Alterations of extracellular calcium elicit selective modes of cell death and protease activation in SH-SY5Y human neuroblastoma cells.
The role of intracellular Ca2+ homeostasis in mechanisms of neuronal cell death and cysteine protease activation was investigated in SH-SY5Y human neuroblastoma cells. Cells were incubated in 2 mM EGTA to lower intracellular Ca2+ or 5 mM CaCl2 to raise it. Cell death and activation of calpain and caspase-3 were measured. Both EGTA and excess CaCl2 elicited cell death. EGTA induced DNA laddering and an increase in caspase-3-like, but not calpain, activity. Pan-caspase inhibitors protected against EGTA-, but not CaCl2-, induced cell death. Conversely, excess Ca2+ elicited necrosis and activated calpain but not caspase-3. Calpain inhibitors did not preserve cell viability. Ca2+ was the death-mediating factor, because restoration of extracellular Ca2+ protected against cell death induced by EGTA and blockade of Ca2+ channels by Ni2+ protected against that induced by high Ca2+. We conclude that the EGTA treatment lowered intracellular Ca2+ and elicited caspase-3-like protease activity, which led to apoptosis. Conversely, excess extracellular Ca2+ entered Ca2+ channels and increased intracellular Ca2+ leading to calpain activation and necrosis. The mode of cell death and protease activation in response to changing Ca2+ were selective and mutually exclusive, demonstrating that these are useful models to individually investigate apoptosis and necrosis.
McGinnis
K M
KM
Department of Pharmacology, University of Michigan, Ann Arbor, USA.
Wang
K K
KK
Gnegy
M E
ME
D002118:Calcium / Q000378:metabolism*; D002154:Calpain / Q000378:metabolism; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D016923:Cell Death / Q000502:physiology; D004273:DNA, Neoplasm / Q000235:genetics / Q000502:physiology; D004533:Egtazic Acid / Q000494:pharmacology; D010450:Endopeptidases / Q000378:metabolism*; D004789:Enzyme Activation / Q000502:physiology; D005110:Extracellular Space / Q000378:metabolism*; D006801:Humans; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D009336:Necrosis; D009474:Neurons / Q000201:enzymology / Q000378:metabolism / Q000502:physiology*; D009994:Osmolar Concentration; D011480:Protease Inhibitors / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D011480:Protease Inhibitors; D004533:Egtazic Acid; D007770:L-Lactate Dehydrogenase; D010450:Endopeptidases; C505407:CASP3 protein, human; D002154:Calpain; D053148:Caspase 3; D020169:Caspases; D002118:Calcium