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