Effects of Low Level Laser Irradiation on Keratinocytes In Vitro

Low-level Laser (LLL) therapies have been used for treatment of several oral diseases, especially to improve tissue repair. Epithelial cells play an important role on healing events promoting coating of damaged tissues and stimulating underlying cells. Therefore, the application of therapies capable to increase proliferation and metabolism of epithelial cells may improve healing of wounds. Objectives: To evaluate the effects of the LLL irradiation on culture of keratinocytes. Methods: Human epithelial cells (HaCaT) were seeded (30,000 cells/cm²) in wells of 24-well dishes for 48 hours. Then, the cells were irradiated by a irradiation device - LASERTable (780 ± 3nm wavelength; 40mW). Three irradiations of cells were performed every 24 hours by application of the following energy doses: 0.5; 1.5; 3; 5; or 7J/cm². Evaluation of cell metabolism (MTT Assay), cell proliferation (Trypan Blue Assay), total protein production as well as the assessment of cell morphology by SEM were performed 24 hours after the last irradiation. Turkey test was used for pairwise comparisons and Kruskal-Wallis complemented by Mann-Whitney test was used for non-parametric data (α=0.05). Results: Despite all LLL energy doses had increased the cellular metabolism of HaCaT cells, the 0.5; 1 and 3J/cm² doses were the most effective. The highest cell proliferation rates were obtained with 0.5; 3 and 5 J/cm² energy doses evaluated. Regarding total protein production, no statistical difference occurred when irradiated and control groups were compared; in addition, no difference was also observed among all tested doses (p>0.05). The SEM images demonstrated that the energy doses used in this study did not promote any morphological alterations in the epithelial cells. Conclusion: Specific energy doses of LLL irradiation applied on cultured HaCaTt cells increase the proliferation as well as the metabolism of this kind of epithelial cell line.