Peripherally located A431 cells are more sensitive to cell death induced by exogenous oxidative stress

Abstract

The effects of hydrogen peroxide, an oxidative agent, on the growth of A431 (an epidermoid carcinoma) cell line were investigated. It was also explored that whether or not the cell localization (peripheral or central position in a cell population) would modify the cell death-inducing effect of hydrogen peroxide. Anti-growth effect of hydrogen peroxide (0.05-1mM) on cell survival was tested by the MTT viability assay while the effect of it on DNA synthesis was measured by [3 H] thymidine incorporation assay. Cell death mode (apoptosis or necrosis) was morphologically determined by double (Hoechst dye 33342/propidium iodide) staining. Effect of hydrogen peroxide on the mitotic figures was visualized by the hematoxylin staining. F-actin fibers were stained by using fluorescein isothiocyanate (FITC)-labeled phalloidine to present the effect of hydrogen peroxide on cell cytoskeleton. It was found that hydrogen peroxide showed anti-growth effects on cells in a dose-dependent manner. It inhibited the rate of DNA synthesis at relatively lower doses (100-400 mu M), while it induced apoptosis (400-600 mu M) and necrosis (6001000 mu M) at relatively higher doses. Interestingly, the cells located at the periphery of cell population were particularly vulnerable to cell death-inducing effect of hydrogen peroxide while those at the center remained relatively unharmed. This effect was confirmed by the visualization of damaged F-actin fibers of these peripherally located cells. Hydrogen peroxide shows a cytostatic effect at relatively lower concentrations, but it is cytotoxic at higher concentrations. In addition, peripherally located cells are much more sensitive to cell death-inducing effect of hydrogen peroxide.