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Evidence for requirement of tyrosine phosphorylation in endothelial P 2y ‐ and P 2u ‐ purinoceptor stimulation of prostacyclin release
Author(s) -
Bowden Abigail,
Patel Viral,
Brown Colin,
Boarder Michael R.
Publication year - 1995
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/j.1476-5381.1995.tb17208.x
Subject(s) - prostacyclin , stimulation , phosphorylation , tyrosine , chemistry , tyrosine phosphorylation , platelet , pharmacology , endocrinology , microbiology and biotechnology , biochemistry , medicine , biology
1 . The release of prostacylin (PGI 2 ) from vascular endothelial cells is stimulated by ATP acting at G protein‐coupled P 2 ‐purinoceptors. Here we investigate the hypothesis that tyrosine protein phosphorylations are involved in this response. 2 . The use of Western blots with anti‐phosphotyrosine antibodies showed that 30 μm 2MeSATP (selective for P 2Y ‐purinoceptors), 300 μm UTP (selective for P 2U ‐purinoceptors) and 300 μm ATP (effective at both these purinoceptors), each stimulate the tyrosine phosphorylation of proteins in bovine cultured aortic endothelial cells. Each of these agonists also stimulates 6‐keto PGF 1α accumulation in the medium (an index of PGI 2 release) in these cells in the same period. 3 . The tyrosine kinase inhibitor, genistein, inhibits the 6‐keto PGF 1α response with the same concentration‐dependency (1–100 μm) as the tyrosine phosphorylation response. 4 . Tyrphostin, a structurally and functionally distinct tyrosine kinase inhibitor, is also a potent inhibitor (0.1–10 μm) of the 6‐keto PGF 1α response. 5 . Neither tyrphostin nor genistein inhibit the phospholipase C response to P 2 ‐purinoceptor stimulation. Furthermore, these inhibitors do not affect the 6‐keto PGF 1α response to ionomycin. 6 . These results show that the regulation of vascular endothelial cells by ATP acting at both P 2Y ‐ and P 2U ‐purinoceptors involves the stimulation of tyrosine phosphorylation, and suggest that this is a necessary event for the purinoceptor‐mediated stimulation of PGI 2 production.