Role of lipid peroxidation in tert-butylhydroperoxide-induced inhibition of endothelial cell calcium signaling.
The potential of lipid peroxidation in inhibition of Ca++ signaling by the membrane-permeant oxidant, tertiary butylhydroperoxide (tert-buOOH), was investigated in calf pulmonary vascular endothelial cells. The oxidant dose-dependently increased lipid peroxidation between concentrations of 10(-5) and 10(-3) M, with an ED50 of approximately 0.05 mM. In addition, the effect of tert-buOOH was time-dependent through the experimental period (3 h). Preincubation of cells with the 21-amino-steroid compound, 21-[4-(5,6-bis(diethylamino)-2-pyridinyl)-1-piperazinyl]-16 alpha-methyl-pregna-1,4,9(11)-triene-3,20-dione hydrochloride (U74500A), reduced tert-buOOH-induced lipid peroxidation to undetectable levels. The effect of U74500A was dose dependent with an IC50 of approximately 10(-6) M. Brief incubation of cells with the aminosteroid resulted in greater than 90% inhibition of lipid peroxidation during subsequent 2-h incubations with tert-buOOH and addition of U74500A during treatment of cells with tert-buOOH halted further lipid peroxidation. In contrast, the iron-containing moiety, hemin, potentiated the effect of tert-buOOH on lipid peroxidation. The ED50 of hemin was approximately 10(-6) M when cells were preincubated with this agent before treatment with tert-buOOH. The potentiating effect of hemin was time-dependent and reached a near maximum upon incubation of cells for 1 h before tert-buOOH. Preincubation of cells with U74500A before treatment with hemin and tert-buOOH decreased lipid peroxidation by 75%. Ca++ signaling was monitored in-cells loaded with the Ca(++)-sensitive fluorescent indicator, fura-2.(ABSTRACT TRUNCATED AT 250 WORDS)
Elliott
S J
SJ
Department of Pediatrics, Baylor College of Medicine, Houston, Texas.
Doan
T N
TN
Schilling
W P
WP
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000378:metabolism; D006427:Hemin / Q000494:pharmacology; D015227:Lipid Peroxidation / Q000187:drug effects*; D010545:Peroxides / Q000494:pharmacology*; D011281:Pregnatrienes / Q000494:pharmacology; D011651:Pulmonary Artery; D015398:Signal Transduction / Q000187:drug effects; D020122:tert-Butylhydroperoxide
D010545:Peroxides; D011281:Pregnatrienes; D006427:Hemin; D020122:tert-Butylhydroperoxide; C053356:U 74500A; D002118:Calcium