane, and nucleus that control gene transcription, cell proliferation, migration necrosis, and inflammation. Cells in blood and lymph, which have been light activated, can travel a distance for systemic effects.,

APPLICATIONS

The four common targets for LLLT are:

  • L – lymph nodes to reduce edema and inflammation.
  • I – site of injury to promote healing and reduce inflammation.,
  • N – nerves to induce analgesia.
  • T – trigger points to reduce muscle spasms.,

LLLT is a transcutaneous procedure with no invasive portion. The physician determines the correct synchronizations of continuous or pulsed laser emission. Penetration depth is determined by wavelength and power. The U.S. Navy research determined 810 nm to be optimal for penetration. Treatment times are in the range of 30 s to 1 min, but there are many areas treated for comprehensible protocol, which often takes approximately 30 min to perform. For stimulating repair and decreasing inflammation, 2.5 Hz pulse is recommended, while a continuous beam is ideal for analgesia and tender points.

ADMINISTRATION

The Federal Drug Administration (FDA) approved the use of LLLT in 2003. In some states, a prescription is mandatory before treatment. Treatment can be administered by a certified therapist, radiology technologist, or a physician. European sports therapists have used LLLT for over a decade; however, they report only a 50% success rate,, which may be due to inconsistent laser parameters and dose. Recent advances by researchers at Harvard Medical School have clarified the mechanism by which there is biphasic dose response.,

Side effects and complications can result from traditional treatments for musculoskeletal pathology. Nonsteroidal antiinflammatories can cause ulcer disease, hypertension, bleeding, and cardiac events. Steroids (oral and/or epidural) can result in infections (including epidural), bleeding, ulcers, avascular necrosis, and tissue fragility. Studies have found LLLT to have no side effects or adverse events beyond those reported for placebo.

With over 4000 basic science research and clinical studies according to pubmed.gov, and low complication rate, LLLT should be considered as a first-line treatment option for conditions such as acute neck or back pain, tendinitis, plantar fasciitis, mild carpal tunnel sndrome, and ligamentous sprains. Its safety profile provides a persuasive argument, with the added benefits of accelerated healing, tissue remodeling, pain relief, and decreased inflammation. LLLT subsequently has been accepted by both the British and Canadian health services. Although approved by the FDA, LLLT has not been recognized or accepted by Medicare or insurance companies because it is viewed as investigational treatment.

Clinical practice guidelines of the American Academy of Orthopaedic Surgeons (AAOS) in 2008 on treatment of carpal tunnel syndrome included laser treatment but carried no recommendations for or against its use because there is insufficient evidence. The literature on LLLT for the treatment of lymphedema, wound healing, prevention of oral mucositis, or for pain demonstrates inconsistent results and methodological weaknesses as per the Blue Cross Blue Shield of Kansas Medical Policy, March 12, 2013. More up-to-date, prospective studies, using newer treatment guidelines by clinicians, are needed to provide a complete picture of efficacy and cost-effectiveness.

CONCLUSION

LLLT will not replace orthopaedic surgery for structural pathology, but it may be useful as an adjunct therapy for patients seeking noninvasive symptomatic treatment or accelerated wound healing.

Footnotes

Financial Disclosure: Dr. Cotler is in private practice and owns Gulf Coast Spine Care Ltd., PA and Laser Health Spa, LLC. He received no financial suport for this manuscript.

REFERENCES

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Original Source: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4272231/

Comparison between cold water immersion therapy (CWIT) and light emitting diode therapy (LEDT) in short-term skeletal muscle recovery after high-intensity exercise in athletes--preliminary results.

Leal Junior EC1, de Godoi V, Mancalossi JL, Rossi RP, De Marchi T, Parente M, Grosselli D, Generosi RA, Basso M, Frigo L, Tomazoni SS, Bjordal JM, Lopes-Martins RA. - Lasers Med Sci. 2011 Jul;26(4):493-501. doi: 10.1007/s10103-010-0866-x. Epub 2010 Nov 19. (Publication)
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Intro: In the last years, phototherapy has becoming a promising tool to improve skeletal muscle recovery after exercise, however, it was not compared with other modalities commonly used with this aim. In the present study we compared the short-term effects of cold water immersion therapy (CWIT) and light emitting diode therapy (LEDT) with placebo LEDT on biochemical markers related to skeletal muscle recovery after high-intensity exercise. A randomized double-blind placebo-controlled crossover trial was performed with six male young futsal athletes. They were treated with CWIT (5°C of temperature [SD ±1°]), active LEDT (69 LEDs with wavelengths 660/850 nm, 10/30 mW of output power, 30 s of irradiation time per point, and 41.7 J of total energy irradiated per point, total of ten points irradiated) or an identical placebo LEDT 5 min after each of three Wingate cycle tests. Pre-exercise, post-exercise, and post-treatment measurements were taken of blood lactate levels, creatine kinase (CK) activity, and C-reactive protein (CRP) levels. There were no significant differences in the work performed during the three Wingate tests (p > 0.05). All biochemical parameters increased from baseline values (p < 0.05) after the three exercise tests, but only active LEDT decreased blood lactate levels (p = 0.0065) and CK activity (p = 0.0044) significantly after treatment. There were no significant differences in CRP values after treatments. We concluded that treating the leg muscles with LEDT 5 min after the Wingate cycle test seemed to inhibit the expected post-exercise increase in blood lactate levels and CK activity. This suggests that LEDT has better potential than 5 min of CWIT for improving short-term post-exercise recovery.

Background: In the last years, phototherapy has becoming a promising tool to improve skeletal muscle recovery after exercise, however, it was not compared with other modalities commonly used with this aim. In the present study we compared the short-term effects of cold water immersion therapy (CWIT) and light emitting diode therapy (LEDT) with placebo LEDT on biochemical markers related to skeletal muscle recovery after high-intensity exercise. A randomized double-blind placebo-controlled crossover trial was performed with six male young futsal athletes. They were treated with CWIT (5°C of temperature [SD ±1°]), active LEDT (69 LEDs with wavelengths 660/850 nm, 10/30 mW of output power, 30 s of irradiation time per point, and 41.7 J of total energy irradiated per point, total of ten points irradiated) or an identical placebo LEDT 5 min after each of three Wingate cycle tests. Pre-exercise, post-exercise, and post-treatment measurements were taken of blood lactate levels, creatine kinase (CK) activity, and C-reactive protein (CRP) levels. There were no significant differences in the work performed during the three Wingate tests (p > 0.05). All biochemical parameters increased from baseline values (p < 0.05) after the three exercise tests, but only active LEDT decreased blood lactate levels (p = 0.0065) and CK activity (p = 0.0044) significantly after treatment. There were no significant differences in CRP values after treatments. We concluded that treating the leg muscles with LEDT 5 min after the Wingate cycle test seemed to inhibit the expected post-exercise increase in blood lactate levels and CK activity. This suggests that LEDT has better potential than 5 min of CWIT for improving short-term post-exercise recovery.

Abstract: Abstract In the last years, phototherapy has becoming a promising tool to improve skeletal muscle recovery after exercise, however, it was not compared with other modalities commonly used with this aim. In the present study we compared the short-term effects of cold water immersion therapy (CWIT) and light emitting diode therapy (LEDT) with placebo LEDT on biochemical markers related to skeletal muscle recovery after high-intensity exercise. A randomized double-blind placebo-controlled crossover trial was performed with six male young futsal athletes. They were treated with CWIT (5°C of temperature [SD ±1°]), active LEDT (69 LEDs with wavelengths 660/850 nm, 10/30 mW of output power, 30 s of irradiation time per point, and 41.7 J of total energy irradiated per point, total of ten points irradiated) or an identical placebo LEDT 5 min after each of three Wingate cycle tests. Pre-exercise, post-exercise, and post-treatment measurements were taken of blood lactate levels, creatine kinase (CK) activity, and C-reactive protein (CRP) levels. There were no significant differences in the work performed during the three Wingate tests (p > 0.05). All biochemical parameters increased from baseline values (p < 0.05) after the three exercise tests, but only active LEDT decreased blood lactate levels (p = 0.0065) and CK activity (p = 0.0044) significantly after treatment. There were no significant differences in CRP values after treatments. We concluded that treating the leg muscles with LEDT 5 min after the Wingate cycle test seemed to inhibit the expected post-exercise increase in blood lactate levels and CK activity. This suggests that LEDT has better potential than 5 min of CWIT for improving short-term post-exercise recovery.

Original Source: http://www.ncbi.nlm.nih.gov/pubmed/21088862

Second International Conference on Near-Field Optical Analysis: Photodynamic Therapy & Photobiology Effect

- Proceedings of the Second International Conference on NOA : May 31 - June 1,2001 (Publication)
This extensive 105 page document showed the ability of the LED to speed the rate of healing, we hypothesized that using the LED for wounds aboard the submarine would increase the rate of healing.
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Please click on the original URL to see the complete study.


Original Source: https://ntrs.nasa.gov/archive/nasa/casi.ntrs.nasa.gov/20030001592.pdf

Is light-emitting diode phototherapy (LED-LLLT) really effective?

Won-Serk Kim1 and R Glen Calderhead2 - Laser Ther. 2011; 20(3): 205–215. (Publication)
This summary publication shows LED phototherapy is proving to have more and more viable applications in many fields of medicine.
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Background: Low level light therapy (LLLT) has attracted attention in many clinical fields with a new generation of light-emitting diodes (LEDs) which can irradiate large targets. To pain control, the first main application of LLLT, have been added LED-LLLT in the accelerated healing of wounds, both traumatic and iatrogenic, inflammatory acne and the patient-driven application of skin rejuvenation.

Rationale and Applications: The rationale behind LED-LLLT is underpinned by the reported efficacy of LED-LLLT at a cellular and subcellular level, particularly for the 633 nm and 830 nm wavelengths, and evidence for this is presented. Improved blood flow and neovascularization are associated with 830 nm. A large variety of cytokines, chemokines and macromolecules can be induced by LED phototherapy. Among the clinical applications, non-healing wounds can be healed through restoring the collagenesis/collagenase imbalance in such examples, and ‘normal’ wounds heal faster and better. Pain, including postoperative pain, postoperative edema and many types of inflammation can be significantly reduced.

Experimental and clinical evidence: Some personal examples of evidence are offered by the first author, including controlled animal models demonstrating the systemic effect of 830 nm LED-LLLT on wound healing and on induced inflammation. Human patients are presented to illustrate the efficacy of LED phototherapy on treatment-resistant inflammatory disorders.

Conclusions: Provided an LED phototherapy system has the correct wavelength for the target cells, delivers an appropriate power density and an adequate energy density, then it will be at least partly, if not significantly, effective. The use of LED-LLLT as an adjunct to conventional surgical or nonsurgical indications is an even more exciting prospect. LED-LLLT is here to stay.

Keywords: Grotthus-Draper law, nonhealing wound, photochemical cascade, photophysical reaction, irritant contact dermatitis, dissecting cellulitis, acne rosacea

INTRODUCTION

High level laser treatment (HLLT) means that high levels of incident laser power are used to deliberately destroy a specific target through a light-heat transduction process to induce photothermal damage of varying degrees. HLLT is used in many surgical fields, but probably most commonly in dermatologic, aesthetic or plastic surgery. On the other hand, when a laser or other appropriate light source is used on tissue at low incident levels of photon energy, none of that energy is lost as heat but instead the energy from the absorbed photons is transferred directly to the absorbing cell or chromophore, causing photoactivation of the target cells and some kind of change in their associated activity. In clinical applications, this was termed ‘low level laser therapy’ (LLLT) by Ohshiro and Calderhead in 1988,) with ‘photobiomodulation’ or ‘photoactivation’ referring to the activity at a cellular and molecular level.

Genesis of LLLT

In the late 1960's, the early days of the clinical application of the laser, there was fear that laser energy could induce carcinogenesis as a side effect of the use of the laser in surgery and medicine. To assess this, in a paper published in 1968, the late Professor Endrè Mester, the recognized father of phototherapy from Semmelweis University, Budapest, applied daily doses of low incident levels of defocused ruby laser energy to the shaved dorsum of rats.) No carcinogenetic changes were noted at all, but Mester incidentally discovered that LLLT accelerated hair regrowth in the laser-irradiated animals. Furthermore, during this period, early adopters of the surgical laser were reporting interesting and beneficial effects of using the laser as a scalpel compared with the conventional cold steel instrument, such as reduced inflammation, less postoperative pain, and better wound healing. Mester's experiments helped to show that it was the ‘L’ of laser, namely light, that was associated with these effects due to the bioactivative levels of light energy which exist simultaneously at the periphery of the photosurgical destructive zone, as illustrated in Figure 1.

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Range of typical bioreactions associated with a surgical laser and their approximate temperature range. Note that some degree of photoactivation almost always occurs simultaneously with HLLT-mediated reactions. (Data adapted from Calderhead RG: Light/tissue interaction in photosurgery and phototherapy. In Calderhead RG. Photobiological Basics of Photosurgery and Phototherapy, 2011, Hanmi Medical Publishers, Seoul. pp 47–89)

In the 1970's, many clinicians, inspired by Mester's major publication in 1969 on the significantly successful use of LLLT for the treatment of nonhealing or torpid crural ulcers, started to apply LLLT clinically, particularly in France and Russia, and this spread to Japan, Korea, and other Asian countries in the early 1980's. However, it was still looked on as ‘black magic’ by the mainstream medicoscientific world in the USA. The first Food and Drug Administration (FDA) approval for laser diode phototherapy was not granted till 2002, but even then the sceptics were not silenced.

LLLT with Lasers

LLLT was first completely limited to treatment with laser sources, such as the helium neon (HeNe) laser in the visible red at 632.8 nm, various semiconductor (diode) lasers (visible red to near infrared, most notable being the GaAlAs at 830 nm) or defocused beams of a surgical laser (Nd:YAG or CO2, for example).) There are several mechanisms which have been reported as to how LLLT can induce a biomodulative effect (Table 1). In the case of LLLT with laser sources, these effects were achieved athermally and atraumatically through the special properties associated with the ‘coherence’ of laser energy, namely monochromaticity, directionality or collimation, and the photons all in phase temporally and spatially. Another phenomenon associated only with laser energy is the so-called ‘speckle’ phenomenon. When the spot from a 670 nm laser pointer is closely examined over a period of time, for example, it appears to be composed of exceptionally brighter spots of light energy which are constantly in motion: these are laser speckles. Speckles have their own characteristics, including high energy and polarization, and these intense spots of polarized light were associated with specific reactions in the absorbing target or chromophore.

Table 1:

Major mechanisms associated with photobioactivation and LLLT
Mild thermal (<40°C) Biochemical Bioelectric Bioenergetic
↑ Nerve conduction (Mitochondrial events) ↑ Electromotive action on membrane bound ion transport mechanisms ↑ Rotational & vibrational changes to membrane molecule electrons
  ↑ ATP production    
  ↑ Release of nitric oxide (NO)    
  ↑ Very low levels of reactive oxygen species (ROS)    

↑ Capillary dilatation ↑ Fibroblast proliferation → Collagen & elastin synthesis ↑ Intracellular extra-cellular ion gradient changes ↑ Stimulation of acupuncture meridian points

  ↑ Mast cell degranulation: cytokine, chemokine and trophic factor release ↑ Depolarization of synaptic cleft → closure of synaptic gate ↑ Increased biophotonic activity

  ↑ Macrophage activity (chemotaxis & internalization) → release of FGF ↑ Activation of the dorsal horn gate control mechanism → pain transmission slowed, pain control increased  

  ↑ Keratinocyte activity cytokine release in epidermis and dermis      

  ↑ Opiate and nonopiate pain control (endorphins, dynorphins and enkephalins)      

  ↑ RNA/DNA synthesis      

  ↑ Enzyme production      

  ↑ Superoxide dismutase (SOD) production      

Up until the end of the 1990's, phototherapy was dominated by these laser sources, because although LEDs were cheap and cheerful, they were highly divergent with low and unstable output powers, and a wide waveband. With very few exceptions, old generation LEDs were incapable of producing really useful clinical reactions in tissue. It was easy to source a ‘red’ LED (output spread over approximately 600 – 700 nm) but it was more or less impossible to source LEDs at specific nominal wavelengths, for example 633 nm, similar to the HeNe laser.

LED PHOTOTHERAPY

Enter the NASA Light-Emitting Diode (LED)

All this changed in 1998 with the development of the so-called ‘NASA LED’ by Prof Harry Whelan and his group at the NASA Space Medicine Laboratory, which offered clinicians and researchers a useful phototherapy source having less divergence, much higher and more stable output powers, and quasimonochromaticity whereby nearly all of the photons were at the rated wavelength.) This new generation of LEDs also had its own phenomenon associated with photon intensity, namely photon interference, whereby intersecting beams of LED energy from individual LEDs produced photon interference, increasing the photon intensity dramatically and thus offering much higher photon intensities than the older generation. For LEDs emitting at visible red and near IR wavelengths, the greatest photon intensity was actually seen beneath the surface of the target tissue, due to the combination of the photon interference phenomenon and the excellent tissue scattering characteristics of light at these wavebands.) This phenomenon, together with quasimonochromaticity, meant that the new generation of LEDs was a clinically viable source for phototherapy.) ‘Low level laser therapy’ was therefore renamed by the US photobiologist, Kendric C Smith, as ‘low level light therapy’, to encompass LED energy.) Accordingly, useful bioreactions could then be achieved with LEDs through cellular photoactivation without heat or damage, as shown by Whelan and colleagues in their early NASA LED wound healing studies.)

Although visible and near-infrared light energy induce the same tri-stage process in target cells, namely photon absorption, intracellular signal transduction and the final cellular photoresponse,) it should be noted that both wavebands have different primary targets and photoreactions in target cells. Visible light is principally a photochemical reaction, acting directly and mostly on cytochrome-c oxidase, the end terminal enzyme in the cellular mitochondrial respiratory chain,) and mainly responsible for inducing adenosine triphosphate (ATP) synthesis, the fuel of the cell and indeed the entire metabolism. Infrared light on the other hand induces a primary photophysical reaction in the cell membrane thereby kick-starting the cellular membrane transport mechanisms such as the Na++K++ pump,) and this in turn induces as a secondary reaction the same photochemical cascade as seen with visible light, so the end result is the same even though the target is different as illustrated schematically in Figure 2.

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The process of cellular photoactivation by low level light therapy (LLLT). Visible light induces a primary photochemical response particularly associated with mitochondrial cytochrome c-oxidase, whereas near IR induces a primary photophysical response in the cellular and organelle membranes. However the eventual photoresponse is the same. (Based on data from Karu & Smith, Refs 6 & 9)

LED phototherapy at appropriate wavelengths and parameters has now been well-reported in a large number of pan-speciality applications.) How and where does LED phototherapy work? When we consider investigating how LED phototherapy or LLLT can bring about and influence the molecular mechanism for cell proliferation, we should recognize that LLLT not only has an effect on various signaling processes, but it can also significantly induce the production of cytokines, such as a number of growth factors, interleukins and various macromolecules (Table 2).)

Table 2:

Molecular level activation by LLLT with appropriate LEDs (From Ref )
Classification Molecules LLLT-Associated Biological Effects
Growth factors BNF, GDNF, FGF, bFGF, IGF-1, KGF, PDGF, TGF-β, VEGF Proliferation
    Differentiation
    Bone nodule formation

Interleukins IL-1α, IL-2, IL-4, IL-6, IL-8 Proliferation
    Migration
    Immunological activation

Inflammatory cytokines PGE2, COX2, IL1β, TNF-α Acceleration/Inhibition of inflammation

Small molecules ATP, cGMP, ROS, CA++, NO, H+ Normalization of cell function
    Pain relief
    Wound healing
    Mediation of cellular activities
    Migration
    Angiogenesis

Journal of Biomedical Science 2009, 16:4

Phototherapy is Becoming Mainstream

The increasing number of papers on LLLT in the Photobiomodulation sessions presented at the 2010 and especially the 2011 meetings of the American Society for Lasers in Medicine and Surgery (ASLMS) bear witness to the fact that LLLT is no longer quite the bête noir it used to be in the USA, although there is still too much skepticism, and it has achieved a reliable status worldwide. LED phototherapy has now been well-proven to work, and is reported to be effective in a large variety of clinical indications such as pain attenuation, wound healing, skin rejuvenation, some viral diseases, allergic rhinitis, other allergy-related conditions and so on.

APPLICATIONS OF LLLT WITH LEDs

When we confirm in what fields LLLT phototherapy has been most used through a review of the literature, the main application is for pain control, with pain of almost all aetiologies responding well.) For example, 830 nm LED phototherapy significantly reduced both acute and chronic pain in professional athletes.) The first author has been using LED in the control of herpes zoster pain for some time, and also for intractable postherpetic neuralgia, corroborating previous studies with 830 nm LLLT for this indication.,) This and other chronic pain entities have been historically very hard to control, but the good efficacy of LED phototherapy has been well recognized. From the large body of work from Rochkind and colleagues in Israel, LED phototherapy can help nerve regeneration, so it has been used for spinal cord injuries,) and many different types of neurogenic abnormality. In the case of the dental clinic and for the osseointegration of implants and prostheses in maxillofacial surgery it has been used for guided bone regeneration.) At present, the research into and development of new applications for LED phototherapy, especially in the processes of inflammatory cell regulation, are being assiduously studied in the dermatology field.

Fast taking over from pain attenuation, and particularly in the dermatology field, wound healing with LED phototherapy has attracted much attention. Reports have shown that, after making uniform burn wounds with a surgical laser, LED phototherapy of experimental wounds induces faster and better organized healing than in the control unirradiated wounds. This is due to the effect of 830 nm phototherapy on raising the action potential the wound-healing cells, at all three phases of the process, particularly mast cells,) macrophages) and neutrophils) in the inflammatory stage; fibroblasts in the proliferative phase (Personal Communication, Prof. Park, Seoul National University, Seoul, South Korea: unpublished data); and fibroblast-myofibroblast transformation in the remodeling phase.) As an additional mechanism, it has also been shown that 830 nm phototherapy increased the early vascular perfusion of axial pattern flaps in a controlled speckle flowmetry Doppler trial in the rat model, with actual flap survival significantly better in the irradiated than in the unirradiated control animals.)

In another very popular indication, studies have reported on the use of LED phototherapy for the rejuvenation of chronologically and photodamaged skin.,) Lee and colleagues, in a randomized controlled study, showed that fibroblasts examined with transmission electron microscopy appeared more active, collagen and elastin synthesis was increased and tissue inhibitors of matric metalloproteinases was increased, as a result of which, effective rejuvenation could be achieved which was maintained up to 12 weeks after the final treatment session. Patient satisfaction scores bore these histopathological findings out (Figure 3).) We must never forget that good skin rejuvenation is firmly based on the wound healing process, particularly neocollagenesis. LED phototherapy has also been reported as being very effective in the prophylaxis against scar formation, due amongst other factors to the response to photomediated interleukin-6 signaling.) Hair loss is another field where LED phototherapy may well have real efficacy, with red and infrared being the wavelengths of choice.) Figure 4 illustrates schematically the mechanisms already confirmed underlying the three main endpoints of 830 nm LLLT, namely wound healing, the anti-inflammatory response through acceleration and quenching of the post-wound inflammatory phase and pain attenuation.

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Patient satisfaction curves compared for LED-mediated skin rejuvenation with 633 nm alone, 633 nm + 830 nm combined and 830 nm on its own, showing the numbers of patients who rated their improvement as excellent on a 5-scale rating. The first set of columns represents the findings immediately after the 8th of 8 weekly sessions, twice per week for 4 weeks. The 2nd, 3rd and 4th sets of columns are the findings at post-treatment weeks 4, 6 and 8 respectively. At all stages, LED phototherapy with 830 nm produced superior satisfaction. The increase over the post-treatment period is interesting, suggesting improved results through continued tissue remodeling as part of the LED-mediate wound healing process. (Data adapted from Ref )

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Mechanisms underlying the three main LLLT endpoints, particularly associated with the wavelength of 830 nm, although 633 nm has beneficial effects as well.

SYSTEMIC EFFECTS OF LED-LLLT

One of the advantages of LLLT with an LED system as compared with a laser source is that LED-based systems offer large planar arrays, so that they can irradiate a large area of the body in a hands-free manner, compared with the point-by-point application of a laser system. In addition, many different cell types can be simultaneously targeted. It may not even be necessary to irradiate every target area. The systemic effect of LED with an 830 nm system (HeaLite II, Lutronic Corp., Goyang, S. Korea, Figure 5) was studied by the first author.) The systemic effect associated with LLLT has already been suggested as far back as Mester's pivotal study on non-healing ulcers in 1969, whereby irradiation of one part of the body could induce effects in another unirradiated area.) To assess this, in the first author's study controlled wounds on the backs of rodents were created with an ablative fractional laser, and rather than irradiating the laser wounds with LED energy (HeaLite system as above), the animals' abdomens in the experimental group were irradiated, and sham irradiation was delivered to the control group. The results clearly indicated that the group which had LED treatment of the abdomen demonstrated significantly better healing than the control group (Figure 6). This means that LED phototherapy could very probably have a systemic effect on inflammatory or immune cells in nonadjacent tissues to the target area, as well as those cells in the irradiated tissues.

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HeaLite II LED phototherapy system, Lutronic Corp, Goyang, South Korea.

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The wound healing value compared between the group treated with 830 nm LED (Tx group) with LED and the unirradiated Con group without LED. Note that the 1 LED-irradiated animal in the 0–25% group had somehow removed the wound dressing very early in the experiment. (Adapted from Ref )

LED LLLT FOR SKIN INFLAMMATORY DISEASES

The anti-inflammatory effect of LED has been generally accepted, but up till now this has not been well shown well in inflammatory skin diseases such as allergic or irritant contact dermatitis, atopic dermatitis or rosacea, although a significant degree of success has been demonstrated and reported for inflammatory acne and recalcitrant treatment-resistant psoriasis.,) In an experimental animal model study the first author was able to demonstrate that when induced dermatitis in rats was treated with 830 nm LED phototherapy (HeaLite II system, Lutronic Corp, as above) at a dose of 60 J/cm2 in continuous wave, compared with an untreated control group, the histopathological findings revealed significant decreased levels of inflammatory cells (Figure 7). Based on the success of that study, treatment-resistant inflammatory contact dermatitis due to a peel compound containing alpha-hydroxy acid (AHA) in a human subject also responded very well to 3 sessions of 830 nm LED therapy, 3 days apart, irradiance of 100 mW/cm2, 10 min/session, dose of 60 J/cm2, continuous wave (Figure 8).

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The changes in dermatitis-associated inflammatory cells following 830 nm LED irradiation in the rat model (A: Control specimen, B: LED irradiated specimen). A marked reduction in inflammatory infiltration is evident.

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Improvement in a patient (24-year-old female) with treatment-resistant post-chemical peel irritant contact dermatitis (AHA-related ICD) seen above at baseline, and below 10 days later following 3 830 nm LED treatment sessions, 3 days apart, 20 minutes per session (60 J/cm2)

Here are another two examples of the clinical success of 830 nm LED phototherapy (continuous wave, 60 J/cm2) in difficult-to-treat conditions. Figure 9 illustrates the dramatic improvement following 830 nm LED phototherapy in a case of dissecting cellulitis of the scalp, a recalcitrant inflammatory problem, treated with 4 sessions over 2 weeks, 20 min/60 J/cm2 per session; and Figure 10 illustrates a typical result 10 weeks after 6 sessions over 6 weeks, 20 min/60 J/cm2 per session, from a clinical trial the first author has conducted on LED therapy for rosacea with neutrophilic dermatitis. This trial is as yet unreported because the full 12-week follow-up time has not yet been reached in all patients. However, preliminary results are very encouraging with no recurrence seen at 10 weeks in those patients who have reached that point.

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Dramatic improvement in a case of dissecting cellulitis of the scalp (34-year-old male) (a) at baseline and (b) following 830 nm LED treatment (twice per week for 2 weeks, 20 min per session to give 60 J/cm2)

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Improvement of acne rosacea (33-year-old female) at baseline (a) and following LED treatments (once per every week for 6 weeks, 20 min and 60 J/cm2 per session) (b). Although not very well noted in the grayscale illustrations, the small acneiform papules have disappeared, with a clear decrease seen in the redness on both cheeks.

CONCLUSIONS

In conclusion, based on the published data and the authors' own experience, LED phototherapy is proving to have more and more viable applications in many fields of medicine. However, it must always be remembered that not any old LED will do. In order to be effective, LED phototherapy must satisfy the following 3 criteria.

  • • The LED system being used must have first of all, and most importantly, the correct wavelength for the target cells or chromophores. At present, the published literature strongly suggests 830 nm for all aspects of wound healing, pain, anti-inflammatory treatment and skin rejuvenation, with a combination of 415 nm and 633 nm for light-only treatment of active inflammatory acne vulgaris. If the wavelength is incorrect, optimum absorption will not occur and as the first law of photobiology states, the Grotthus-Draper law, without absorption there can be no reaction.
  • • Secondly, the photon intensity, i.e., spectral irradiance or power density (W/cm2), must be adequate, or once again absorption of the photons will not be sufficient to achieve the desired result. If the intensity is too high, however, the photon energy will be transformed to excessive heat in the target tissue, and that is undesirable.
  • • Finally, the dose or fluence must also be adequate (J/cm2), but if the power density is too low, then prolonging the irradiation time to achieve the ideal energy density or dose will most likely not give an adequate final result, because the Bunsen-Roscoe law of reciprocity, the 2nd law of photobiology, does not hold true for low incident power densities.

Provided these three criteria are met, LED phototherapy does indeed work, and has many useful aspects in clinical practice for practitioners in many surgical specialities. As an exciting extension of the monotherapy approach with LED-LLLT, and even more importantly, the combination of appropriate LED phototherapy as an adjunct to many other surgical or nonsurgical approaches where the architecture of the patient's skin has been altered will almost certainly provide the clinician with even better results with less patient downtime, in a shorter healing period, and with excellent prophylaxis against obtrusive scar formation.

References

1. Ohshiro T, Calderhead RG: Low Level Laser Therapy: a Practical Introduction. 1988. John Wiley and Sons, Chichester, UK
2. Mester E, Szende B, Spiry T, Scher A. (1972): Stimulation of wound healing by laser rays. Acta Chir Acad Sci Hung. Acta Chir Acad Sci Hung: 315–324 [PubMed]
3. Calderhead RG, Ohshiro T, Ito E, Okada T, Kato K: The Nd:YAG and GaAlAs lasers: a comparative analysis in pain therapy. In Atsumi K, Nimsakul N, editors. , Eds: Laser Tokyo ‘81. 1981, Japan Society for Laser Surgery and Medicine, Tokyo: Section 21, pp 1–4
4. Whelan HT, Houle JM, Whelan NT. et al. (2000): The NASA Light-Emitting Diode Medical Program-Progress in Space Flight and Terrestrial Applications. Space Tech. & App. Int'l. Forum. Space Tech. & App. Int'l. Forum: 37–43
5. Calderhead RG. (2007): The photobiological basics behind light-emitting diode (LED) phototherapy. Laser Therapy, 16: 97–108
6. Smith KC. (2005): Laser (and LED) therapy is phototherapy. Photomed Laser Surg. 23: 78–80 [PubMed]
7. Smith KC. (2010): Laser and LED photobiology. Laser Therapy, 19: 72–78
8. Whelan HT, Smits RL, Buchmann EV. et al (2001): Effect of NASA Light-Emitting Diode (LED) Irradiation on Wound Healing. J Clin Laser Med Surg, 2001. 19: 305–314 [PubMed]
9. Karu T. (1999): Primary and secondary mechanisms of action of visible to near-IR radiation on cells. J Photochem Photobiol B, J Photochem Photobiol B: 1–17 [PubMed]
10. Karu T: Identification of the photoreceptors. In: Karu T, editor. Ten Lectures on Basic Science of Laser Phototherapy. 2007, Prima Books AB, Grangesberg, Sweden
11. Tunér J, Hode L: The New Laser Therapy Handbook. 2010, Prima Press, Grangesborg, Sweden
12. Gao X, Xing D. Molecular mechanisms of cell proliferation induced by low power laser irradiation. J Biomedical Science. 2009. 16:4 [PMC free article] [PubMed]
13. Baxter GD, Bleakley C, Glasgow P, Calderhead RG. (2005): A near-infrared LED-based rehabilitation system: initial clinical experience. Laser Therapy, Laser Therapy: 29–36
14. Moore KC, Hira N, Kumar PS, Jayakumar CS, Ohshiro T. (1988): A double blind crossover trial of low level laser therapy in the treatment of postherpetic neuralgia. Laser Therapy, Pilot Issue: 7–10
15. Numazawa R, Kemmotsu O, Otsuka Hi, Kakehata J, Hashimoto T, Tamagawa S. (1996): The rôle of laser therapy in intensive pain management of postherpetic neuralgia. Laser Therapy, Laser Therapy: 143–148
16. Rochkind S. (2009): Review of 30-years experience: laser phototherapy in neuroscience and neurosurgery part II-nerve cells, brain and spinal cord. Laser Therapy, 18: 127–136
17. Asanami S, Shiba H, Ohtaishi M, Okada Y, Ohshaka F, Tanaka Y. (1993): The activatory effect of low incident energy hene laser irradiation on hydroxyapatite implants in rabbit mandibular bone. Laser Therapy, Laser Therapy: 29–32
18. Calderhead RG, Kubota J, Trelles MA, Ohshiro T. (2008): One mechanism behind LED phototherapy for wound healing and skin rejuvenation: key role of the mast cell. Laser Therapy, 17: 141–148
19. Young S, Bolton P, Dyson M, Harvey W, Diamantopoulos C. (1989): Macrophage responsiveness to light therapy. Lasers Surg Med, Lasers Surg Med: 497–505 [PubMed]
20. Osanai T, Shiroto C, Mikami Y, Kudou E. et al. (1990): Measurement of GaAlAs diode laser action on phagocytic activity of human neutrophils as a possible therapeutic dosimetry determinant. Laser Therapy, 1990. 2: 123–134
21. Enwemeka CS, Cohen-Kornberg E, Duswalt EP, Weber DM, Rodriguez IM. (1994): Biomechanical effects of three different periods of GaAs laser photostimulation on tenotomized tendons. Laser Therapy, Laser Therapy: 181–188
22. Kubota J. (2002): Effects of diode laser therapy on blood flow in axial pattern flaps in the rat model. Lasers Med Sci, 17: 146–153 [PubMed]
23. Goldberg DJ, Amin S, Russell BA, Phelps R. et al: Combined 633-nm and 830-nm led treatment of photoaging skin. J Drugs Dermatol, 2006. 5: 748–753 [PubMed]

Original Source: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3799034/

Home Search Introduction

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Welcome to the laser-therapy.us research tool. This tool is a searchable collection of technical publications, books, videos and other resources about the use of lasers for photobiomodulation. This tool includes almost the entire U.S. library of medicine research papers on LLLT, videos from Youtube associated with therapy lasers and the tables of contents from laser therapy books. This allows users to search for a keyword or condition and see resources about using lasers to treat that condition. All the resources include links to the original source so we are not making any statement about the use of lasers for treating non-FDA cleared application, we are simple summarizing what others have said.  Where every possible, we have included a link to the orginal publication.

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This tool uses a broad match query so:

  • It does not correct spelling and searches only cold laser related subjects so do not use LLLT, cold or laser in the search bar
  • It works better with shorter search terms or even parts of search terms
  • It searches all the available fields so you can enter a body part, author, condition or laser brand.
  • Where ever possible, the detailed section about the resource will link to the sources.
  • This system is only for photobiomodulation or cold laser therapy research (including LLLT, laser acupuncture and high power laser therapy) only. It does NOT include photodynamic laser therapy (where the laser is used to react with a pharmaceutical), hot surgery lasers or cosmetic lasers. It does include some resources on weight loss and smoking cessation.

The results of the search are sorted based on 3 quality factors on a scale of 1 to 10 with 10 being the best score. Originally all the resources were given a 5-5-5 until they could be individually evaluated. These scores are purely opinion and are only used to simplify the rank of the results from more valuable to least valuable. This should not be considered a critique of any work. This system was created to help researchers (including ourselves) find the most usable resources for any cold laser therapy research. The resources are assigned values based on the following 3 factors:

  • Efficacy: The resource (especially research papers) should show a significant improvement in the condition being treated. Resources that show better results are given a higher quality score.
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Over the past few years of working with research, we found that a majority of the published resources are lacking in one of these three ranking factors.
The original goal of this research tool was to tie published resources to the protocols in the laser-therapy.us library. This connection allows users to trace each protocol back to a list of resources so the protocol can be researched and improved.

General Comments


POWER
When many of the first research papers were published, the most power laser available for therapy were less than 100mW and many systems had to be pulsed to keep the laser from burning out too quickly. Today, system are available that will deliver up to 60,000mW of continuous output. Because of these power limitation, many early studies were limited to extremely low dosages by today’s standards. It takes a 50mW system 17 minutes to deliver 50 joules at the surface of the skin. If this was spread over a large area of damage or was treating a deeper problem, the actual dosages were much less than 1J/cm2.  Today, we know that these dosages typically produce very little or no results.
WAVELENGTH
About 80% of the resources in this database are in the near infrared wavelength. There is also some interest in the red wavelength (600 to 660nm) . Other wavelengths like blue, purple, and green have very little scientific research behind them and have not gotten much traction in the core therapy market with the exception of some fringe consumer products.
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