bcl-2 protects HL-60 cells from apoptosis by stabilizing their intracellular calcium pools

bcl-2 has been shown to enhance cell survival by inhibiting apoptosis induc ed under different circumstances. In this study we investigated the effects of bcl-2 overexpression on the homeostasis of subcellular organelles such as ER and mitochondria. In our study, HL-60/bcl-2 and control HL-60/neo cel ls were obtained by transfection of bcl-2 cDNA or the neomycin-resistant ge ne, respectively. Apoptosis was evaluated by both DNA fragmentation and how cytometry qualitatively and quantitatively, and the intracellular calcium by Fura-2/AM. Thapsigargin (TG), a highly specific inhibitor of the ER-asso ciated Ca2+ pump, and Br-A23187, a calcium ionophore, were used in this stu dy. Our results showed that overexpression of bcl-2 significantly blocked T G- and Br-A23187-induced apoptosis in calcium containing buffer. Measuremen t of intracellular calcium showed that bcl-2 overexpression could reduce su stained elevation of cytosolic Ca2+ induced by these agents. However, in ca lcium-free medium, bcl-2 overexpression maintained Ca2+ uptake in ER of bot h TG- and Br-A23187-treated cells. Moreover, the depletion of Ca2+ by EGTA enhanced TG- and Br-A23187-induced apoptosis, and reduced the anti-apoptoti c action of bcl-2, suggesting that cytosolic Ca2+ elevation may be required for optimal ER pool refilling. These findings suggest that bcl-2 facilitat es and maintains the replenishment of Ca2+ in intracellular stores and, as a result, influences the intracellular calcium, thus protecting the cells f rom death. In addition, there were no cytochrome c release from mitochondri a into the cytosol in TG- and Br-A23187- induced apoptosis, suggesting that cytochrome c release is not a universal phenomenon in the apoptotic proces s, (C) 2001 Elsevier Science Inc. All rights reserved.