Abstract
UVA irradiation penetrates deep into the epidermis producing reactive oxygen species (ROS) that are responsible for damaging cellular components. Many phenolic compounds scavenge ROS and chelate transition metal ions that promote oxidation. In this thesis the effect of five polyphenols, namely, epicatechin gallate (ECG), luteolin, luteolin-4'-O-glucoside, luteolin-7-O-glucoside and trolox were investigated in terms of their ability to modulate UVA-mediated oxidative damage in human skin fibroblasts (HSF) at two different doses of irradiation (250 and 500 kJ/m2). The cytotoxicity of each compound was evaluated by both the MTT and LDH assays using a polyphenol concentration of 30 muM which was not cytotoxic. At both doses of UVA irradiation, an increase in apoptosis was observed. Each of the phenolic compounds tested reduced the extent of apoptosis except for luteolin-4'-O-glucoside and trolox. UVA irradiation of HSF increased lipid peroxidation (PV and MDA-TBARS) Overall, luteolin and its glucosides demonstrated the greatest protection in terms of reducing lipid peroxidation at both UVA doses. These flavones were more effective than ECG, which in turn was more protective than trolox. Trolox only reduced PV at the lower irradiation dose. UVA irradiation caused an increase in protein cross-linkage (dityrosine formation) shown by increased fluorescence, and in collagen I and IV degradation in HSF. All (poly)phenols reduced protein cross-linkage at the high UVA dose but did not prevent collagen I and IV degradation. Both luteolin and its 7-O-glucoside interacted effectively with Fe3+ and Cu2+ indicating the importance of both 3,4-dihydroxy group and the 4-oxo group on the B and C rings, respectively. Luteolin-4'-O-glucoside showed a lower interaction with the two transition metals highlighting the importance of the B ring catechol group. ECG also interacted effectively with Cu2+ but not Fe3+ whereas trolox showed the least interaction with these metal ions. These results show that UVA causes apoptosis and damage to lipids and proteins in HSF. This damage may be reduced by the inclusion of luteolin and its associated glucosides, as well as ECG, to HSF prior to irradiation. However, trolox was generally ineffective. Overall the efficacy of the (poly)phenols investigated in terms of their ability to prevent UVA induced damage was related to their structural features and their ability to interact with metal ions.