Retinyl palmitate (RP) is an
ester of
retinol (
vitamin A) and the predominant form of
retinol found endogenously in the skin. We have previously reported that photoirradiation of RP with UVA light resulted in the formation of
anhydroretinol (AR),
5,6-epoxyretinyl palmitate (5,6-epoxy-RP) and other photodecomposition products. While AR was formed through an ionic photodissociation mechanism, 5,6-epoxy-RP was formed through a light-mediated,
free radical-initiated chain reaction. In the current study, the
phototoxicity of RP, AR and 5,6-epoxy-RP in human skin Jurkat T-cells with and without light irradiation was determined using a
fluorescein diacetate assay. Under similar conditions, the Comet assay was used to assess damage to cellular
DNA. Nuclear
DNA was not significantly damaged when the cells were irradiated by UVA plus visible light in the absence of a
retinoid; however, when the cells were illuminated with UVA plus visible light in the presence of either RP, 5,6-epoxy-RP or AR (50, 100, 150 and 200 microM), DNA fragmentation was observed. Cell death was observed for
retinoid concentrations of 100 microM or higher. When treated with 150 microM of RP, 5,6-epoxy-RP or AR, cell death was 52, 33 and 52%, respectively. These results suggest that RP and its two photodecomposition products, AR and 5,6-epoxy-RP, induce DNA damage and cytotoxicity when irradiated with UVA plus visible light. We also determined that photoirradiation of RP, AR and 5,6-epoxy-RP causes single strand breaks in supercoiled phi chi 174 plasmid
DNA. Using a constant dose of UVA light (50 J/cm2), the level of DNA cleavage was highest in the presence of AR, followed by 5,6-epoxy-RP, then RP. The induced
DNA strand cleavage was inhibited by
NaN3. These results suggest that photoirradiation of RP, 5,6-epoxy-RP and AR with UVA light generates
free radicals that initiate
DNA strand cleavage.