Photo-induced DNA damage and photocytotoxicity of retinyl palmitate and its photodecomposition products

Abstract

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 mM), DNA fragmentation was observed. Cell death was observed for retinoid concentrations of 100 mM or higher. When treated with 150 mM 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 FX174 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.

Keywords

5,6-epoxy-retinyl palmitate anhydroretinol Comet assay DNA strand cleavage retinyl palmitate

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References

Ahmad, H. and Mukhtar, H. 2004: Toxicology of the skin: new and emerging concepts. Toxicology and Applied Pharmacology 195, 265-266.

Basu-Modak, S. and Tyrrell, R. M. 1993: Singlet oxygen: a primary effector in the ultraviolet A/near-visible light induction of the human heme oxygenase gene. Cancer Research 53, 4505-4510.

Boehnlein, J. , Sakr, A. and Lichtin, J. L. 1994: Characterization of esterase and alcohol dehydrogenase activity in skin. Metabolism of retinyl palmitate to retinol (vitamin A) during percutaneous absorption. Pharmaceutical Research 11, 1155-1159.

Cherng, S.-H. , Xia, Q. , Blankenship, L. R. et al. 2005: Photodecomposition of retinyl palmitate in ethanol by UVA light-formation of photodecomposition products, reactive oxygen species, and lipid peroxides. Chemical Research in Toxicology 18, 129-138.

Cosmetic, Toiletry and Fragrance Association ; Wenninger, J. , Canterbery, R. C. and McEwen, G. N. Jr. , editors, 1999: International cosmetic ingredient dictionary and handbook, eighth edition. Washington, DC: CTFA, 1279-1279.

Data from the US Food and Drug Administration’s Voluntary Cosmetics Registration Program (2000).

Dong, S. , Hwang, H.-M. , Shi, X. et al. 2000: UVA-induced DNA single-strand cleavage by 1-hydroxypyrene and formation of covalent adducts between DNA and 1- hydroxypyrene. Chemical Research in Toxicology 13, 585-593.

Duell, E. A. , Kang, S. and Voorhees, J. J. 1997: Unoccluded retinol penetrates human skin in vivo more effectively than unoccluded retinyl palmitate or retinoic acid. The Journal of Investigative Dermatology 109, 301-305.

Forbes, P. D. 1981: Photocarcinogenesis: an overview. The Journal of Investigative Dermatology 77, 139-143.

10.

Fu, P. P. , Howard, P. C. , Culp, S. et al. 2002: Do topically applied skin creams containing retinyl palmitate affect the photocarcinogenicity of simulated solar light? Journal of Food and Drug Analysis 10, 262-268.

11.

Fu, P. P. , Cherng, S.-H. and Coop, L. 2003: Photoreaction, phototoxicity, and photocarcinogenicity of retinoids. Journal of Environmental Science and Health. Part C, Environmental Carcinogenesis & Ecotoxicology Reviews C21(2), 165-197.

12.

International Agency for Research on Cancer . 1992: Solar and ultraviolet radiation. IARC Monographs on the Evaluation of Carcinogenic Risks to Humans, Volume 55, IARC.

13.

International Agency for Research on Cancer . 1998: Handbooks of cancer prevention, vitamin A, Volume 3, Lyon, France: IARC.

14.

Ihara, H. , Hashizume, N. , Hirase, N. et al. 1999: Esterification makes retinol more labile to photolysis. Journal of Nutritional Science and Vitaminology 45, 353-358.

15.

Mukhtar, H. , editor, 1995: Skin cancer: mechanisms and human relevance. Boca Raton, FL: CRC Press.

16.

Sortino, S. , Condorelli, G. and de Guidi, G. 1998: Molecular mechanism of photosensitization XI. Membrane damage and DNA cleavage photoinduced by Enoxacin. Photochemistry and Photobiology 68, 652-659.

17.

Tice R. R. , Agurell E. , Anderson D. et al. 2000: Single cell gel/Comet assay: guidelines for in vitro and in vivo genetic toxicology testing. Environmental and Molecular Mutagenesis 35, 206-221.

18.

Wang, L , Yan, J. , Cohly, H. et al. 2004: Photo-acute toxicity and genotoxicity of azulene on human Jurkat T-cells. Mutation Research 562, 143-150.