In vitro Photooxidation of Nucleic Acids by Ultraviolet A Radiation

Abstract

Although ultraviolet A radiation (UVA, 315-400 nm) has been shown to induce oxidative stress in mammalian cells and skin, the critical chromophore(s) and molecular target(s) involved have not been identified. We examined the role of oxidative damage to nucleic acids induced by UVA. To assess photooxidation of cellular DNA and RNA by UVA, we irradiated human skin fibroblasts with up to 765 kJ/m2 UVA. Cellular DNA and RNA were isolated immediately and enzymatically hydrolyzed to nucleosides. 8-Oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), a primary oxidation product in DNA, and 8-oxo-7,8-dihydroguanosine (8-oxoG), resulting from hydroxylation of guanine in RNA, were measured by HPLC with electrochemical detection. We determined that irradiation of skin fibroblasts with levels of UVA that produced moderate toxicity also resulted in significant levels of guanine hydroxylation in RNA. Lower levels of photooxidation were observed in DNA. These results suggest that measurement of guanine hydroxylation in nucleic acids, particularly in cellular RNA, may be a powerful tool for investigating the photobiological activity of UVA.