A new role of phase I and phase II enzyme in keratinocytes UVB induced apoptosis.

A new role of Phase I and Phase II enzymes in keratinocytes UVB induced apoptosis.UVB is the major cause for cutaneous malignancies in the human population. The skin is able to activate anti-oxidants and enzymatic detoxification reactions to neutralize reactive photochemical products. Apoptosis, removing irreversibly DNA-damaged and potentially neoplastic cells, represents a major defence mechanism towards malignant transformation. Very little is known about the role of cutaneous cytochrome P450 (CYP450) isoenzymes and glutathione S-transferase (GST) in UVB response. Although not yet fully investigated, it has been studied a possible relationship between CYP450 inductors and UVB initiated apoptosis in rat hepatocytes. Altered high levels of GST has been directly correlated to resistance to chemotherapeutic drugs suggesting that GST plays a role in prevention of apoptosis. Our study is focused on phase I and phase II enzyme activities in normal human keratinocytes. In the first part of the study we demonstrated that CYP450 (1A1 and 2B1) and GST are induced not only by classical inducers such as \u3b2-naphthoflavone, 3-methylcholanthrene, phenobarbital but also by UVB radiations (50 mJ/cm2). Differentiated keratinocytes were employed for all the experiments as confirmed by immunoblotting with Cytokeratin 10, a specific marker of the basal spinous transition. In the second part we evaluated a possible involvement of these enzyme in UVB mediated apoptosis process. Western blot analysis of Bcl2 expression and PARP cleavage showed that inhibition of CYP450 by Proadifen prevented UVB induced apoptotic cell death. In contrast, the diuretic drug, ethacrinic acid, a GST inhibitor, was able to improve UVB induced apoptosis. In order to confirm the anti-apoptotic activity displayed by GST, when GST activity was increased by Phenobarbital, UVB apoptosis was prevented.These results suggest that UVB radiations may play a critical role as tumour promoters even through the regulation of CYP450 and GST metabolising enzymes