The role of calcium in the toxic effects of tert-butyl hydroperoxide on adult rat cardiac myocytes.
Oxidant stress has been implicated in reoxygenation damage following hypoxia and can lead to loss of membrane integrity and cell death. In this study the effects of oxidant stress, induced by tert-butyl hydroperoxide (tBHP), on cell conformation and intracellular free calcium ([Ca2+]i) of cardiac myocytes isolated from rat ventricles were examined. Incubation in the presence of 1 mM tBHP lead to a rise in [Ca2+]i, hypercontracture and loss of membrane integrity (as judged by trypan blue staining and loss of fluorescence of fura-2 loaded cells). Incubation in calcium-free medium or medium containing 2,3 butanedione-monoxime (BDM), which decreases myofibrillar calcium sensitivity, delayed but did not prevent the cell shape changes and loss of membrane integrity. In the presence of BDM, hypercontracture occurred at a higher [Ca2+]i than in control cells, indicating a possible role for [Ca2+]i in the generation of hypercontracture in this model. Treatment with calcium antagonists (10(-6) or 10(-7) M nisoldipine or 10(-6) M amlodipine) did not afford any protection against tBHP. ATP depletion did not accelerate loss of membrane integrity. Pretreatment of cells with the iron chelator, desferrioxamine mesylate greatly attenuated the effect of tBPH, delaying the rise in [Ca2+]i, cell shape changes and loss of membrane integrity. It appears, therefore, that tBHP-induced changes are mediated by the iron dependent generation of butyl alkoxyl radicals. The evidence suggests that tBHP-induced contracture is [Ca2+]i dependent rather than ATP dependent. Calcium modifies, but is not essential for the action of tBHP on isolated myocytes. During reoxygenation of hypoxic hearts calcium overload and free radical generation may act synergistically resulting in the characteristic changes associated with this condition, including loss of sarcolemmal integrity.
Daly
M J
MJ
Department of Medicine, University of Melbourne, Austin Hospital, Heidelberg, Victoria, Australia.
Young
R J
RJ
Britnell
S L
SL
Nayler
W G
WG
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000037:antagonists & inhibitors / Q000502:physiology*; D002478:Cells, Cultured; D003676:Deferoxamine / Q000494:pharmacology; D005260:Female; D006321:Heart / Q000187:drug effects*; D007700:Kinetics; D009206:Myocardium / Q000166:cytology / Q000378:metabolism; D016877:Oxidants / Q000633:toxicity*; D010545:Peroxides / Q000633:toxicity*; D051381:Rats; D011919:Rats, Inbred Strains; D020122:tert-Butylhydroperoxide
D016877:Oxidants; D010545:Peroxides; D000255:Adenosine Triphosphate; D020122:tert-Butylhydroperoxide; D003676:Deferoxamine; D002118:Calcium