Capsazepine elevates intracellular Ca 2+ in human osteosarcoma cells, questioning its selectivity as a vanilloid receptor antagonist

Abstract

Capsazepine is thought to be a selective antagonist of vanilloid type 1 receptors; however, its other in vitro effect on different cell types is unclear. In human MG63 osteosarcoma cells, the effect of capsazepine on intracellular Ca 2+ concentrations ([Ca 2+] i) and cytotoxicity was explored by using fura-2 and tetrazolium, respectively. Capsazepine caused a rapid rise in [Ca 2+] i in a concentration-dependent manner with an EC 50 value of 100 μM. Capsazepine-induced [Ca 2+] i rise was partly reduced by removal of extracellular Ca 2+, suggesting that the capsazepine-induced [Ca 2+] i rise was composed of extracellular Ca 2+ influx and intracellular Ca 2+. In Ca 2+-free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca 2+-ATPase, caused a monophasic [Ca 2+] i rise, after which the increasing effect of capsazepine on [Ca 2+] i was inhibited by 75%. Conversely, pretreatment with capsazepine to deplete intracellular Ca 2+ stores totally prevented thapsigargin from releasing more Ca 2+. U73122, an inhibitor of phospholipase C, abolished histamine (an inositol 1,4,5-trisphosphate-dependent Ca 2+ mobilizer)-induced, but not capsazepine-induced, [Ca 2+] i rise. Overnight treatment with 1–100 μM capsazepine inhibited cell proliferation in a concentration-dependent manner. These findings suggest that in human MG63 osteosarcoma cells, capsazepine increases [Ca 2+] i by stimulating extracellular Ca 2+ influx and also by causing intracellular Ca 2+ release from the endoplasmic reticulum via a phospholiase C-independent manner. Capsazepine may be mildly cytotoxic.