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Articles in PresS, published online ahead of print May 2, 2002
Am J Physiol Heart Circ Physiol, 10.1152/ajpheart.01067.2001
Submitted on December 5, 2001
Accepted on April 29, 2002
1 Servei de Cardiologia, Hospital Universitari Vall d'Hebron, Barcelona, Spain
* To whom correspondence should be addressed. E-mail: dgdorado{at}hg.vhebron.es.
To characterize the effects of ischemia on cyclic GMP (cGMP) synthesis in microvascular endothelium, cultured endothelial cells from adult rat hearts were exposed to hypoxia or normoxia at pH 6.4 or 7.4. Cellular cGMP and soluble (sGC) and membrane (mGC) guanylyl cyclase activities were measured after stimulation of sGC (SNAP) or mGC (urodilatin), or after no stimulation. Cell death (LDH release) was negligible in all experiments. Hypoxia at pH 6.4 induced a rapid ~90% decrease in cellular cGMP after sGC- and mGC-stimulation. This effect was reproduced by acidosis. Hypoxia at pH 7.4 elicited a less pronounced (~50%) and slower reduction in cGMP synthesis. Reoxygenation after 2 h of hypoxia at either pH 6.4 or 7.4 normalized the response to mGC stimulation, but further deteriorated sGC response; normalization of pH rapidly reversed the effects of acidosis. At pH 7.4 response to GC stimulation correlated well with cellular ATP. We conclude that simulated ischemia severely depresses cGMP synthesis in microvascular coronary endothelial cells through ATP depletion and acidosis without intrinsic protein alteration.
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