Mechanism of endothelial cell shape change in oxidant injury.
Changes in endothelial cell morphology induced by neutrophil-generated hydrogen peroxide (H2O2) may account for the capillary leak of the adult respiratory distress syndrome (ARDS). The relationship of H2O2 effects on the concentration of intracellular Ca2+ [( Ca2+]i) and ATP to changes in microfilaments and microtubules, important determinants of cell shape, was examined. Bovine pulmonary artery endothelial cells were injured over a 2-hr time course with a range of H2O2 doses (0-20 mM). The higher concentrations of H2O2 consistently produced contraction and rounding of greater than 50-75% of cells by 1-2 hr. The range of 1-20 mM H2O2 produced rapid, significant reductions in endothelial ATP levels over the time course of injury. Although there were significant increases in mean endothelial [Ca2+]i in response to 5, 10, and 20 mM H2O2, 1 mM H2O2 did not affect the [Ca2+]i. Fluorescence microscopy revealed that microfilament disruption occurred as ATP levels fell and preceded depolymerization of microtubules which developed after [Ca2+]i approached 1 X 10(-6) M. H2O2 at 1 mM injury caused microfilament disruption but did not depolymerize microtubules. Microfilament disruption occurred without oxidant exposure, when ATP levels were reduced by glucose depletion and mitochondrial inhibition with oligomycin (650 nM). If a Ca2+ ionophore, ionomycin (5 microM), was then added, [Ca2+]i rose to greater than 1 X 10(-6) M, microtubules fragmented and depolymerized, and cell contraction and rounding very similar to that induced by H2O2 occurred. These results suggest that endothelial cell dysfunction and capillary leak in ARDS may be due to H2O2-mediated changes in cellular ATP and [Ca2+]i.
Hinshaw
D B
DB
Department of Surgery, University of Michigan, Ann Arbor.
Burger
J M
JM
Armstrong
B C
BC
Hyslop
P A
PA
D008841:Actin Cytoskeleton / Q000187:drug effects; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D002463:Cell Membrane Permeability / Q000187:drug effects; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology* / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008870:Microtubules / Q000187:drug effects; D010084:Oxidation-Reduction / Q000187:drug effects*
D000255:Adenosine Triphosphate; D006861:Hydrogen Peroxide; D005978:Glutathione; D002118:Calcium