7 beta-hydroxycholesterol induces Ca2+ oscillations, MAP kinase activationand apoptosis in human aortic smooth muscle cells

In the present study, we characterize the early cytotoxic effects of 7 beta -hydroxycholesterol, a major cytotoxin in oxidized LDL, in human aortic sm ooth muscle cells. Within a few minutes after addition, 7 beta -hydroxychol esterol induced Ca2+ oscillations with a frequency of approximate to 0.3-0. 4 min(-1). A few hours later, thapsigargin-sensitive Ca2+ pools were deplet ed, indicating that 7 beta -hydroxycholesterol perturbs intracellular Ca2homeostasis. The mitogen-activated protein kinases (MAPKs) ERK1 and ERK2 (b ut not JNK) were activated within 5 min after addition of 7 beta -hydroxych olesterol. The side-chain hydroxylated oxysterols 25-hydroxycholesterol and 27-hydroxycholesterol were more potent in inducing apoptosis than 7 beta - hydroxycholesterol and cholesterol-5 alpha ,6 alpha -epoxide, as determined by TUNEL staining. Addition of TNF alpha. (10 ng/ml) and IFN gamma (20 ng/ ml) enhanced the cytotoxicity of oxysterols and potentiated apoptosis. The cytokines alone were not toxic to smooth muscle cells at these concentratio ns. 25-Hydroxycholesterol and 7 beta -hydroxycholesterol but not cholestero l inhibited protein synthesis at 4-8 h as determined by [S-35]methionine in corporation assay. Morphologically, oxysterol-induced cell death was charac terized by disorganization of the ER and Golgi membranes. The Ca2+ and ERK signals preceded the ultrastructural changes induced by 7 beta -hydroxychol esterol, (C) 2000 Elsevier Science Ireland Ltd. All rights reserved.