title: Effect of Bcl-2 on oxidant-induced cell death and intracellular Ca2+ mobilization.
abstract: The mechanism by which Bcl-2 inhibits cell death is unknown. It has been suggested that Bcl-2 functions as an antioxidant. Because Bcl-2 is localized mainly to the membranes of the endoplasmic reticulum (ER) and the mitochondria, which represent the main intracellular storage sites for Ca2+, we hypothesized that Bcl-2 might protect cells against oxidant injury by altering intracellular Ca2+ homeostasis. To test this hypothesis, we examined the effect of oxidant treatment on viability in normal rat kidney (NRK) cells and in NRK cells stably transfected with Bcl-2 in the presence or absence of intracellular Ca2+, and we compared the effect of Bcl-2 expression on oxidant-induced intracellular Ca2+ mobilization and on ER and mitochondrial Ca2+ pools. NRK cells transfected with Bcl-2 (NRK-Bcl-2) were significantly more resistant to H2O2-induced cytotoxicity than control cells. EGTA-AM, an intracellular Ca2+ chelator, as well as the absence of Ca2+ in the medium, reduced H2O2-induced cytotoxicity in both cell lines. Compared with controls, cells overexpressing Bcl-2 showed a delayed rise in intracellular Ca2+ concentration ([Ca2+]i) after H2O2 treatment. After treatment with the Ca2+ ionophore ionomycin, Bcl-2-transfected cells showed a much quicker decrease after the maximal rise than control cells, suggesting stronger intracellular Ca2+ buffering, whereas treatment with thapsigargin, an inhibitor of the ER Ca2+-ATPases, transiently increased [Ca2+]i in control and in Bcl-2-transfected cells. Estimates of mitochondrial Ca2+ stores using an uncoupler of oxidative phosphorylation show that NRK-Bcl-2 cells have a higher capacity for mitochondrial Ca2+ storage than control cells. In conclusion, Bcl-2 may prevent oxidant-induced cell death, in part, by increasing the capacity of mitochondria to store Ca2+.
authors.affiliation: Department of Pathology, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.
authors.forename: M
authors.initials: M
authors.lastname: Ichimiya
authors.forename: S H
authors.initials: SH
authors.lastname: Chang
authors.forename: H
authors.initials: H
authors.lastname: Liu
authors.forename: I K
authors.initials: IK
authors.lastname: Berezesky
authors.forename: B F
authors.initials: BF
authors.lastname: Trump
authors.forename: P A
authors.initials: PA
authors.lastname: Amstad
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D016923:Cell Death / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D003587:Cytomegalovirus / Q000235:genetics; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D006801:Humans; D006861:Hydrogen Peroxide / Q000633:toxicity*; D015759:Ionomycin / Q000494:pharmacology; D007668:Kidney; D007700:Kinetics; D008928:Mitochondria / Q000378:metabolism*; D016877:Oxidants / Q000633:toxicity*; D018384:Oxidative Stress; D011401:Promoter Regions, Genetic; D011419:Propidium / Q000633:toxicity; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis; D014162:Transfection
chemical_list: D016877:Oxidants; D019253:Proto-Oncogene Proteins c-bcl-2; D011994:Recombinant Proteins; D011419:Propidium; D004533:Egtazic Acid; D015759:Ionomycin; C046770:EGTA acetoxymethyl ester; D006861:Hydrogen Peroxide; D002118:Calcium