BACKGROUND： This study aimed to explore the effects of TNF-α on the expression of IP3R1 mRNA and protein in human mesangial cells （HMCs）, and to elucidate the mechanism of TNF-α relating to IP3R1 expression in th...BACKGROUND： This study aimed to explore the effects of TNF-α on the expression of IP3R1 mRNA and protein in human mesangial cells （HMCs）, and to elucidate the mechanism of TNF-α relating to IP3R1 expression in the occurrence of hepatorenal syndrome （HRS）.METHODS： HMCs were stimulated by tumor （TNF-α） with 100 ng/mL for different hours （2, 4, 8, and 24 hours）. The expression changes of IP3R1 mRNA and protein were detected by quantitative real-time polymerase chain reaction and immunoblotting. Several inhibitors including D609, U73122, PP1, safingol, rottlerin and non-radioactive protein kinase C （PKC） were used to examine the mechanism of signal transduction of TNF-α-regulated IP3R1 in HMCs.RESULTS： The levels of IP3R1 mRNA at 2 hours after TNF-α exposure were significantly enhanced and peaked at 8 hours in HMCs （P〈0.01）, then descended at 24 hours （P〈0.01）. The levels of IP3R1 protein at 4 hours after TNF-α exposure were obviously increased and peaked at 24 hours after TNF-α exposure （P〈0.01）. Compared to the control group, safingol （PKCa inhibitor） and D609 （phosphatidylcholine-specific phospholipase C inhibitor） significantly blocked the TNF-α- induced expression of IP3R1 mRNA （3.30±0.81 vs. 1.95±0.13, P〈0.05; 2.10±0.49, P〈0.01） and IP3R1 protein （3.09±0.13 vs. 1.86±0.39, P〈0.01; 1.98±0.02, P〈0.01）. TNF-α promoted PKCa activation with maximal PKCa phosphorylation that occurred 8 hours after stimulation measured by non-radioactive PKC assay, and the effect was markedly attenuated by pretreatment with D609 or safingol.CONCLUSION： TNF-α increased the expression of IP3R1 and this was mediated, at least in part, through the PC-PLC/PKCa signaling pathways in HMCs.展开更多
Background and objectives Proliferation of human vascular smooth muscle cells(VSMCs)induced by hyperinsulinemia is a very common clinical pathology.Extensive research has focused on PKC(Protein kinase C)-MAPK(mitogen-...Background and objectives Proliferation of human vascular smooth muscle cells(VSMCs)induced by hyperinsulinemia is a very common clinical pathology.Extensive research has focused on PKC(Protein kinase C)-MAPK(mitogen-activated protein kinase)intracellular signal transduction and the phenotypic modulation accompanied by reorganization of intracellular F-actins in VSMCs.Methods DNA synthesis,signaling of ERK1/2 MAPKs,and changes inα-smooth muscle(SM)actin and F-actin were studied in hypertensive and normotensive human arterial VSMCs exposed to insulin and PMA with and without the PKC inhibitor,GF109203X.Results Differences among cell types in MAPK signaling,α-SM actin,and F-actin isoforms in VSMCs harvested from the arteries of patients with essential hypertension(EH)and normotension(NT)were identified in response to insulin treatment.Proliferation and activation of MAPK were more pronounced in EH VSMCs than in NEH VSMCs.Insulin exposure decreased expression ofα-SM actin and was accompanied by rearrangement of intracellular F-actins in VSMCs,especially in the EH group.These effects were reversed by treatment with the PKC inhibitor.Conclusions Human mesenteric VSMCs of EH and NT patients differed in proliferation,MAPK signaling,and degree of changes inα-SM actin and F-actin isoforms immediately following insulin exposure in vitro.展开更多
The effect of lipopolysaccharide (LPS) on the exposure of platelet fibrinogen receptors was investigated. The results showed that:l)LPS increased the binding of fibrinogen-gold complexes to platelets and the labels we...The effect of lipopolysaccharide (LPS) on the exposure of platelet fibrinogen receptors was investigated. The results showed that:l)LPS increased the binding of fibrinogen-gold complexes to platelets and the labels were primarily limited to shape-changed platelets; 2) LPS caused a dose-dependent rise in intracellular Ca2+ concentration in platelets; 3)LPS induced the activation of platelet protein kinase C (PKC) and the phosphorylation of glycoprotein llla (GPllla) which was inhibited by H-7. All these results suggest that stimulation of platelets with LPS causes a conformational change in glycoprotein llb/llla (GPllb/llla)through platelet shape change and /or phosphorylation of GPllla via PKC, which serves to expose the fibrinogen binding sites of GPllb/llla on human platelets.展开更多
基金supported by a grant from Health Bureauof Jiangxi Province
文摘BACKGROUND： This study aimed to explore the effects of TNF-α on the expression of IP3R1 mRNA and protein in human mesangial cells （HMCs）, and to elucidate the mechanism of TNF-α relating to IP3R1 expression in the occurrence of hepatorenal syndrome （HRS）.METHODS： HMCs were stimulated by tumor （TNF-α） with 100 ng/mL for different hours （2, 4, 8, and 24 hours）. The expression changes of IP3R1 mRNA and protein were detected by quantitative real-time polymerase chain reaction and immunoblotting. Several inhibitors including D609, U73122, PP1, safingol, rottlerin and non-radioactive protein kinase C （PKC） were used to examine the mechanism of signal transduction of TNF-α-regulated IP3R1 in HMCs.RESULTS： The levels of IP3R1 mRNA at 2 hours after TNF-α exposure were significantly enhanced and peaked at 8 hours in HMCs （P〈0.01）, then descended at 24 hours （P〈0.01）. The levels of IP3R1 protein at 4 hours after TNF-α exposure were obviously increased and peaked at 24 hours after TNF-α exposure （P〈0.01）. Compared to the control group, safingol （PKCa inhibitor） and D609 （phosphatidylcholine-specific phospholipase C inhibitor） significantly blocked the TNF-α- induced expression of IP3R1 mRNA （3.30±0.81 vs. 1.95±0.13, P〈0.05; 2.10±0.49, P〈0.01） and IP3R1 protein （3.09±0.13 vs. 1.86±0.39, P〈0.01; 1.98±0.02, P〈0.01）. TNF-α promoted PKCa activation with maximal PKCa phosphorylation that occurred 8 hours after stimulation measured by non-radioactive PKC assay, and the effect was markedly attenuated by pretreatment with D609 or safingol.CONCLUSION： TNF-α increased the expression of IP3R1 and this was mediated, at least in part, through the PC-PLC/PKCa signaling pathways in HMCs.
基金This work was supported by grants from the National Science Foundation of China(No.30170384 and No.30570764)
文摘Background and objectives Proliferation of human vascular smooth muscle cells(VSMCs)induced by hyperinsulinemia is a very common clinical pathology.Extensive research has focused on PKC(Protein kinase C)-MAPK(mitogen-activated protein kinase)intracellular signal transduction and the phenotypic modulation accompanied by reorganization of intracellular F-actins in VSMCs.Methods DNA synthesis,signaling of ERK1/2 MAPKs,and changes inα-smooth muscle(SM)actin and F-actin were studied in hypertensive and normotensive human arterial VSMCs exposed to insulin and PMA with and without the PKC inhibitor,GF109203X.Results Differences among cell types in MAPK signaling,α-SM actin,and F-actin isoforms in VSMCs harvested from the arteries of patients with essential hypertension(EH)and normotension(NT)were identified in response to insulin treatment.Proliferation and activation of MAPK were more pronounced in EH VSMCs than in NEH VSMCs.Insulin exposure decreased expression ofα-SM actin and was accompanied by rearrangement of intracellular F-actins in VSMCs,especially in the EH group.These effects were reversed by treatment with the PKC inhibitor.Conclusions Human mesenteric VSMCs of EH and NT patients differed in proliferation,MAPK signaling,and degree of changes inα-SM actin and F-actin isoforms immediately following insulin exposure in vitro.
文摘The effect of lipopolysaccharide (LPS) on the exposure of platelet fibrinogen receptors was investigated. The results showed that:l)LPS increased the binding of fibrinogen-gold complexes to platelets and the labels were primarily limited to shape-changed platelets; 2) LPS caused a dose-dependent rise in intracellular Ca2+ concentration in platelets; 3)LPS induced the activation of platelet protein kinase C (PKC) and the phosphorylation of glycoprotein llla (GPllla) which was inhibited by H-7. All these results suggest that stimulation of platelets with LPS causes a conformational change in glycoprotein llb/llla (GPllb/llla)through platelet shape change and /or phosphorylation of GPllla via PKC, which serves to expose the fibrinogen binding sites of GPllb/llla on human platelets.
