Abstract
Treatment of intact human umbilical vein endothelial cells with NaF results in a dose-dependent biphasic response in both prostacyclin and inositol phosphate production: the stimulation observed with 10-20 mM NaF decreases with higher concentrations. High concentrations of NaF furthermore reduce thrombin- or A23187-stimulated prostacyclin production. Direct assay of phospholipase C activity in cell homogenates shows a similar biphasic response to NaF, also after chelation of Ca2+; addition of AlCl3 shifts the inhibition toward lower NaF concentrations. Guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) also causes a dose-dependent biphasic response in inositol phosphate formation in permeabilized cells and homogenates; a higher inhibitory concentration of GTP gamma S abolishes the stimulation of inositol phosphate production by low NaF concentrations. A high concentration of NaF furthermore inhibits the non-G-protein-dependent activation of phospholipase C by deoxycholate. NaF also induces a dose-dependent biphasic response in cyclic AMP formation in intact cells, indicating that the inhibition of phospholipase C at higher NaF concentrations does not result from a rise in cyclic AMP. The data are compatible with the existence of a guanine nucleotide-dependent, cyclic AMP-independent, phospholipase C-inhibitory pathway in endothelial cells.
Highlights
Treatment of intact human umbilical vein endothelial cells with NaF results in a dose-dependent biphasic response in both prostacyclin and inositol phosphate production: the stimulation observed with lo-20 mM
HUVECs were incubated for 2 h with [sH]adenosine and stimulated with increasing concentrations of NaF in the presence of
Values are the mean of three experiments and are expressed as percentage of [3H] adenosine incorporated by the HUVECs
Summary
Treatment of intact human umbilical vein endothelial cells with NaF results in a dose-dependent biphasic response in both prostacyclin and inositol phosphate production: the stimulation observed with lo-20 mM. (GTPyS) causes a dose-dependent biphasic response in inositol phosphate formation in permeabilized cells and homogenates; a higher inhibitory concentration of GTPyS abolishes the stimulation of inositol phosphate production by low NaF concentrations. NaF induces a dose-dependent biphasic response in cyclic AMP formation in intact cells, indicating that the inhibition of phospholipase C at higher NaF concentrations does not result from a rise in cyclic AMP. Evidence exists that stimulation of PLC quickly follows the binding of specific agonists to cell surface receptors through the signal-transducing action of a GTP-binding protein [3, 4]. This process in accordance with 18 U.S.C. Section 1734 solely to indicate this fact
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