7beta-hydroxycholesterol induces Ca(2+) oscillations, MAP kinase activation and apoptosis in human aortic smooth muscle cells.
In the present study, we characterize the early cytotoxic effects of 7beta-hydroxycholesterol, a major cytotoxin in oxidized LDL, in human aortic smooth muscle cells. Within a few minutes after addition, 7beta-hydroxycholesterol induced Ca(2+) oscillations with a frequency of approximately 0.3-0.4 min(-1). A few hours later, thapsigargin-sensitive Ca(2+) pools were depleted, indicating that 7beta-hydroxycholesterol perturbs intracellular Ca(2+) homeostasis. The mitogen-activated protein kinases (MAPKs) ERK1 and ERK2 (but not JNK) were activated within 5 min after addition of 7beta-hydroxycholesterol. The side-chain hydroxylated oxysterols 25-hydroxycholesterol and 27-hydroxycholesterol were more potent in inducing apoptosis than 7beta-hydroxycholesterol and cholesterol-5alpha,6alpha-epoxide, as determined by TUNEL staining. Addition of TNFalpha (10 ng/ml) and IFNgamma (20 ng/ml) enhanced the cytotoxicity of oxysterols and potentiated apoptosis. The cytokines alone were not toxic to smooth muscle cells at these concentrations. 25-Hydroxycholesterol and 7beta-hydroxycholesterol but not cholesterol inhibited protein synthesis at 4-8 h as determined by [35S]methionine incorporation assay. Morphologically, oxysterol-induced cell death was characterized by disorganization of the ER and Golgi membranes. The Ca(2+) and ERK signals preceded the ultrastructural changes induced by 7beta-hydroxycholesterol.
Ares
M P
MP
Wallenberg Laboratory, University Hospital MAS, Lund University, S-20502, Malmö, Sweden. mikko.ares@medforsk.mas.lu.se
Pörn-Ares
M I
MI
Moses
S
S
Thyberg
J
J
Juntti-Berggren
L
L
Berggren
P
P
Hultgårdh-Nilsson
A
A
Kallin
B
B
Nilsson
J
J
D001011:Aorta / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D017209:Apoptosis / Q000187:drug effects*; D002118:Calcium / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002843:Chromatin / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D006888:Hydroxycholesterols / Q000494:pharmacology*; D008854:Microscopy, Electron; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology
D002843:Chromatin; D006888:Hydroxycholesterols; D011500:Protein Synthesis Inhibitors; C011724:cholest-5-en-3 beta,7 alpha-diol; D004247:DNA; D020928:Mitogen-Activated Protein Kinases; D002118:Calcium