Comparative Effects of Two Different Doses of Low-Level Laser Therapy on Wound Healing Third-Degree Burns in Rats PATRICIA BRASSOLATTI, 1 * PAULO S  ERGIO BOSSINI, 1 MARIA CAROLINA DER ^ ENCIO OLIVEIRA, 2 HUELITON WILIAN KIDO, 3 CARLA ROBERTA TIM, 3 LUCIANA ALMEIDA-LOPES, 4 LUCIMAR RETTO DA SILVA DE AV  O, 5 FERNANDO M. ARA  UJO-MOREIRA, 6 AND NIVALDO ANTONIO PARIZOTTO 1 1 Department of Physiotherapy, Post-Graduate Program of Biotechnology, Federal University of S~ ao Carlos (UFSCar), S~ ao Carlos, SP, Brazil 2 Department of Physiotherapy, Graduate of Physiotherapy, Federal University of S~ ao Carlos (UFSCar), S~ ao Carlos, SP, Brazil 3 Department of Biosciences, Federal University of S~ ao Paulo (UNIFESP), Santos, SP, Brazil 4 DMC Equipment Import and Export- Co. Ltda, S~ ao Carlos, SP, Brazil 5 Department of Medicine, Federal University of S~ ao Carlos (UFSCar), S~ ao Carlos, SP, Brazil 6 Department of Physics, Post-Graduate Program of Biotechnology Federal University of S~ ao Carlos (UFSCar), S~ ao Carlos, SP, Brazil KEY WORDS burns; wound healing; low-level laser therapy ABSTRACT Burns are injuries caused by direct or indirect contact to chemical, physical, or bio- logical agents. Low-level laser therapy (LLLT) is a promising treatment since it is low-cost, non- invasive, and induces cell proliferation. This study aimed to investigate the effects of LLLT (660 nm) at two different fluences (12.5 J/cm 2 and 25 J/cm 2 ) per point of application on third-degree burns in rats. Thirty rats (Wistar) divided into GC, GL12.5, and GL25 were used in the study, and submitted to burn injury through a soldering iron at 1508C, pressed on their back for 10 s. LLLT was applied immediately, and 2, 4, 6, and 8 days after wound induction. Histological analysis revealed a decreased inflammatory infiltrate in the group treated with 25 J/cm 2 , and intense inflammatory infiltrate in the control group and in the group treated with 12.5 J/cm 2 . The immunostaining of COX-2 was more intense in the control groups and in the group treated with 12.5 J/cm 2 than in the group treated with 25 J/cm 2 . Conversely, VEGF immunomarking was more expressive in the group treated with 25 J/cm 2 than it was in the other two groups. Therefore, our findings suggest that the use of 25 J/cm 2 and 1 J of energy was more effective in stimulating the cellular processes involved in tissue repair on third-degree burns in rats by reducing the inflammatory phase, and stimulating angiogenesis, thus restoring the local microcirculation which is essential for cell migration. Microsc. Res. Tech. 79:313–320, 2016. VC 2016 Wiley Periodicals, Inc. INTRODUCTION Burns are caused by direct or indirect contact to chemical, physical or biological agents (de Moraes et al., 2013; Turatti Pessolato et al., 2011) and are asso- ciated with tissue damage, infection, pain, and even death (Ezzati et al., 2009; Renno et al., 2011). Burn lesions are classified according to depth and extension, and are divided into first, second and third degrees (Souza et al., 2014). Although third-degree injuries are considered less common than other forms of traumatic injuries, they are classified as the most serious lesions for they involve the total destruction of epidermis and dermis, and induce high physiological stress in the body (Fiorio et al., 2014; Vasheghani et al., 2008). It is estimated that approximately 1 million burns happen annually in the USA, and therefore they are associated with considerable socioeconomic problems due to high treatment costs (Renno et al., 2009). Among the treatment approaches which have been presented in recent years, such as grafts (Shevchenko, James, James; 2010), biological dressings (Saska et al., 2011), and collagen-based dressings (Braziulis et al., 2012), low-level laser therapy (LLLT) is highlighted (Ezzati et al., 2009; Fiorio et al., 2013; Renno et al., 2009). Laser action is based on absorption of laser light by tissue specific photoreceptors, among which the most known is cytochrome c oxidase. After light absorption, important changes in cellular metabolism are observed. Firstly, an increase in cellular ATP produc- tion which happens due the stimulus to the electron transport chain that is inside the mitochondria is observed. Immediately afterwards, there is an increase in DNA and RNA synthesis, which then acts directly on the cell cycle regulation and favors protein synthe- sis, proliferation cell (Avci et al., 2013; Karu, 2010; Novaes et al., 2014; Renno et al., 2009), the angiogene- sis process (Bossini et al., 2009; Colombo et al., 2013), *Correspondence to: Patricia Brassolatti, University of S~ ao Carlos (UFSCar), Rod. Washington Luis, Km 235., CEP: 13565-905, S~ ao Carlos, S~ ao Paulo, Brazil. E-mail: patty.brassolatti@gmail.com Received 5 July 2015; accepted in revised form 17 January 2016 REVIEW EDITOR: Prof. Alberto Diaspro Contract grant sponsors: Coordination for the Improvement of Higher Level Education (CAPES) and the Research and Education Center for Photo Therapy in Health Science (NUPEN). DOI 10.1002/jemt.22632 Published online 8 February 2016 in Wiley Online Library (wileyonlinelibrary.com). V VC 2016 WILEY PERIODICALS, INC. MICROSCOPY RESEARCH AND TECHNIQUE 79:313–320 (2016)