Toxicity of cholesterol oxidation products to Caco-2 and HepG2 cells: modulatory effects of α- and γ-tocopherol

Cholesterol can be oxidized to form a variety of cholesterol oxidation products also known as oxysterols. The aims of the present study were to compare the cytotoxic effects of four oxysterols, namely 25‐hydroxycholesterol (25‐OHC), 7β‐hydroxycholesterol (7β‐OHC), cholesterol‐5β,6β‐epoxide (β‐epox) and cholesterol‐5α,6α‐epoxide (α‐epox), in two human cell culture models. Further, the ability of 10 and 100 µm α‐ and γ‐tocopherol (α‐TOC and γ‐TOC, respectively) to protect against oxysterol‐induced cytotoxicity was also assessed. Human colonic adenocarcinoma Caco‐2 and human hepatoma HepG2 cells were supplemented with increasing concentrations of 25‐OHC, 7β‐OHC, β‐epox and α‐epox (0–25 µg ml−1) for 24, 48 or 96 h. Following 24‐h and 48‐h exposure, test media were replaced with normal growth media and the cells were maintained for 72 and 48 h, respectively. The 96‐h exposure represented a constant challenge to the cells. Cytotoxicity was assessed using the neutral red uptake assay. The concentration of compound that inhibited cell viability by 50% (ic50 value) was calculated. All four oxysterols investigated induced the greatest cytotoxic effects following 96 h of exposure. 25‐Hydroxycholesterol exhibited the greatest cytotoxicity in both cell lines. Both β‐epox and α‐epox were more toxic to HepG2 cells than to Caco‐2 cells after the 48‐h exposure. Pretreatment of cells with either α‐ or γ‐TOC did not protect against oxysterol‐induced cytotoxicity. The caco‐2 cells treated with the high concentration (100 µm) of γ‐TOC were found to be more susceptible to oxysterol‐induced toxicity under the conditions employed in this study. Copyright © 2003 John Wiley & Sons, Ltd.