Endothelial cell ER stress and Ca^<2+> signaling pathway

点击翻译按钮获取中文摘要

Apoptosis of endothelial cells (ECs) is now regarded to be an initial step inducing atherosclerosis. Recent studies have reported that the depletion of endoplasmic reticulum (ER) Ca^<2+> stores plays an important role in apoptosis. Caspase-12 is a key signal to lead ER stress-induced apoptosis. However, it is not known whether the depletion of ER Ca^<2+> is linked to caspase-12 signaling in ECs. Here we have investigated the interaction of Ca^<2+> signaling and caspase-12 cleavage in apoptosis of cultured porcine aortic ECs. Cytosolic Ca^<2+> concentration ([Ca^<2+>]i) was measured using fura-2/AM. Apoptosis was assessed by DNA ladder formation, and cleavage of caspase-12 by Western blotting. Thapsigargin (6μM), an inhibitor of the ER-associated Ca^<2+>-ATPase, increased [Ca^<2+>]i (F340/F380 ratio 0.717±0.137 to 6.133±0.710 : p<0.001) and induced persistent ER calcium depletion, cleavage of caspase-12 and apoptosis. Bradykinin (10 nM) increased [Ca^<2+>]i (F340/F380 ratio 0.717±0.053 to 6.133±0.528 : p<0.001) but induced neither cleavage of caspase-12 nor apoptosis. However, when ECs were treated with BAPTA/AM (100μM), BK caused the Ca^<2+> depletion of ER and apoptosis without the cleavage of caspase-12. Moreover non-selective caspase inhibitor, zVAD-fmk (100μM), inhibited apoptosis and cleavage of caspase-12 in TG-stimulated ECs, and calpain inhibitor, MDL 28170 (120μM), inhibited cleavage of caspase-12 but not inhibited apoptosis. These results suggested that the increase of [Ca^<2+>]i did not play an important role in inducing apoptosis in ECs, and ER Ca^<2+> depletion induced apoptosis, which is independent from caspase-12 linked signaling pathway.