BW755C or staurosporine inhibits collagen-stimulated phosphoinositide phosphorylation in platelets.
Stimulation of platelets by collagen results in increased formation of the polyphosphoinositides, phosphatidylinositol phosphate (PtdInsP) and phosphatidylinositol bisphosphate (PtdInsP2) through stimulation of phosphoinositide kinase activities. We investigated a possible regulatory role of endogenous thromboxane formation and protein kinase C (PKC) activation in the induction of phosphoinositide phosphorylation following collagen stimulation, as well as following stimulation by the thromboxane mimetic, U-46619. Human platelets were prelabeled with [3H]inositol and stimulated with collagen (2 micrograms/mL) or U-46619 (1 microM), in the absence or presence of either the cyclo-oxygenase/lipoxygenase inhibitor, BW755C, or staurosporine, a putative inhibitor or PKC. Collagen stimulation resulted in a time-dependent increase in [3H]inositol-labeled PtdInsP and PtdInsP2 which was completely inhibited in the presence of BW755C. Addition of U-46619 to BW755C-treated, collagen-stimulated platelets restored the increased polyphosphoinositide formation. Stimulation of platelets with U-46619 alone also resulted in increased formation of [3H]PtdInsP and [3H]PtdInsP2, but this was not affected by the presence of BW755C. These results suggest that the collagen-induced activation of phosphoinositide kinases was dependent upon thromboxane formation, but that U-46619-induced phosphoinositide formation was rather independent of further thromboxane production. Pretreatment of platelets with staurosporine, prior to agonist addition, completely blocked the collagen-stimulated rise in radiolabeled PtdInsP and the U-46619-induced PtdInsP and PtdInsP2 generations, suggesting that protein kinase, possibly PKC, may play a role in the activation of phosphoinositide kinases by these agonists.
Thomas
L M
LM
Department of Nutritional Sciences, University of Guelph, Ontario, Canada.
Holub
B J
BJ
D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; D015772:4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyrazol-3-amine / Q000494:pharmacology*; D000470:Alkaloids / Q000494:pharmacology*; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D003094:Collagen / Q000378:metabolism*; D006801:Humans; D007553:Isotope Labeling; D010716:Phosphatidylinositols / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011450:Prostaglandin Endoperoxides, Synthetic; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D019311:Staurosporine; D014316:Tritium
D000470:Alkaloids; D010716:Phosphatidylinositols; D011450:Prostaglandin Endoperoxides, Synthetic; D014316:Tritium; D015772:4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyrazol-3-amine; D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; D003094:Collagen; D011493:Protein Kinase C; D019311:Staurosporine