Cytotoxic Mechansims for Cholesterol Oxidation Products in Fibroblasts and Endothelial Cells

Abstract

Several cholesterol oxides form during free radical oxidation of membrane lipids. These cholesterol oxides have been recovered from tissues and at least eight of these are products consistently detected after oxy-radical induced oxidation of membrane lipids. The proportions of these products can serve as a “fingerprint” of the oxidation process in membranes. Three common cholesterol oxides have been tested for their cytotoxic potency in cell culture systems. The isomeric cholesterol 5,6 epoxides, 7-keto-cholesterol and cholestane-3β,5α,6β-triol are potent cytotoxins with remarkable stability under conditionsof cell culture. All three compounds are toxic at concentrations half of that seen with bile acids such as lithocholic, cholic or deoxycholic acids. Their toxicity appears to be directed towards the cell membrane and includes marked perturbations of calcium homeostasis. The potency of each compound follows the order: pulmonary endothelial cells > lung fibroblasts (V79 cells) > hepatocytes. The cytotoxic potency of each cholesterol oxide is reduced more than 50% in calcium deficient media (<0.05 mM) indicating the involvement of calcium in cytotoxicity. The LD50 for cholestane triol (20 uM) approaches the toxicity of linoleic acid hydroperoxides (10 uM) in endothelial cells. Although endothelial cells represent the most sensitive cell type examined for virtually all cholesterol oxides, each cell type displays approximately equal sensitivity to linoleic acid hydroperoxides. In addition to their membrane toxicity, the cholesterol epoxides also produce DNA damage which correlates with extensive DNA binding. The epoxides are also weak mutagens as demonstrated in V79 cells. A unique aspect of cholesterol epoxide toxicity is the equal LD50’s and IC50’s.(ie. 50% inhibition of DNA synthesis) in both V79 and endothelial cells, suggesting that the two events may proceed via a common mechanism. Since cholesterol epoxides are major products derived from lipid peroxidation, rapidly accumulation on treatment of membranous cholesterol with linoleic and other organic hydroperoxides, it is concievable that a number of toxic effects of lipid peroxides may be attributable to the more long-lived cholesterol oxides which are produced.