Effect of vitamins A, C and E on normal and HPV-immortalized human oral epithelial cells in culture

Abstract

There is experimental and epidemiological evidence that antioxidant vitamins can inhibit carcinogenesis. Since immortalization by Human Papilloma Virus (HPV) is one possible early step towards carcinogenesis in oral epithelia, we studied the differential effect of vitamins A, C and E on HPV-immortalized oral epithelial cells (IHGK) as compared to the normal counterpart. The dose response was determined by morphology, cell cycle by flow cytometry, and growth curve by cell number. The optimum dose in terms of inhibitory effect vs. toxicity was determined for each vitamin by morphology. Optimum doses were: vitamin A - 1.4 x 10 -5 M, vitamin C - 10 -3 M, and vitamin E - 10 -6 M for both HPV-immortalized and normal cells. Growth curve showed reduction of proliferation by all three vitamins, with vitamins A and E more effective than C for both cell types. Flow cytometry showed that vitamins A and E reduced the percentage of cells at G2 phase of cell cycle and indicated arrest in the S phase. This effect was greatest in the immortalized cells with a 50% and 35% decrease of G2 for vitamins A and E respectively, whereas the normal counterpart showed a 48% decrease for A and a 12% increase for E. By organotypic culture, the morphology was not markedly different between the vitamin-treated and the control cells, except for a slight increase in the keratinization of normal cells with vitamin A. Also noted was a reduction in number of cell layers from five layers or more for controls to only one or two for vitamin E. In conclusion, we have demonstrated that the antioxidant vitamins inhibit proliferation, and show a preferential effect on IHGK cells.