scholarly journals Sphingosine enhances platelet aggregation through an increase in phospholipase C activity by a protein kinase C-independent mechanism

1992 ◽  
Vol 282 (1) ◽  
pp. 243-247 ◽  
Author(s):  
T Hashizume ◽  
T Sato ◽  
T Fujii
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Platelet Activating Factor ◽  
Membrane Phospholipids ◽  
Stimulus Response ◽  
Protein Kinase C Inhibitors ◽  
Kinase C ◽  
Low Concentrations ◽  
Positive Modulator

Sphingosine (a potent inhibitor of protein kinase C) at 5-10 microM, which are concentrations lower than those that inhibit this enzyme activity, enhanced the aggregation of rabbit platelets induced by low concentrations of U46619, platelet-activating factor, thrombin and arachidonic acid, whereas H-7 and staurosporine, other protein kinase C inhibitors, failed to do so. Of the sphingosine analogues which also inhibit protein kinase C, psychosine and lyso-GM3 did not show such an enhancing effect. Sphingosine promoted both Ins(1,4,5)P3 formation and an increase in the cytoplasmic free Ca2+ concentration in response to all the agonists used. Furthermore, the hydrolytic action of exogenously added phospholipase C (from Clostridium perfringens) on platelet membrane phospholipids was dose-dependently enhanced by pretreatment of the platelets with sphingosine. These results imply that sphingosine, at relatively low concentrations, brings about hyperaggregability of the platelets by the agonists employed, probably owing to enhancement of the phospholipase C activity. Such an effect appears to be induced by a mechanism independent of protein kinase C inhibition. We suggest that sphingosine might act as a positive modulator for the stimulus-response coupling in the platelets.

scholarly journals Protein kinase C promotes restoration of calcium homeostasis to platelet activating factor-stimulated human neutrophils by inhibition of phospholipase C

2009 ◽  
Vol 6 (1) ◽  
pp. 29 ◽  
Author(s):  
Gregory R Tintinger ◽  
Annette J Theron ◽  
Helen C Steel ◽  
Riana Cockeran ◽  
Lynette Pretorius ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Calcium Homeostasis ◽  
Platelet Activating Factor ◽  
Human Neutrophils ◽  
Kinase C

Trypsin stimulation of aldosterone and 18-hydroxycorticosterone production by rat adrenal zona glomerulosa tissue is mediated by activation of protein kinase C

1990 ◽  
Vol 5 (1) ◽  
pp. 85-93 ◽  
Author(s):  
G. P. Vinson ◽  
S. M. Laird ◽  
J. P. Hinson ◽  
N. Mallick ◽  
S. Marsigliante ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Polymyxin B ◽  
Zona Glomerulosa ◽  
Protein Kinase C Inhibitors ◽  
Kinase C ◽  
Protein Kinase C Activity ◽  
Tissue Sensitivity ◽  
Protein Components

ABSTRACT When rat adrenal whole capsules, containing the zona glomerulosa, were incubated, addition of the protein kinase C inhibitors TMB-8 (10 μmol/l), W7, H7, polymyxin-B and sphingosine (all 1 μmol/l) was found to inhibit the steroidogenic response to trypsin. Aldosterone and 18-hydroxycorticosterone were strongly, and corticosterone moderately, affected, while the production of 18-hydroxydeoxycorticosterone was neither stimulated by trypsin nor inhibited by the protein kinase C inhibitors. Addition of neomycin, which prevents substrate interaction with phospholipase C, also inhibited the response to trypsin, while addition of phospholipase C itself stimulated aldosterone, 18-hydroxycorticosterone and corticosterone production with the same tissue sensitivity as trypsin. Addition of phospholipase A2 had no effect. Direct assay of protein kinase C activity showed that trypsin stimulation effected the translocation of Ca2+/phospholipid-activated protein kinase C from the cytosolic to the membrane fraction. When glomerulosa tissue was incubated with [32P]ATP, and cytosolic proteins were subjected to isoelectric focusing on polyacrylimide gels, autoradiography showed that incorporation of 32P into several protein components was increased by trypsin stimulation. It was concluded that trypsin exerts its stimulatory effects on steroidogenesis by activating protein kinase C; not, however, by generating the Ca2+/phospholipid-independent fragment, but possibly by enhancing the activity of phospholipase C.

Differential megakaryocytic desensitization to platelet agonists

1992 ◽  
Vol 263 (4) ◽  
pp. C864-C872 ◽  
Author(s):  
G. W. Dorn ◽  
M. G. Davis
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Calcium Signaling ◽  
Phospholipase C ◽  
Platelet Activating Factor ◽  
Dienoic Acid ◽  
Cytosolic Calcium ◽  
Peripheral Circulation ◽  
Human Platelets ◽  
Kinase C

Platelets are released into the peripheral circulation from the bone marrow where they arise as fragments of megakaryocyte cytoplasm. To characterize the effects of platelet agonists on megakaryocytes, we examined calcium signaling and desensitization to thrombin, the thromboxane A2 (TxA2) mimetic (15S)-hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5Z,13E-dienoic acid (U46619), and platelet-activating factor (PAF) in cultured CHRF-288-11 megakaryocytic cells. Initially, we compared agonist-stimulated calcium transients in fura-2-loaded CHRF-288-11 cells and human platelets. The 50% effective concentration values for the agonists to increase free cytosolic calcium were as follows: thrombin (0.11 +/- 0.02 U/ml in CHRF, 0.19 +/- 0.03 U/ml in platelets), U46619 (147 +/- 33 nM in CHRF, 157 +/- 5 nM in platelets), and PAF [15 +/- 2 nM in CHRF, 16 +/- 2 nM in platelets (n = 4 each)]. CHRF-288-11 thrombin, TxA2, and PAF receptors were demonstrated to be coupled to phospholipase C because each of the agonists stimulated phosphatidylinositol hydrolysis in myo-[3H]inositol-loaded CHRF-288-11 cells and pharmacological inhibition of phospholipase C-blunted agonist-stimulated calcium signaling. CHRF-288-11 cells exposed to the three agonists for 1 h showed different patterns and extent of homologous and heterologous desensitization. Protein kinase C activation appeared to be necessary but not sufficient for desensitization because 1) activation of protein kinase C with phorbol 12-myristate 13-acetate inhibited the calcium responses to all three agonists, 2) inhibition of protein kinase C with staurosporine attenuated subsequent desensitization to each agonist, and 3) each agonist increased protein kinase C activity in CHRF-288-11 cell homogenates.

scholarly journals Mechanism of hepatic glycogen synthase inactivation induced by Ca2+-mobilizing hormones. Studies using phospholipase C and phorbol myristate acetate

1986 ◽  
Vol 237 (1) ◽  
pp. 235-242 ◽  
Author(s):  
P F Blackmore ◽  
W G Strickland ◽  
S B Bocckino ◽  
J H Exton
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Glycogen Synthase ◽  
Partial Purification ◽  
Hepatic Glycogen ◽  
Kinase C ◽  
Low Concentrations ◽  
Dependent Inactivation

Incubation of hepatocytes with the protein kinase C activator and tumour promoter 4 beta-phorbol 12 beta-myristate 13 alpha-acetate (PMA) produced a time- and concentration-dependent inactivation of glycogen synthase, but no change in phosphorylase. The same rate and extent of inactivation occurred in hepatocytes depleted of Ca2+ by treatment with the Ca2+ chelator EGTA. When hepatocytes were treated with the Ca2+-mobilizing hormone vasopressin (10 nM), the rate of glycogen synthase inactivation was similar to that observed with PMA (1 microM). Depletion of intracellular Ca2+ stores with EGTA abolished the ability of vasopressin to mobilize Ca2+ and activate phosphorylase without abolishing its ability to inactivate glycogen synthase and increase 1,2-diacylglycerol (DAG), the endogenous activator of protein kinase C. Protein kinase C, either in membranes or after partial purification, was shown to be activated in vitro by PMA in the presence of very low concentrations of Ca2+. Exogenous phospholipase C from Clostridium perfringens, at low concentrations, inactivated glycogen synthase and increased DAG without affecting cell Ca2+ or phosphorylase. It is proposed that the inactivation of glycogen synthase elicited by the Ca2+-mobilizing hormones is due, at least in part, to generation of DAG and activation of protein kinase C.

scholarly journals Ethanol-induced phospholipase C activation is inhibited by phorbol esters in isolated hepatocytes

1988 ◽  
Vol 251 (3) ◽  
pp. 865-871 ◽  
Author(s):  
J B Hoek ◽  
R Rubin ◽  
A P Thomas
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Phorbol Esters ◽  
Inositol Phosphates ◽  
Isolated Hepatocytes ◽  
Intact Cells ◽  
Protein Kinase C Inhibitors ◽  
Kinase C ◽  
Activate Protein Kinase

Ethanol causes a transient activation of the phosphoinositide-specific phospholipase C in intact hepatocytes and mimics the action of receptor-mediated agonists [Hoek, Thomas, Rubin & Rubin (1987) J. Biol. Chem. 262, 682-691]. Preincubation of the hepatocytes with phorbol esters which activate protein kinase C prevented this effect of ethanol: phorbol ester treatment inhibited the ethanol-induced phosphorylase activation, the increase in intracellular free Ca2+ concentrations measured in quin 2-loaded hepatocytes, and the changes in concentrations of inositol phosphates, phosphoinositides and phosphatidic acid. Several lines of evidence indicate that these effects were mediated by protein kinase C. Phorbol esters acted in a concentration range where they activate protein kinase C; phorbol esters that do not activate protein kinase C were not effective in inhibiting the effects of ethanol. The permeant diacylglycerol oleoyl-acetylglycerol also inhibited the effects of ethanol, but other diacylglycerols were not effective in the intact cells. The inhibition of ethanol-induced Ca2+ mobilization by phorbol esters was prevented by preincubating the cells with the protein kinase C inhibitors 1-(5-isoquinolinesulphonyl)-2-methylpiperazine (H7) and sphingosine. H7 also enhanced the Ca2+ mobilization induced by ethanol in cells that were not pretreated with phorbol esters, indicating that the transient nature of the ethanol-induced Ca2+ mobilization may be due to an activation of protein kinase C caused by the accumulation of diacylglycerol. These data support a model whereby ethanol activates the phosphoinositide-specific phospholipase C, possibly by affecting receptor-G-protein-phospholipase C interactions in the membrane.

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