Effect of thapsigargin on Ca2+ fluxes and viability in human prostate cancer cells

Effect of the carcinogen thapsigargin on human prostate cancer cells is unclear. This study examined if thapsigargin altered basal [Ca2+](i) levels in suspended PC3 human prostate cancer cells by using fura-2 as a Ca2+-sensitive fluorescent probe. Thapsigargin at concentrations between 10 nM and 10 mu M increased [Ca2+](i) in a concentration-dependent fashion. The Ca2+ signal was reduced partly by removing extracellular Ca2+ indicating that Ca2+ entry and release both contributed to the [Ca2+](i) rise. This Ca2+ influx was inhibited by suppression of phospholipase A2, but not by inhibition of store-operated Ca2+ channels or by modulation of protein kinase C activity. In Ca2+-free medium, pretreatment with the endoplasmic reticulum Ca2+ pump inhibitor 2,5-di-(t-butyl)-1,4-hydroquinone (BHQ) nearly abolished thapsigargin-induced Ca2+ release. Conversely, pretreatment with thapsigargin greatly reduced BHQ-induced [Ca2+](i) rise, suggesting that thapsigargin released Ca2+ from the endoplasmic reticulum. Inhibition of phospholipase C did not change thapsigargin-induced [Ca2+](i) rise. At concentrations of 1-10 mu M, thapsigargin induced cell death that was partly reversed by chelation of Ca2+ with BAPTA/AM. Annexin V/propidium iodide staining data suggest that apoptosis was partly responsible for thapsigargin-induced cell death. Together, in PC3 human prostate cancer cells, thapsigargin induced [Ca2+](i) rises by causing phospholipase C-independent Ca2+ release from the endoplasmic reticulum and Ca2+ influx via phospholipase A2-sensitive Ca2+ channels. Thapsigargin also induced cell death via Ca2+-dependent pathways and Ca2+-independent apoptotic pathways.