Influence of the laserphototherapy (LPT) on proliferation and migration of human dental pulp stem cells

Abstract

Purpose: to evaluate in vitro the influence of different parameters of laserphototherapy (LPT) on proliferation and migration of human dental pulp stem cells. First, different concentrations of fetal bovine serum (fbs) on the supplementation of the culture medium will be tested to achieve the best condition of nutritional deficit to evidence the possible effects of LPT on cell proliferation. The concentrations of 10%, 5% and 2.5% will be tested, adopting the optimum concentration (15% fbs) as positive control. Hereafter, using the nutritional deficit condition determined on previous experiment, the effects of LPT on cell proliferation and migration will be studied. The cells will be plated on 96-well tissue-culture plates and then they will be or not (controls) submitted to the LPT with the diode laser, at a wavelength of 660 nm or 808 nm; power of 20 mW and energy densities of 5J/cm² and 20J/cm² with punctual and contact mode. The cell proliferation will be based on the cell counting and on the mitochondrial activity analysis (MTT reduction-based cytotoxicity assay) at the experimental times of 24, 48, 72 and 96 hours. The cell migration will be evaluated with the "Scratch" assay in plated cultures on 24-well plates and treated or not with mitomycin. So, the confluent monolayers will be scratched with a pipette tip and, after that, submitted or not (controls) to the LPT. The closure of the wounds will be followed at the times of 0, 12, 24 and 48 hours. Finally, the data will be analyzed by the appropriate statistical test adopting the significance level of 5% (pd 0.05). (AU)