title: BW755C or staurosporine inhibits collagen-stimulated phosphoinositide phosphorylation in platelets.
abstract: 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.
authors.affiliation: Department of Nutritional Sciences, University of Guelph, Ontario, Canada.
authors.forename: L M
authors.initials: LM
authors.lastname: Thomas
authors.forename: B J
authors.initials: BJ
authors.lastname: Holub
mesh_terms: 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
chemical_list: 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