Determining optimal dose of laser therapy for attachment and proliferation of human oral fibroblasts cultured on titanium implant material

Abstract

The purpose of this study was to investigate the influence of single or multiple doses of low-level laser therapy (LLLT) on attachment and proliferation of human gingival fibroblasts in a standardized, reproducible in vitro model. Titanium discs were randomly allotted to one of three groups: group I served as a control, group II was exposed to a single laser dose of 3 J/cm2, and the three subgroups in group III were exposed to laser doses of 0.75, 1.5, and 3 J/cm2. To examine the possible thermal effects of laser exposure on the cell culture, the temperature in the Petri dish was measured for every dose used, before and during irradiation. For attachment assays, groups II and III were exposed to laser irradiation and then seeded onto titanium discs. In group III, the exposures were repeated after 3 and 6 h. Cells were cultured for 6 and 24 h and stained with Hoechst and Propidium. Attached cells were counted under a light microscope. To investigate the effect of LLLT on cell proliferation after 48 h, 72 h, and 7 days, cells were cultured on titanium discs for 24 h and then exposed to laser irradiation for 1 day and 3 consecutive days, respectively. Cell proliferation was determined by counting cells under the microscope and by a cell proliferation enzyme-linked immunosorbent assay system. No increase of temperature of the cell cultures occurred before or during laser exposure at any of the doses tested. Both single and multiple doses of LLLT significantly enhanced cellular attachment (p<0.05). The proliferation assays showed higher cell proliferation (p<0.05) in group III at doses of 1.5 and 3 J/cm2 after 72 h and 7 days, with agreement between staining and enzyme-linked immunosorbent assay. It is concluded that, in this cellular model, the attachment and proliferation of human gingival fibroblasts are enhanced by LLLT in a dose-dependent manner.