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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: bayatenator@gmail.com

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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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