Co-mutagenic and pro-oxidant effects of Vitamin E promotes DNA damage and cell transformation in prostate

Introduction The dietary supplements use has increased over the past years in the global landscape and vitamins are by far the products which record the greater appeal. Following the well-rooted idea that the vitamin E promotes health status, many of people assume it not to treat diagnosed deficiencies, but to prevent new occurrences of several diseases, including cancer. However, recent trials and meta-analysis showed no significant effects against cancer aetiology, whereas data from SELECT study reported an increased risk of prostate cancer among healthy men. To date, the mechanism underlying these observations is largely unknown. In this work, we show in in vitro and in vivo models, how vitamin E through co-mutagenic and pro-oxidant mechanisms promotes both DNA damage and cell transformation frequency. Materials and Methods In vitro models Human prostate cells RWPE-1 were exposed to α-tocopherol (100 μM) for 48 h. The gene expression of COX2, CYPA1, CYPA4, CYPB6, CYPC9, and CYP4F2 gene was studied. PGE2 were determinate by ELISA test. The measure of reactive oxygen species (ROS) was carried out by the use of the Electron Paramagnetic Resonance (EPR) radical-probe technique. Human fibroblasts IMR90 were exposed to α-tocopherol (100 μM) for 48 h then cells were fixed, stained with DAPI. Micronuclei were scored. IMR90 cultured as previously described were employed for the Immunofluorescence determination of 53BP1 and γH2AX foci. BALB/c 3T3 cells were treated with α-tocopherol (100 μM), after 48 h exposition the medium was removed and cells were incubated with benzo[a]pyrene with a final concentrations ranged from 1 µg/ml to 0.01 µg/ml. Cells were maintained in culture for 4 weeks with twice weekly medium changes, then fixed and stained. Positive transformation foci were counted. In vivo model Vitamin E was administered to Sprague-Dawley rats at doses of 100 or 200 mg/kg b.w. dose daily for 7 or 14 consecutive days. Controls received vehicle (corn oil). Prostate was resected and the CYP1A1 CYP1A2, CYP1B1 gene expression was studied along with the catalytic activity through specific probes. ROS yields was measured by EPR spectroscopy. Finally, the malonyldialdehyde (MDA) development, as a marker of lipid peroxidation as well as protein carbonylation were assessed. Results Here we show that in RWPE-1 cell line vitamin E produces an overexpression of various phase-I bioactivating cytochrome P450 (CYP) isoforms, including activators of polycyclic aromatic hydrocarbons (PAHs), such as CYPA1, CYPA4, CYPB6, CYPC9, and CYP4F2 coupled with a significant yield of radical oxygen species (ROS). These observations have been supported by an in vivo model; in vitamin E supplemented rats we reported a significant booster effect on CYP superfamily along with oxidative damage to lipids and proteins. Furthermore, we show that vitamin E causes DNA damage as exemplified through micronuclei and H2AX - 53BP1 foci based-assays. Finally, we find that the pre-exposure of BALB/c 3T3 cells with vitamin E significantly increase the transformation frequency of the PAH-prototype benzo[a]pyrene. Discussion and Conclusions Taken together, these results can contribute to explain the increased cancer risk found in the SELECT trial. Our data support the hypothesis that vitamin E could act as a co-mutagenic/co-carcinogenic agent increasing the bioactivation of chemicals joint with its pro-oxidant activity resulting in higher risk of cancer through DNA insults and epigenetic alterations. Our study point out one more time the potentially harmful effects of recommending supplementations with micronutrient as a chemopreventive strategy on a large scale.