Melanin density affects photobiomodulation outcomes in cell culture

Objective: This study investigated the influence of melanin on the outcome of photoradiation at 670 nm in a cell culture model. Background Data: Melanins are naturally occurring cutaneous pigments. Human skin is classified into six skin types based on melanin content. Methods: Gelatin photo filters were fabricated with varying melanin contents. Human HEP-2 and murine L929 cell lines were cultured in complete Dulbecco's Modified Eagle's Medium (DMEM) media. Photoradiation at 670 nm delivering 5.0 J/cm(2) per treatment/24 h ( 50 J/cm(2) total fluence) was carried out with melanin filters placed between the light source and the wells using a light-emitting diode (LED) device. Five groups based on percent melanin were treated: group 1, no filter; group 2, gelatin alone; group 3, 0.0125%; group 4, 0.025%; and group 5, 0.050%. Cell proliferation was measured using CyQuant and 3-(4,5-dimethylthiazol-2-yl)-2,5-disphenyl tetrasodiumbromide (MTT) assays for 240 h post-photoradiation. Results: The Proliferation Index ( PI) as measured by CyQuant assay was not statistically different amongst the groups in either cell line. MTT assay results demonstrated a significant dose response effect ( p <= 0.05) in both cell lines with activity inversely proportional to melanin concentration. The relative PI values by MTT assay at 144 h for groups 1, 2, 3, and 4, respectively, were 1.44 +/- 0.06, 1.28 +/- 0.05, 1.20 +/- 0.07, and 1.06 +/- 0.04 for the L-929 cells, and 1.61 +/- 0.03, 1.47 +/- 0.06, 1.35 +/- 0.03, and 1.19 +/- 0.06 for the HEP-2 cells (n = 4; p < 0.05). Conclusion: These results demonstrate that cutaneous melanin content should be taken into consideration in photobiomodulation paradigms. Further studies to quantify these effects are warranted.