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Indian Journal of Experimental BiologyVol. 49, May 2011, pp. 357-361
Effect of low-level laser therapy on experimental wounds of hard palate
mucosa in mice
Farahnaz Fahimipour
Shahid Beheshti University MC, Tehran, Iran
Mohsen Nouruzian
Department of Anatomy and Biology, Shahid Beheshti University MC, Tehran, Iran
Morteza Anvari.1, Mahmood Akhavan Tafti
.2& Mehdimalek Yazdi
.3
Anatomy1, Pathology2& Medical Engineering3DepartmentShahid Sadoghi University of Medical Science, Yazd, Iran
Mahsa Khosravi & Zahra DehghannayeriMedical Faculty, Shahid Sadoghi University of Medical Faculty, Yazd, Iran
Shabnam S Sabounchi
Dental Faculty, Shahid Beheshti University MC, Tehran, Iran
and
Mohammad Bayat *
Department of Anatomy and Biology, Medical Faculty, Shahid Beheshti University MC, Tehran 1985717443, Iran
Received 8 November 2010; revised 9 February 2011
Under general anesthesia and sterile conditions, incision wound was induced in the hard palate mucosa of adult male
mice. The wounds of groups 1 and 2 were irradiated daily with He-Ne laser at 3 and 7.5 J/cm2 for 120 and 300 s,respectively, while the incision wound of group 3 not exposed served as controls. On day 3 of injury, the laser-treatedwounds contained significantly lower neutrophils than the wounds in the control group. By day 7 after injury, the laser-treated wounds contained significantly more fibroblasts and at the same time contained significantly fewer macrophages. In
conclusion, an acceleration of the wound healing process of experimental wounds in the hard palate mucosa of mice at low-level laser therapy with a He-Ne laser at energy densities of 3 and 7.5 J/cm2was observed.
Keywords: Hard palate mucosa, Low-level laser therapy, Wound healing
Craniomaxillofacial injuries occur in a significant
proportion of trauma patients, either in isolation or in
combination with other serious injuries1. Transverse
maxillary deficiencies are relatively common
dentofacial deformity in combination with other
maxillary and/or mandibular deformities2. Orofacial
clefts are defined as congenital defects in which thefusion between two or more of the following
structures have failed: the palatal shelves, the
maxillary prominences, and the medial nasal
prominences. The clefts are surgically closed to
restore the integrity of the oral and nasal cavity and to
allow for normal feeding and speech development3.
Healing of these wounds is associated with
the disadvantageous effects in maxillary growth
and dento-alveolar development often seen in cleft
palate patient3-5.
Low-level laser therapy (LLLT) has been used for
more than 25 years in clinical practice and is knownto modulate various biological processes6-9
. LLLT is a
non-thermal modality; usually the temperature
changes associated with treatment are negligible10
. Anumber of different laser light sources, including
helium-neon, ruby, and gallium arsenide have been
used to deliver LLLT in different treatments and
schedules.
In patients, biostimulation with low-level laser
radiation has accelerated the healing of gingival
incisions and gingivectomy11-13
, reduced the severity
*Correspondent authorTelephone/Fax: 98 21 22439949.
E-mail: [email protected]
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INDIAN J EXP BIOL, MAY 2011358
of conditioning-induced oral mucositis in bonemarrow transplantation patients14. Low-level laser
radiation induced formation of new periodontal
ligament, cementum, and bone
in dogs
15
, and alsoimproved macromolecular clearance via the lymph
flow in the hamster gingiva16
. At the cellular level,studies on cultured human gingival fibroblasts and
human oral mucosal fibroblasts have suggested a
number of LLLT effects: increased DNA synthesis17,
induced rapid generation of myofibroblasts from
fibroblasts7, enhanced proliferation
18,19 and attachment
of human gingival fibroblasts20.
The effects of LLLT on the gingiva have been
studied quite extensively in vivoand in vitro; however
there is a dearth of data on the effect of LLLT on the
mucosa of soft and hard palates
14
. Barasch et al.
14
irradiated He-Ne laser to patients' oral mucositis,
including the mucosa of the soft palate. Oralmucositis and pain scores were significantly lower for
the treated versus the untreated side. They concluded
that He-Ne laser treatment was well tolerated and
reduced the severity of oral mucositis in patients14
. Be
that as it may, clinical trial studies have failed to showbeneficial effects from LLLT on wound healing after
gingivoplasty in humans21 and also have failed to
show beneficial effects from LLLT on the protein
synthesis of human gingival fibroblasts in vitro22
.
The use of LLLT has still not been widely acceptedby physicians and a number of controlled studies13
,
and the FDA withdrew its approval for its therapeutic
use in the USA in 198323.
The present study sought to address the problem by
evaluating whether He-Ne laser radiation at energy
densities of 3 and 7.5 J/cm2accelerated the healing of
the experimental wounds of the hard palate mucosa in
mice.
Materials and Methods
Ninety adult male Albino N mari mice (3 months
old, weighing 6010g) were used. The study protocolwas approved by the Medical Ethics Committees ofShahid Sadoghi Medical University,Yazd, Iran. The
mice were provided with a standard animal food and
water ad libitum. On day zero, all the mice were
anesthetized with 50 mg/kg ketamine hydrochloride
injected intramuscularly along with 5 mg/kg
diazepam. In the hard palate mucosa, 2 mm posterior
to the incisive tooth of the upper jaw a 4-mm long and
2-mm deep incision wound was made under sterile
conditions. The animals were randomly divided into
three groups, of 30 each. The incisions in groups 1and 2 were treated daily with a He-Ne laser tube with
3 and 7.5J/cm2 energy densities, respectively. LLLT
was commenced 24 h after surgery under generalanesthesia. The laser used was a He-Ne laser
(Meredith Co, Turkey) rated at 25 mW of power.
Groups 1 and 2 were exposed to 120 and 300s of
laser light, respectively. LLLT was applied to one
point, covering the whole length of the incision in
each session (day). The laser was held 1 cm away
from the surface of the target tissue.
The incision wounds of group 3 were treated with
the LLLT probe without the laser being switched on,
and the group was thus considered the control group.
Thereafter, 10 mice of each group were sacrificed 3
days after surgery, 10 mice of each group weresacrificed 7 days after surgery, and the remaining 10
mice were sacrificed on the 15 th postoperative day.
The mice were sacrificed via the inhalation of
chloroform in a closed space.
For histological examinations, a sample was
excised from the wound bed and the normal adjacent
mucosa of each mouse and was fixed in formaldehyde
saline before it was embedded in paraffin blocks.
Sagittal sections (5 m thickness) were cut and stainedwith the hematoxylin and eosin method. Ten zones
from each sample were examined morphometrically
using a calibrated ocular on a Nikon light microscopeat a magnification of 400X for counting the fibroblasts,
macrophages, neutrophils, and endotheliums of theblood vessels. The histological examinations were
performed in a blind fashion.
The data were subjected to the one way analysis of
variance (ANOVA) and were expressed as mean
SE. Multiple comparisons were performed by the
least significant difference (LSD) test. Pvalues
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FAHIMIPOUR et al.: LASER THERAPY OF WOUNDS OF HARD PALATE MUCOSA 359
mean number of neutrophils in group 3 wassignificantly higher than those of the other groups
(P
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INDIAN J EXP BIOL, MAY 2011360
midline. The wounds on the left side were irradiateddaily with He-Ne laser at 4J/cm2, whereas the wounds
on the right side were not exposed and thus served as
controls. Early epithelialisation with increasedfibroblastic reaction and neovascularization was seen
in the lasertreated wounds. Wound histology alsoshowed a significant increase in leukocyte infiltration
until day 9. Bisht et al.6reported that their results had
established the biostimulatory effects of low-intensity
laser radiation on the healing of the skin wound.
Increased leukocytic reaction, reported by Bisht et al.,
is in disagreement with decreased neutrophil and
macrophage counts, observed in the present study.
The literature, however, shows that
photobioactivation accelerates inflammation and
promotes fibroblast proliferation, and even accelerates
the healing process26.
Recent investigations using advanced experimentaltechniques have tried to define the precisebiostimulatory effects of LLLT. Recently, anupregulation of genes leading to an inverse in theproliferation of laser-treated fibroblasts has beendemonstrated
27.
Irradiation of human fibroblasts at 628 nm wasreported to have caused an increase in 7 categories ofgenes already known to play roles in the enhancementof cell proliferation and suppression of apoptosis
27.
Another related study succeeded in demonstrating in
real time that LLLT could cause intracytosolicincrease in pH and calcium as well as changes inreactive oxygen species
28.
These finding have laid a new groundwork forattempting to understand the comprehensive effects ofLLLT on the biochemical milieu of the injured tissue29.
To conclude, acceleration in the healing ofexperimental wounds in the hard palate mucosa ofmice at LLLT with He-Ne laser at energy densities of3 and 7.5 J/cm2was observed.
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