The Effects Of Angiotensin-(1-7) On Rat Hepatic Stellate Cell Contraction Mediated By Rho-Rock Pathway

BackgroundIt is well known,various kinds of chronic liver disease can develop into hepatic cirrhosis from hepatic fibrosis,while portal hypertension is the serious complication of hepatic cirrhosis.Increased intrahepatic vascular resistance is the important factor under the state of hepatic cirrhosis.Hepatic stellate cells is the major cells within extracellular matrix,its activation plays a key role in the trail process of the occurrence and development of hepatic fibrosis as well as the portal hypertension formation.The activated HSC can convert to myofibroblast and fibroblast,and is able proliferate and gain contractility which results in the diminution of the anteroposterior diameter of sinus hepaticus,and then,the intrahepatic vascular resistance will increase.On the other hand,activated HSC can cause the hepatic tissue cicatrice to contract,and increase the intrahepatic vascular resistance progressly,and result in portal hypertension finally.Therefore,the regulatory mechanism for the contraction of HSC is the important component element for the research to the mechanism of portal hypertension.Recently,renin- angiotensin- aldosterone system(RAAS) intrahepatic is the hot spot of investigation for hepatic fibrosis,With the awareness of the constant deepening of RAAS,RAAS discovered many new components,such as angiotensin-converting enzyme2(ACE2),angiotensin-(1-7)(Ang-(1-7)), Ang(1-7)receptor Mas and so on.They constitute the ACE2-Ang(1-7)-Mas receptor axis has become another important branch of RAAS,the ACE-Angâ…¡-AT1R axis significantly inhibited.Intrahepatic ACE-Angâ…¡-AT1R axis and the formation of portal hypertension have a close relationship.At present,there are no report between ACE2-Ang(1-7)-Mas receptor axis and HSC.contraction.Activated HSC into myofibroblast-like cells with characteristics of smooth muscle cells,suggesting that HSC contraction and contraction of smooth muscle cells of the molecular mechanisms are similar and can be calcium-dependent and non-calcium-dependent contraction of the two pathways to complete.In the non-calcium pathway,Rho-Rock pathway on HSC contraction control mechanism is a hot research in recent years.In this study,by observing the Ang-(1-7) on HSC contraction and Rho-Rock signaling pathway on.the expression of various components to clarify its regulation of HSC contraction.Objective To investigate the effects of Angiotensinl-7 on rat hepatic stellate cells contraction mediated by Rho-Rock pathway.Methods HSC-T6 cell line was under pre-disposal treatment with Ang-(1-7) 1μmol/L.The cell contraction was detected by silicone-rubber- membrane cultivation directly.Besides,HSC-T6 cell. line was under pre-disposal treatment with Ang-(1-7) 10μmol/L.And the protein level of MLC,phosphorylation MLC andα-Smooth muscle actin(α-SMA )were detected by Western blotting.Then observated the difference of phosphorylation MLC andα-SMA protein level after using the blocking agent of Ang-(1-7),Angâ…¡(angiotensinâ…¡),Ang-(1-7) mas receptor-A779,Angâ…¡and Ang-(1-7),respectively.RT-PCR was used to detect the expression of Rock2, RhoAGTP and RhoGEF in Ca²⁺-independent pathways mediated by Rho-kinase. Results Ang-(1-7) inhibited Angâ…¡-induced HSC contraction;Ang-(1-7) inhibited Angâ…¡-induced phosphorylation MLC andα-SMA protein expression. Ang-(1-7)+Angâ…¡inhibited Angâ…¡-induced phosphorylation MLC level,Ang-(1-7) and Angâ…¡treatment phosphorylation MLC protein expression markedly increased. And Ang-(1-7) phosphorylation MLC protein levels were significantly higher than Ang-(1-7)+A779(P = 0.000).Ang-(1-7)+Angâ…¡inhibited Angâ…¡-inducedα-SMA protein expression level,Ang-(1-7) and Angâ…¡phosphorylation MLC protein expression markedly increased.And Ang-(1-7)α-SMA protein level was significantly higher than that Ang-(1-7)+A779(P=0.000).Ang-(1-7)+Angâ…¡deal with the expression of Rock2,RhoAGTP,RhoGEF mRNA significantly reduced. Ang-(1-7)+A779 treatment Rock2,RhoAGTP,RhoGEF mRNA expression decreased, Ang-(1-7) and Angâ…¡Rock2,RhoAGTP,RhoGEF mRNA expression markedly increased.Conclusion Ang-(1-7) could reduce Angâ…¡-induced HSC contraction.