Calcium signaling in endothelial cells involves activation of tyrosine kinases and leads to activation of mitogen-activated protein kinases.
The activation of endothelial cells following exposure to a variety of receptor-dependent and -independent stimuli is associated with the release of Ca2+ from intracellular stores as well as the influx of Ca2+ from the extracellular space. In the present study, we investigated the interaction between Ca2+ signaling in cultured human umbilical vein endothelial cells and tyrosine phosphorylation. Stimulation of endothelial cells with either bradykinin (100 nmol/L), histamine (1 mumol/L), or the Ca(2+)-ATPase inhibitor thapsigargin (30 nmol/L) resulted in a slightly delayed but prolonged tyrosine phosphorylation of two low molecular weight proteins (approximately 42 and approximately 44 kD). These proteins were identified by immunoprecipitation as the 42- and 44-kD isoforms of mitogen-activated protein kinase (MAP kinase). The agonist-induced tyrosine phosphorylation of the 42-/44-kD doublet was sensitive to the tyrosine kinase inhibitors genistein (100 mumol/L) and piceatannol (10 mumol/L) and was inhibited by the removal of Ca2+ from the extracellular medium. In fura 2-loaded endothelial cells, inhibition of tyrosine kinases attenuated Ca2+ signaling after stimulation with either bradykinin (30 nmol/L) or thapsigargin (30 nmol/L). Since inhibition of tyrosine kinases specifically attenuates the plateau phase of the Ca2+ response after stimulation, the effect of tyrosine kinase inhibition appeared to be mostly associated with the influx of Ca2+ from the extracellular space.(ABSTRACT TRUNCATED AT 250 WORDS)
Fleming
I
I
Zentrum der Physiologie, Klinikum der JWG-Universität, Frankfurt/Main, Germany.
Fisslthaler
B
B
Busse
R
R
D001920:Bradykinin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000494:pharmacology; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D004789:Enzyme Activation; D019833:Genistein; D006632:Histamine / Q000494:pharmacology; D006801:Humans; D015151:Immunoblotting; D007529:Isoflavones; D019950:Mitogen-Activated Protein Kinase 1; D010766:Phosphorylation / Q000187:drug effects; D011233:Precipitin Tests; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011994:Recombinant Proteins; D015398:Signal Transduction; D013729:Terpenes / Q000494:pharmacology; D019284:Thapsigargin; D014471:Umbilical Veins
D007529:Isoflavones; D011994:Recombinant Proteins; D013729:Terpenes; D019284:Thapsigargin; D006632:Histamine; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D019950:Mitogen-Activated Protein Kinase 1; D000252:Calcium-Transporting ATPases; D001920:Bradykinin; D002118:Calcium