Iron-induced Lipid Peroxidation in Human Skin-derived Cell Lines: Protection by a Red Orange Extract
Flora Morini, Fabiola Dusatti, Francesco P. Bonina, Antonella Saija and Margherita Ferro
Although photodamage and photoprotection have already been extensively studied in cultured cells, few data have been reported in the literature regarding the in vitro behaviour of skin cells toward a chemical stress, such as iron-induced lipid peroxidation. We investigated the susceptibilities of two human skin-derived cell lines (NCTC 2544 keratinocytes and HFFF2 fibroblasts) to lipid peroxidation induced by FeSO4/histidine, FeSO4/ascorbate and Fe2(SO4)3/ADP. NCTC 2544 cells were more susceptible than HFFF2 cells to lipid peroxidation (assessed by measuring the content of malondialdehyde [MDA]) with iron/ascorbate and iron/ADP as pro-oxidants whereas, with iron/histidine, the same level of MDA production was achieved (about 10nmol/mg protein) in the two cell populations. On the basis of these results, one experimental model (iron/histidine) was selected to assess the protective effect of a mixture of two classical antioxidants, Trolox C™ (50μM) and Vitamin C (1mM), added to the cell cultures according to various protocols. The maximal decrease of MDA production in both cell lines was obtained when the antioxidant mixture and the pro-oxidant were added simultaneously to the cultures. By using the same experimental design, NCTC 2544 and HFFF2 cells were exposed to a standardised extract of red oranges (ROE; 0.025–0.5mg/ml), the main active principles of which (anthocyanins 3.1%, hydroxycinnamic acid 2.07%, and flavanone glycosides 8.1%) possess antioxidant activity. Cells treated with ROE, that were still over 90% viable, as evaluated by means of neutral red uptake and tetrazolium salt reduction tests, showed a significant and dose-dependent inhibition of MDA production. This study provides new information about the behaviour of cultured skin cells exposed to chemically induced oxidative stress, and provides further support to the possibility of using skin-derived human cell lines in the evaluation of the effectiveness of antioxidant ingredients for new drugs and/or cosmetics.