Lasers in Surgery and Medicine 29:179–184 (2001) Comparison of the Low Level Laser Therapy Effects on Cultured Human Gingival Fibroblasts Proliferation Using Different Irradiance and Same Fluence Luciana Almeida-Lopes, DDS, MSD, 1,2 * Josepa Rigau, MD, PhD, 3 Renato Amaro Zaˆ ngaro, PhD, 1 Joa˜ o Guidugli-Neto, MD, PhD, 4 and Ma´ rcia Martins Marques Jaeger, DDS, PhD 4 1 Institute for Research and Development, Universidade Vale do Parai ´ba-SJC, Brazil 2 School of Dentistry, University Camilo Castelo Branco-SP, Brazil 3 Department of Histology, School of Medicine, University Rovira i Virgili, Reus, Spain 4 Department of Oral Pathology, School of Dentistry, University of Sa˜o Paulo-SP, Brazil 05508–900 Background and Objective: The low level laser therapy (LLLT) has been used in Dentistry to improve wound healing. In order to analyse the effect of LLLT on the in vitro proliferation of gingival fibroblasts we developed a primary culture of human gingival fibroblasts. Study Design/Materials and Methods: The cell line named LMF was grown in Dulbecco’s Modified Eagle’s medium (DME) with either 5% (nutritional deficit) or 10% fetal bovine serum (fbs). Laser irradiation was carried out with diode lasers with the following wavelengths: 670 nm (L1), 780 nm (L2), 692 nm (L3), and 786 nm (L4). The fluence was fixed in 2 J/cm 2 . For growth analysis, control (not irradiated) and treated cultures (irradiated) were plated in 60 mm diameter culture dishes for 12 h before the irradiation. Results: We found that cells cultured in nutritional deficit condition grown in medium supplemented by only 5% fbs presented a cell proliferation rate significantly smaller that cell grown in ideal culture conditions (10% fbs). However, when irradiated, cells in nutritional deficit presented cell growth similar or higher than that of control cells grown in ideal culture conditions. Using the same fluence, the infrared laser induced a higher cell proliferation than visible laser when the power outputs were different. However, lasers of equal power output presented similar effect on cell growth independently of their wavelengths. Conclusions: The LLLT acts by improving the in vitro fibroblast proliferation and a smaller laser exposure time results in higher proliferation. Lasers Surg. Med. 29:179– 184, 2001. ß 2001 Wiley-Liss, Inc. Key words: cell culture; diode laser; human fibroblast; wound healing INTRODUCTION The low level laser therapy (LLLT) has been used in Dentistry to improve wound healing. However, most of the studies on the effects of LLLT on wound healing were undertaken in skin [1–11]. These studies showed the laser effects on wound such as increasing cellular metabolic processes, increasing the regenerative potential of the biological tissues, increasing neovascularizationl and formation of regenerative tissue [1–11]. However, there are a lesser number in mucous membranes [11–16]. Additionally, these former studies were generally carried out in animal [15]. More recently, in vitro studies of laser effects have been performed [17–29]. The greater advantage of this kind of study is that one can isolate a specific part of a determinate process [30,31]. The vast majority of in vivo studies were carried out either on established cell lines or on primary- cultured skin fibroblasts [27,28,32–40]. However, only a small number of studies were done with cell lines derived from human mucous membranes [11,17,18,41–43]. It is known that lasers with different wavelengths produce different effects on fibroblasts. In the literature, the studies comparing the effects of laser using visible and near-infrared wavelengths showed differences most of them indicating that the best results were obtained when visible wavelength was used [27–29,33,34,41,43,44]. Two other aspects of LLLT on fibroblasts are related to the dosage and culture conditions. Lower fluence rates present no significant effect, whereas high fluence rates present inhibitory effect. An ideal fluence to obtain response using fibroblasts in vitro is around 2 J/cm 2 [5,11,24,25,40,43,45–49]. Additionally, the response of the tissue in vivo to the LLLT is directly correlated to stress conditions, thus it is lacking when applied to healthy tissue [32,43,50,51]. The aim of this work was to study the effect of LLLT on fibroblasts primary cultured from human oral mucosa. We compared the effect on cell proliferation of visible lasers vs. infrared lasers, keeping the fluence constant at 2 J/cm 2 and using different irradiance. We believe that the results of this study will be of clinical relevance helping the This work was carried out at School of Dentistry, University of Sa˜o Paulo, Sa˜ o Paulo, Brazil. Contract grant sponsor: Fundac¸a˜o de Amparo a` Pesquisa do Estado de Sa˜ o Paulo (FAPESP), Sa˜ o Paulo, Brazil. Contract grant sponsor: Conselho Nacional de Pesquisa (CNPq), Brazil. *Correspondence to: Luciana Almeida-Lopes, Rua Tiba˜es, 22, 02525-030 Sa˜o Paulo, SP, Brazil. E-mail: lal@apcd.org.br Accepted 13 February 2001 ß 2001 Wiley-Liss, Inc.