80 - Lipofuscin Generated by UVA Exposure makes Human Skin Keratinocytes Sensitive to Visible Light

Abstract

The effects of UV radiation on skin cells are well known, while the effect of other fractions of the solar spectrum remains ignored. Recently, we observed that both neonatal primary normal human keratinocyte (NHK) and in immortalized human keratinocytes (HaCaT) cells exposed to UVA and after exposure to visible light had reduction in cell viability, increasing level of redox misbalance as monitored by the increase in DCF fluorescence, and the generation of singlet oxygen. We also detected strand breaks of nuclear DNA and premutagenic Fpg and Endo III-sensitive DNA lesions by Comet assay. Our data showed that UVA radiation (λ= 366 nm, 12 J.cm-2) generates lipofuscin, which is an autofluorescent visible-photosensitizer and a subproduct of incomplete lysosomal digestion of oxidized biomolecules, organelles, and membranes. Visible light photosensitizes lipofuscin inducing photooxidatives processes through the generation of several oxidizing species, which can damage DNA and generate premutagenic lesions, as 8-oxo-dG. Lipofuscinogenesis was also shown to occur more effectively through specific oxidative damages in lysosomes and mitochondria by with photosensitization of 1,9-dimethyl methylene blue (DMMB) 10 nM with red light (λ= 633 nm, 11 J.cm-2). We have then characterized the lipofuscin from human skin keratinocytes by fluorescence lifetime imaging and fluorescence spectroscopy (fluorescence lifetime = 2 ns; excitation at 450 nm and emission at 640 nm). Concluding, UVA and visible light can be dangerous to skin health, raising the importance of not only protecting ourselves against UV radiation, but also against visible light.