Effects of Ca2+ deregulation on mitochondrial membrane potential and cell viability in nucleated cells following lytic complement attack.
We have previously shown [Papadimitriou JC. Ramm LE. Drachenberg CB. Trump BF. Shin ML. (1991) J. Immunol., 147, 212-217] that formation of lytic C5b-9 channels on Ehrlich ascites tumor cells induced rapid depletion of adenine nucleotides associated with prelytic leakage preceding cell death. Extracellular Ca2+ concentration ([Ca2+]e) reduction by chelation markedly delayed the onset of cell death, although the adenine nucleotide leakage was enhanced. In the present study, we examined the temporal relationships between ionized cytosolic Ca2+ ([Ca2+]i), mitochondrial membrane potential (delta psi m) and cell death in individual cells by digital imaging fluorescence microscopy (DIFM), during the earliest phase of C5b-9 attack. The results showed an immediate, > 20-fold rise in [Ca2+]i, rapidly followed by dissipation of delta psi m and subsequent acute cell death. These events were markedly delayed by chelation of Ca2+e, but not by nominally Ca2+ free medium. Differing from previous reports indicating propidium iodide labeling of viable cells bearing C5b-9 channels, with DIFM we observed nuclear fluorescence with that marker only in association with cell death. These findings indicate that Ca2+ influx through lytic C5b-9 channels is responsible for the massive increase in [Ca2+]i, as well as for the rapid loss of delta psi m, followed by acute cell death. When this [Ca2+]i increase is prevented, the cell death is probably related to metabolic depletion.
Papadimitriou
J C
JC
Department of Pathology, University of Maryland School of Medicine, Baltimore.
Phelps
P C
PC
Shin
M L
ML
Smith
M W
MW
Trump
B F
BF
D000227:Adenine Nucleotides / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D016923:Cell Death / Q000502:physiology*; D003167:Complement Activation / Q000502:physiology*; D015938:Complement Membrane Attack Complex / Q000502:physiology*; D007091:Image Processing, Computer-Assisted; D007425:Intracellular Membranes / Q000502:physiology; D008564:Membrane Potentials / Q000502:physiology; D051379:Mice; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods; D008928:Mitochondria / Q000502:physiology*; D014407:Tumor Cells, Cultured
D000227:Adenine Nucleotides; D015938:Complement Membrane Attack Complex; D002118:Calcium