TY - JOUR
T1 - Cytosolic free Ca2+ and proteolysis in lethal oxidative injury in endothelial cells
AU - Geeraerts, M. D.
AU - Ronveaux-Dupal, M. F.
AU - Lemasters, J. J.
AU - Herman, B.
PY - 1991
Y1 - 1991
N2 - Oxygen free radicals (OFR) are thought to mediate ischemia-reperfusion injury to endothelium of heart, lung, brain, liver, and kidney and contribute to development of atherosclerosis, pulmonary O2 toxicity, and adult respiratory distress syndrome. Increased cytosolic free Ca2+ (Ca(i)2+) has been proposed as a mechanism of injury from oxidative stress, yet the pathways by which an increase in Ca(i)2+ may cause OFR-mediated endothelial cell injury remain unknown. Using multiparameter digitized video microscopy and the fluorescent probes, fura-2 acetoxymethyl ester and propidium iodide, we measured Ca(i)2+ and cell viability in human umbilical endothelial cells during oxidative stress with xanthine (50 μM) plus xanthine oxidase (40 mU/ml). Oxidative stress caused a sustained increase in Ca(i)2+ from a resting level of 90-100 nM to near 500 nM, which was preceded by formation of plasma membrane blebs. The increase in Ca(i)2+ was prevented by removal of extracellular Ca2+ (Ca(o)2+). Prevention of the increase in Ca(i)2+ was associated with prolonged cell viability. Readdition of Ca(o)2+ resulted in an immediate large increase in Ca(i)2+ and rapid onset of cell death. The protease inhibitors, leupeptin and pepstatin, delayed the increase in Ca(i)2+ and prolonged cell viability. The results are consistent with the hypothesis that endothelial cell injury due to oxidative stress may be the result of Ca(i)2+ influx and resultant activation of Ca2+-dependent proteases.
AB - Oxygen free radicals (OFR) are thought to mediate ischemia-reperfusion injury to endothelium of heart, lung, brain, liver, and kidney and contribute to development of atherosclerosis, pulmonary O2 toxicity, and adult respiratory distress syndrome. Increased cytosolic free Ca2+ (Ca(i)2+) has been proposed as a mechanism of injury from oxidative stress, yet the pathways by which an increase in Ca(i)2+ may cause OFR-mediated endothelial cell injury remain unknown. Using multiparameter digitized video microscopy and the fluorescent probes, fura-2 acetoxymethyl ester and propidium iodide, we measured Ca(i)2+ and cell viability in human umbilical endothelial cells during oxidative stress with xanthine (50 μM) plus xanthine oxidase (40 mU/ml). Oxidative stress caused a sustained increase in Ca(i)2+ from a resting level of 90-100 nM to near 500 nM, which was preceded by formation of plasma membrane blebs. The increase in Ca(i)2+ was prevented by removal of extracellular Ca2+ (Ca(o)2+). Prevention of the increase in Ca(i)2+ was associated with prolonged cell viability. Readdition of Ca(o)2+ resulted in an immediate large increase in Ca(i)2+ and rapid onset of cell death. The protease inhibitors, leupeptin and pepstatin, delayed the increase in Ca(i)2+ and prolonged cell viability. The results are consistent with the hypothesis that endothelial cell injury due to oxidative stress may be the result of Ca(i)2+ influx and resultant activation of Ca2+-dependent proteases.
KW - Calcium
KW - Endothelial cell
KW - Leupeptin
KW - Oxidative stress
KW - Pepstatin
KW - Protease
KW - Toxic oxygen species
KW - pH
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U2 - 10.1152/ajpcell.1991.261.5.c889
DO - 10.1152/ajpcell.1991.261.5.c889
M3 - Article
C2 - 1951673
AN - SCOPUS:0026332408
SN - 0002-9513
VL - 261
SP - C889-C896
JO - American Journal of Physiology - Cell Physiology
JF - American Journal of Physiology - Cell Physiology
IS - 5 30-5
ER -