Different Power Settings of LLLT on the Repair of the Calcaneal Tendon

Abstract

The purpose of this study was to evaluate the effect of an 830-nm GaAlAs diode laser operating at output powers of 40, 60, 80, and 100 mW and energy density of 30 J/cm(2) on the repair of partial calcaneal tendon ruptures in rats.
A partial tendon rupture was induced in all animals, which were treated with laser irradiation for 5 consecutive days. Six days after injury, the injured tendons were removed and examined by polarized light microscopy. Collagen fiber organization was evaluated by birefringence measurements, and collagen content was determined by Picrosirius Red staining.
It was observed that the higher the output power (60-100 mW) the greater the amount of type III collagen (p<0.01). The amount of type I collagen was significantly greater (p=0.05) in the 80 mW group than in the control group (sham stimulation). A non-statistically significant improvement in the realignment of collagen fibers was observed in the irradiated groups.
Low-level laser therapy resulted in significantly greater amounts of type III collagen (output powers of 60 mW or more) and type I collagen (output power of 80 mW), however, no significant differences between groups were found in the realignment of collagen fibers.

Full text

Different Power Settings of LLLT on the Repair
of the Calcaneal Tendon
Marco Aurelio Invaldi Neves, P.T., M.S.,
1
Carlos Eduardo Pinfildi, P.T., Ph.D.,
1,2
Viviane Timm Wood, P.T., M.S.,
1
Rafael Correˆ a Gobbato, P.T., M.S.,
1
Fabio Mendes da Silva, M.D.,
1
Nivaldo Antoˆ nio Parizotto, P.T., Ph.D.,
3
Bernardo Hochman, M.D., Ph.D.,
1
and Lydia Masako Ferreira, M.D., Ph.D.
4
Abstract
Objective: The purpose of this study was to evaluate the effect of an 830-nm GaAlAs diode laser operating at
output powers of 40, 60, 80, and 100 mW and energy density of 30 J/cm
2
on the repair of partial calcaneal tendon
ruptures in rats. Methods: A partial tendon rupture was induced in all animals, which were treated with laser
irradiation for 5 consecutive days. Six days after injury, the injured tendons were removed and examined by
polarized light microscopy. Collagen fiber organization was evaluated by birefringence measurements, and
collagen content was determined by Picrosirius Red staining. Results: It was observed that the higher the output
power (60–100 mW) the greater the amount of type III collagen ( p<0.01). The amount of type I collagen
was significantly greater ( p=0.05) in the 80 mW group than in the control group (sham stimulation). A non-
statistically significant improvement in the realignment of collagen fibers was observed in the irradiated groups.
Conclusions: Low-level laser therapy resulted in significantly greater amounts of type III collagen (output
powers of 60 mW or more) and type I collagen (output power of 80 mW), however, no significant differences
between groups were found in the realignment of collagen fibers.
Introduction
The calcaneal tendon is one of the most commonly
injured tendons in the human body.
1,2
Due to the
increasing physical activities of the population, the tendon
lesion like tendinopathies had increased and the correlation
with this lesion can be related to microtraumas and over-
use.
3–5
The tendinous tissue is a dense connective tissue, whose
function is to transmit the force of a muscle to a bone, to
produce movement.
6,7
Tendons consist of fibroblasts and an
extracellular matrix in which fibrous proteins of collagen,
elastine, proteoglycans, glycoproteins, and multiple saccha-
rides are immersed.
8
Collagen is the main structural protein
and major component of the extracellular matrix (86–95%
wet weight). Collagen fibrils are long, highly aligned, and
have a crystalline structure, which gives them high me-
chanical resistance to tension.
9–12
The healing of a calcaneal tendon may take weeks or even
months to complete, making adherence to the treatment
regimen difficult.
13–16
Because of the high incidence of these
injuries, there is a need for studies focusing on the im-
provement of tendon repair, reducing recovery time and the
time to return to normal activities.
Satisfactory results have been obtained in studies con-
ducted with the purpose of finding the means to accelerate
the healing of the calcaneal tendon
17
using physical agents
such as ultrasound,
3,18
electrical stimulation,
19
and low-level
laser therapy (LLLT).
20,21
LLLT has provided relevant results such as an increase in
fibroblast proliferation and collagen synthesis,
22
cutaneous
neovascularization,
23,24
and tendon repair.
21,25
However,
there are variations in the parameters used (e.g., wavelength,
energy density, output power) by different investigators.
Because only a few studies on LLLT have investigated the
effect of different output powers, there is no consensus on
which is the optimal output power for use in tendon repair.
Therefore, the aim of the present study was to evaluate the
effect of output powers of 40, 60, 80, and 100 mW on the
repair of partial calcaneal tendon rupture in rats.
1
Department of Plastic Surgery, Universidade Federal de Sa
˜o Paulo (UNIFESP), Sa
˜o Paulo, Brazil.
2
Department of Health Science, Universidade Federal de Sa
˜o Paulo (UNIFESP), Campus Baixada Santista, Santos, Sa
˜o Paulo, Brazil.
3
Department of Physiotherapy, Universidade Federal de Sa
˜o Carlos, UFSCar, Sao Paulo, Brazil.
4
Division of Surgery Department and Post-Graduate Program in Plastic Surgery, Universidade Federal de Sa
˜o Paulo (UNIFESP), Sa
˜o
Paulo, Brazil.
Photomedicine and Laser Surgery
Volume 29, Number 10, 2011
ªMary Ann Liebert, Inc.
Pp. 663–668
DOI: 10.1089/pho.2010.2919
663

Methods
All animal experiments were approved by the Animal
Care and Use Committee at the Federal University of Sa
˜o
Paulo (UNIFESP-EPM). All animals received humane care in
strict compliance with the Guide for the Care and Use of
Laboratory Animals (National Research Council, 1996).
The sample consisted of 50 adult male Wistar rats (Rattus
norvegicus), weighing 260–320 g. The animals were obtained
from the Central Laboratory Animal Facility of the Federal
University of Sa
˜o Paulo (UNIFESP), Sa
˜o Paulo, Brazil. The
rats were housed in individual polypropylene cages, on a
12:12 h light–dark cycle, and fed standard rat chow and
water ad libitum.
The animals were randomly divided into 5 groups of
10 rats each using the Statistical Software Biostat 5.0 (Brazil-
Para
´-Bele
´m). A partial tendon rupture was induced by direct
trauma in all animals. After, each group was subjected to the
following procedures: the control group received sham stim-
ulation, and the experimental groups received laser treatment
for 5 consecutive days. Six days after trauma-induced injury,
the animals were killed, and the calcaneal tendons were re-
moved and examined by polarized light microscopy.
Procedure to induce partial calcaneal tendon rupture
The animals were weighed and anesthetized with an in-
traperitoneal injection of ketamine hydrochloride (100 mg/
kg) and xylazine hydrochloride (50 mg/kg). The skin over
and around both the right and left calcaneal tendons was
shaved; the paw subjected to trauma-induced injury was
selected by lottery (Statistical Software BioStat 5.0). An injury
device was developed at the machine shop of the Federal
University of Sa
˜o Carlos (UFSCar).
The selected paw was positioned on the injury device and
a light tensile force was applied to the calcaneal region in
such way that the ankle was maintained in dorsiflexion and
the dorsal surface of the paw was in contact with the injury
device. Following, a 186 g weight was dropped on the ten-
don of the animal from a height of 20 cm (potential energy,
364.9 mJ), (Fig. 1).
Immediately after this procedure, the weight was removed
and the injury site was marked with a circle using a skin pen
for the laser treatment. Next, the animals were returned to
their cages and were observed until the anesthesia wore off.
Laser treatment
The laser treatment was started after trauma-induced in-
jury, and all animals were subjected to the treatment for 5
consecutive days. Treatment sessions were conducted on
each animal at the same time of day as the first laser appli-
cation. The laser probe was placed in direct contact with the
animal on only one treatment point at the injury site and
positioned perpendicularly to the calcaneal tendon.
Laser treatment was delivered using a GaAlAs diode laser
(DMC

, Sao Carlos, Sao Paulo, Brazil) emitting at a wave-
length of 830 nm (infrared), with output powers of 40, 60, 80,
and 100 mW, energy density of 30 J/cm
2
, total energy dose of
0.84 J, beam cross-sectional area of 0.028 cm
2
, and operating
in a continuous mode.
The animals were randomized into five groups and each
group was subjected to laser treatment and irradiated as follows:
Control group – sham stimulation
40 mW group – output power of 40 mW and power
density of 1.4 W/cm
2
for 21 sec
60 mW group – output power of 60 mW and power
density of 2.14 W/cm
2
for 14 sec
80 mW group – output power of 80 mW and power
density of 2.8 W/cm
2
for 10.5 sec
100 mW group – output power of 100 mW and power
density of 3.5 W/cm
2
for 8.4 sec
Tendon excision and sample preparation
Six days after trauma-induced injury, the animals were
killed by anesthesia overdose. For removal of the calcaneal
tendon, two incisions were made in each animal: one in the
musculotendinous junction, and another proximal to the
calcaneal insertion. Five seconds after removal, the calcaneal
tendon was affixed to a paraffin surface, which had been
previously molded in a Petri dish. Two pins were used to
affix the tendon to the paraffin; the first pin was placed in the
base of the tendon (osteotendinous junction), and the second
pin was placed in the gastrocnemius muscle. Following, 10%
buffered formaldehyde was poured into the Petri dish until
the sample was covered by liquid. The sample was main-
tained immersed for 40 min, after which the formaldehyde
was removed, an incision was made 2 mm below the mus-
culotendinous junction, and another incision was made
2 mm above the osteotendinous junction using a shaving
blade.
All samples were cut to the same length of 0.8 cm. Next,
each sample was wrapped in paper filter (3 ·2 cm) and
placed inside a small box, which was immersed in 10%
buffered formaldehyde for 24 h, and then in 70% alcohol for
48 h, initiating the preparation of histological slides for po-
larized light microscopy.
Following this, the organization, state of aggregation and
molecular arrangement of collagen fibers were evaluated by
birefringence measurements and the collagen content was
determined by Picrosirius Red staining.
FIG. 1. Injury device for partial calcaneal tendon rupture.
Overall view of the weight over the tendon at the moment of
the injury after the weight was dropped.
664 NEVES ET AL.

Birefringence measurements
Birefringence optical retardation (OR) measurements were
made in monochromatic light (546 nm) using a light micro-
scope (Leica, Ernst-Leitz, Strabe, Wetzlar, Germany) with a
POL 10 ·/0.22 eyepiece, a 0.9 condenser, and a compensator
(k/4, Senarmont).
Prior to the birefringence analysis, all histological slides
were immersed in distilled water for 30 min. During the
measurements, the longitudinal axis of the tendon was po-
sitioned at the angle of 45with respect to the microscope
polarizer; at this position, the optimum value of optical re-
tardation for collagen fibers was obtained (OR =brightness).
Picrosirius Red staining
The tissue stained with Picrosirius Red was examined by
polarized light microscopy (Nikon E-800 microscope, London,
UK) for the presence of thin type III collagen fibers (green)
and thick type I collagen fibers (red and yellow). Ten micro-
scopic fields (field area, 303598.2lm
2
; total area, 3035982 lm
2
)
were analyzed per slide. Images were imported to the Image-
Pro Plus 4.5 software (Bethesda, MD) for quantitative analysis;
results are expressed as mean percentages of the total area
occupied by each type of collagen fiber.
Statistical analysis
The reproducibility of the measurements made by two
observers was assessed using the intraclass correlation co-
efficient (ICC) and the Bland-Altman graphs.
The one-way analysis of variance (ANOVA) was used for
group comparison. Variance equality hypothesis was veri-
fied by the Levene’s test. When the variances were different,
the Brown-Forsythe correction and the Dunnett’s multiple
comparison methods were used. Significance was set at
p<0.05 for all analyses
Results
Birefringence results indicated that groups treated with
LLLT showed a non-statistically significant improvement
(p>0.05) in the realignment of collagen fibers associated
with an increase in the output power (Figs. 2 and 3).
It was observed that the higher the output powers (60–
100 mW) the greater was the amount of type III collagen.
Significant differences in the amount of type III collagen
were found between the control group and the 60 mW
(p<0.01), 80 mW ( p<0.01), and 100 mW ( p=0.02) groups
(Table 1).
Significantly greater amounts of type I collage fibers were
found in 80 mW group than in the control group ( p=0.05).
No significant differences in the amounts of type I collagen
were found between the control group and the other ex-
perimental groups ( p>0.05) (Figs. 4 and 5).
Discussion
Van Breugel and Bar
26
reported that most studies have
considered energy density as the most important variable
FIG. 2. Quantitative birefrin-
gence measurements showing
the organization of collagen
fibers in the different groups:
(A) Control group (sham
stimulation), (B) 40-mW
group, (C) 60-mW group,
(D) 80-mW group, and (E)
100-mW group. The longitu-
dinal axis of the calcaneal
tendon was positioned at a
45angle for optimum value
of optical retardation. Arrows
and marks (Aand *) show
the high degree of parallelism
of collagen fibers in the cal-
caneal tendon.
FIG. 3. Box plot showing mean optical retardation (OR)
values for the different groups. Mean OR values were non-
statistically significant higher in the 40-, 60-, 80- and 100-mW
groups than in the control group ( p>0.05).
LLLT POWER TENDON HEALING 665

with respect to the therapeutic effects of laser irradiation.
Few studies have been conducted to evaluate the effect of
output power on the results of laser therapy.
Tune
´r and Hode
27
suggested that a high output power
may lead to a high energy density. Many studies on laser
therapy
25,27,28
were conducted with lasers with output
powers ranging from 10 to 40 mW, but there are few studies
in the literature using lasers with output powers ranging
from 40 to 100 mW.
29
Based on this fact, four different power
settings were used in the present study (an intermediate
value of 40 mW, and three higher values of 60, 80, and
100 mW) in order to determine which power setting would
deliver the best results.
According to Chan et al. (2007),
19
the tendon repair pro-
cess is similar to other repair processes that occur in bio-
logical tissues. The repair process is associated with the
proliferation and migration of different types of cells, colla-
gen synthesis, angiogenesis, and granulation tissue forma-
tion, and with the orientation of tendon cells and collagen
fibers in a highly organized manner as an effort to restore the
structure and function of the injured tendon.
In our study, the degree of orientation of collagen fibers
and the type of collagen present in the initial phase of the
inflammatory process were assessed by Picrosirius Red
staining.
Birefringence analysis was conducted to assess the align-
ment of collagen fibers, as suggested by Vidal,
7
who con-
sidered this method ideal to detect and describe the
orientation of collagen fibers of the tendon.
Birefringence results indicated a non-statistically signifi-
cant trend ( p>0.05) toward a gradual increase in mean OR
values associated with an increase in the output power as
follows: control group (52.1 nm), 40 mW group (59.42 nm),
60 mW group (61.82 nm), 80 mW group (61.59 nm), and
100 mW group (63.52 nm). These results disagree with the
Oliveira et al.
21
results about treatment days, which reported
a better organization of collagen fibers using an LLLT 850 nm
with 1.4 W/cm
2
and 4 J/cm
2
on the 5th day, whereas our
study showed better results on the 6th day after injury. In the
present study, the tendon repair probably began to occur
prematurely (from about the 3rd to the 5th day after injury)
showed by assessment of OR values, for high output powers,
should be made in a shorter period of time after injury. We
believe that the nonsignificant results may be attributed to
the small size of the sample and due evaluation period, like
7th day post lesion. However, we observed a gradual in-
crease in the organization of collagen fibers associated with
increase in the output power.
Significant differences in the amount of type III collagen
were found between the control group and the 60, 80, and
Table 1. Mean Values and Standard Deviation (SD) of Optical Retardation (OR)
and Percentage of Types I and III Collagen in the Study Groups
OR (nm) Type III collagen (%) Type I collagen (%)
Groups Mean SD Groups Mean SD Groups Mean SD
G1 (Sham) 52.10 11.97 G1 (Sham) 31.96 12.10 G1 (Sham) 2.60 1.79
G2 (40 mW) 59.42 8.64 G2 (40 mW) 48.02 15.40 G2 (40 mW) 9.12 5.41
G3 (60 mW) 61.82 13.18 G3 (60 mW)* 55.85 12.58 G3 (60 mW) 9.01 5.36
G4 (80 mW) 61.59 7.01 G4 (80 mW)* 60.90 9.24 G4 (80 mW)* 15.40 6.11
G5 (100 mW) 63.52 5.80 G5 (100 mW)* 61.55 9.57 G5 (100 mW) 6.50 4.96
p=0.139 *p£0.01 *p=0.05
FIG. 4. Box plot showing
the percentage of type I
and III collagen for the
different groups. The per-
centage of type I collagen
was significantly higher
in the 80-mW group than
in the control group ( p=
0.05). The percentage of
type III collagen was sig-
nificantly higher in the 60-,
80- and 100-mW groups
than in the control group
(p<0.05).
666 NEVES ET AL.

100 mW groups ( p<0.01), with the 100 mW group showing
the best results. On the other hand, significant differences in
the amount of type I collagen fibers were found only be-
tween the 80 mW group and the control group; this is an
isolated case, as there is no study in the literature reporting
similar results. Maffulli et al.,
30
in a previous study on an
in vitro model of human tendon healing, reported greater
amounts of type III collagen in ruptured and tendinopathic
Achilles tendons than in normal Achilles tendons. The
amount of type III collagen in injured calcaneal tendons may
be related to the phase of lesion development. Many studies
in the literature reported a reduced amount of type I collagen
and an increased amount of type III collagen in injured cal-
caneal tendons.
30,31
The greater amounts of type III collagen found in the 60,
80, and 100 mW groups may be explained by the fact that
LLLT with high output powers may deliver a signal to the
extracellular matrix that induces changes in fibril structures.
These changes may occur because of the interaction between
electromagnetic energy and collagen molecules. The azi-
muthal orientation of the collagen along the longitudinal axis
of the tendon is a good evidence of structural changes in col-
lagen fibers after irradiation.
18,32
The results from the present
study suggest that LLLT using infrared lasers operating at
output powers of 40, 60, 80, and 100 mW has a positive effect
on the treatment of partial calcaneal tendon rupture. Our re-
sults are consistent with the findings of other studies reporting
on the use of LLLT with longer time periods as a treatment for
tissue repair, in which the response of fibroblasts and collagen
molecules to laser irradiation were evaluated.
33–35
The results of this study raise an important question re-
garding the parameters. Is power or power density the im-
portant factor to improve the tissue repair? In 2005 the World
Association for Laser Therapy (WALT) published the dosage
guidelines showing the importance of the energy (joules) and
power density. The maximum power density recommended to
treat the Achilles tendon is 100 mW/cm
2
, but this study used
power densities of 1.4–3.5W/cm
2
and still produced positive
effects. Further studies are necessary to clarify the validity of
the 100 mW/cm
2
limit stated in the WALT guidelines.
Further studies are necessary to study the power densities
such as 80, 90, or 100 mW/cm
2
.
Conclusions
The results also revealed that the laser output power af-
fects tissue repair. However, it is still premature to state that
output power is a fundamental factor in accelerating tendon
repair. Further studies on LLLT using infrared lasers are
necessary to better understand the effect of output power on
tissue repair.
Acknowledgments
We thank the Division of Plastic Surgery of the UNIFESP,
Lamav-UFSCar and the Coordination for the Improvement of
Higher Education Personnel (CAPES) for supporting this study.
Author Disclosure Statement
No conflicting financial interests exist.
References
1. Suchak, A.A., Bostick, G., Reid, D., Blitz, S., and Jomha, N.
(2005). The incidence of Achilles tendon ruptures in Ed-
monton Canada. Foot Ankle Int. 26,932–936.
2. Tumilty, S. (2007). Achilles tendon ruptures: rising incidence
in New Zealand follows international trends. Phys. Ther.
Rev. 12, 59–65.
3. Demir, H., Menku, P., Kirnap, M., et al. (2004). Comparison
of the effects of laser, ultrasound and combined laser +
ultrasound treatments in experimental tendon healing. La-
sers Surg. Med. 35, 84–89.
4. Tumilty, S., Munn, J., Abbott, J.H. et al. (2008). Laser therapy
in the treatment of Achilles tendinopathy: a pilot study.
Photomed. Laser Surg. 26, 25–30.
5. Wren, T.A.L., Yerby, A.S., Beaupre, G.S., and Carter, D.R.
(2001). Mechanical properties of the human Achilles tendon.
Clin. Biomech. 16, 245–251.
6. Arruda, E.R.B., Rodrigues, N.C., Tarciro, C., and Parizotto,
N.A. (2007). Influence of different low-intensity laser ther-
apy wavelengths for rat tendon regeneration following te-
notomy. Rev. Bras. Fisioter. 11, 283–288.
7. Vidal, B.C. (2003). Image analysis of tendon helical super-
structure using interference and polarized light microscopy.
Micron 34, 423–432.
8. Enwemeka, C.S. (1989). Inflammation, cellularity, and fi-
brillogenesis in regenerating tendon: implications for tendon
rehabilitation. Phys. Ther. 69, 816–826.
9. Fillipin, L.I., Mauriz, J.L., Vedovelli, K. et al. (2005). Low-
level laser therapy (LLLT) prevents oxidative stress and re-
duces fibrosis in rat traumatized Achilles tendon. Lasers
Surg. Med. 37, 293–300.
10. Matsumoto, F., Trudel, G., Uhthoff, H.S., and Backman, D.S.
(2003). Mechanical effects of immobilization on the Achilles’
tendon. Arch. Phys. Med. Rehabil. 84, 662–667.
FIG. 5. Tissue stained with
Picrosirius Red examined by
polarized light microscopy.
(A) Slide from the control
group (sham stimulation)
showing a small amount of
type I collagen, and (B) slide
from an irradiated group
showing an increased amount
of type III collagen.
LLLT POWER TENDON HEALING 667

11. Naim, F., Simsek, A., Sipahioglu, S. et al. (2005). Evaluation
of the surgical results of Achilles tendon ruptures by gait
analysis and isokinetic muscle strength measurements. Acta
Orthop. Traumatol. Turc. 39, 1–6.
12. Oliveira, R.F., Oliveira, D.A.A.P., Monteiro, W. et al. (2008).
Comparison between the effect of low–level laser therapy
and low–intensity pulsed ultrasonic irradiation in vitro.
Photomed. Laser Surg. 26, 6–9.
13. Agaiby, A.D., Ghali, L.R., Wilson, R., and Dyson, M. (2000).
Laser modulation of angiogenic factor production by T–
lymphocytes. Lasers Surg. Med. 26, 357–363.
14. Battegay, E.J. (1995). Angiogenesis: mechanistic insights,
neovascular diseases, and therapeutic prospects. J. Mol. Med.
73, 333–346.
15. Reddy, G.K., Gum, S., Stehno–Bitel, L., and Enwemeka, C.S.
(1998). Biochemistry and biomechanics of healing tendon:
Part II. Effects of combined laser therapy and electrical
stimulation. Med. Sci. Sports Exerc. 30, 794–800.
16. Stehno–Bittel, L., Reddy, G.K., Gum, S., and Enwemeka, C.S.
(1998). Biochemistry and biomechanics of healing tendon:
Part I. effects of rigid plaster casts and functional casts. Med.
Sci. Sports Exerc. 30, 788–793.
17. Enwemeka, C.S., and Reddy, G.K. (2000). The biological ef-
fects of laser therapy and other modalities on connective
tissue repair processes. Laser Therapy 12, 22–30.
18. Cunha, A., Parizotto, N.A., and Vidal, B.C. (2001). The effect of
therapeutic ultrasound on repair of the Achilles tendon (ten-
don calcaneus) of the rat. Ultrasound Med. Biol. 27,1691–1696.
19. Chan, H.K.F., Fung, D.T.C., and Ng, G.Y.F. (2007). Effects of
low–voltage microamperage stimulation on tendon healing
in rats. J. Orthop. Sports Phys. Ther. 37, 399–403.
20. Ng, G.Y.F., and Fung, D.T.C. (2008). The combined treat-
ment effects of therapeutic laser and exercise on tendon re-
pair. Photomed Laser Surg. 26, 137–141.
21. Oliveira, F.S., Pinfildi, C.E., and Parizoto N.A. (2009). Effect
of low level laser therapy (830 nm) with different therapy
regimes on the process of tissue repair in partial lesion cal-
caneous tendon. Lasers Surg. Med. 41, 271–276.
22. Almeida–Lopes, L., Rigau, J., Za
˜ngaro, R.A. et al. (2001).
Comparison of the low level laser therapy effects on cultured
human gingival fibrobalts proliferation using different irra-
diance and same fluence. Lasers Surg. Med. 29, 179–184.
23. Pinfildi, C.E., Liebano, R.E., Hochman, B.S., and Ferreira,
L.M. (2005). Helium–neon laser in viability of random skin
flap in rats. Lasers Surg. Med. 37, 89–91.
24. Prado, R.P., Liebano, R.E., Hochman, B., Pinfildi, C.E., and
Ferreira, L.M. (2006). Experimental model for low level laser
therapy on ischemic random skin flap in rats. Acta Cir. Bras.
21, 258–262
25. Salate, A.C.B., Barbosa, G., Gaspar, P. et al. (2005). Effect of
In–Ga–Al–P diode laser irradiation on angiogenesis in par-
tial ruptures of Achilles tendon in rats. Photomed. Laser
Surg. 23, 470–475.
26. Van Breugel, H.H., and Bar, P.R. (1992). Power density and
exposure time of He–Ne laser irradiation are more important
than total energy dose in photo–biomodulation of human
fibroblasts in vitro. Lasers Surg. Med. 12, 528–537.
27. Tune
´r, J., and Hode, L. (1999). Low level laser therapy.
Clinical practice and scientific background. Gra
¨ngesberg,
Sweden: Prima Books
28. Young, S., Bolton, P., and Dyson, M. (1989). Macrophage
responsiveness to light therapy. Lasers Surg. Med. 9,
497–505.
29. Corazza AV, Jorge J, Kurachi C, and Bagnato VS. (2007).
Photobiomodulation on the angiogenesis of skin wounds in
rats using different light sources. Photomed. Laser Surg. 25,
102–106.
30. Maffulli, N., Ewen, S.W.B., Waterston, S.W. et al. (2000).
Tenocytes from ruptured and tendinopathic Achilles ten-
dons produce greater quantities of type III collagen than
tenocytes from normal Achilles tendons an in vitro model of
human tendon healing. Am. J. Sports Med. 28, 499–505.
31. Coombs, R.R.H., Klenerman, L., Narcisi, P. et al. (1980).
Collagen typing in Achilles tendon rupture. J. Bone Joint
Surg. Br. 62, 258.
32. Enwemeka, C.S., Cohen–Korneberg, G.E., Duswalt, E.P.
et al. (1994). Biomechanical effects of three different periods
of GaAs laser photostimulation on tenotomized tendons.
Laser Therapy 6, 181–188.
33. Longo, L., Evangelista, S., Tinacci, and Sesti, A.G. (1987).
Effect of diodes–laser silver arsenide–aluminium (Ga–Al–
As) 904 nm on healing of experimental wounds. Laser Surg.
Med. 7, 444–447.
34. Vinck, E.M., Cagnie, B., Cornelissen, M.J., Declercq, H.A.,
and Cambier, D.C. (2003). Increased fibroblast proliferation
induced by light emitting diode and low power laser irra-
diation. Lasers Med. Sci. 18, 95–99.
35. Azevedo, L.H., Eduardo, F.P., Moreira, M.S. et al. (2006).
Influence of different power densities of LILT on cultured
human fibroblast growth: A pilot study. Lasers Med. Sci. 21,
86–89.
Address correspondence to:
Marco Aurelio I. Neves
Division of Plastic Surgery
UNIFESP
Rua Napolea
˜o de Barros, 715 — 4
o
andar
CEP 04024–002 Sa
˜o Paulo
Sa
˜o Paulo
Brazil
E-mail: marcoain@gmail.com
and cepinfildi@hotmail.com
668 NEVES ET AL.

This article has been cited by:
1. L M G Neves, R L Matheus, G M T Santos, M A M Esquisatto, M E C Amaral, F A S Mendonça. 2013. Effects of microcurrent
application and 670 nm InGaP low-level laser irradiation on experimental wound healing in healthy and diabetic Wistar rats.
Laser Physics 23:3, 035604. [CrossRef]
2. Murilo Xavier, Renato Aparecido Souza, Viviane Araújo Pires, Ana Paula Santos, Flávio Aimbire, José Antônio Silva, Regiane
Albertini, Antonio Balbin Villaverde. 2013. Low-level light-emitting diode therapy increases mRNA expressions of IL-10 and
type I and III collagens on Achilles tendinitis in rats. Lasers in Medical Science . [CrossRef]
3. Danillo Barbosa, Renato Aparecido Souza, Wellington Roberto Gomes Carvalho, Murilo Xavier, Paula Kariluce Carvalho, Tereza
Cristina Rodrigues Cunha, Emilia Ângela Loschiavo Arisawa, Landulfo Silveira, Antonio Balbin Villaverde. 2013. Low-level laser
therapy combined with platelet-rich plasma on the healing calcaneal tendon: a histological study in a rat model. Lasers in Medical
Science . [CrossRef]
4. Erica Martinho Salvador Laraia, Iandara Schettert Silva, Daniel Martins Pereira, Filipe Abdalla dos Reis, Regiane Albertini,
Patrícia de Almeida, Ernesto Cesar Pinto Leal Junior, Paulo de Tarso Camillo de Carvalho. 2012. Effect of Low-Level Laser
Therapy (660 nm) on Acute Inflammation Induced by Tenotomy of Achilles Tendon in Rats. Photochemistry and Photobiology
no-no. [CrossRef]