The Role Of Activin A In CCl₄-induced Acute And Chronic Liver Injury In Mice

Activin, a member of the TGF-β super family, has been demonstrated to beresponsible for many biological function. It plays important biological roles onvarious organs and systems. Activin receptors have been proved as one kind ofserine/threonine （Ser/Thr） kinase receptors families. Activin directly combines withtype II receptor （ActRII）, then couples type I receptor （ActRI）, and cascades Smadproteins reaction, which will contribute to the signaling transduction to activation oftarget gene in nucleus.Activin contributes to liver tissue fibrosis through inducing hepatocytesapoptosis and production of extracellular matrix （ECM） derived from activatedhepatic stellate cells （HSC）. Activin has three isoforms, activin A （βAβA）, activin B（βBβB） and activin AB （βAβB）. Activin A has been indicated to liver diseases due toabnormal expression levels of activin A in virus hepatitis and liver fibrosis, as wellas hepatocellular carcinoma. The increased expression of activin A in HSC may bethe main reason of liver fibrosis. However, the effects of activin A and itssignaling transduction in carbon tetrachloride （CCl₄）-induced acute and chronic liverinjury are still unclear.In order to explore the expression and role of activin A and its signalingpathway in liver，acute and chronic liver injury of C57BL/6mouse model wereinduced by CCl₄. The expressions of activin A, activin type IIA receptor（ActRIIA）,Smad2and Smad3in liver tissues of acute and chronic liver injury models wereanalyzed by the enzyme linked immunosorbent assay （ELISA）,immunohistochemical staining and real-time quantitative PCR （qRT-PCR）. Further,the role of activin A in the process of acute liver injury was confirmed by blockadeof activin A action with anti-activin A and anti-ActRIIA antibodies in vivo. Preparation of CCl₄-induced model mouse of acute and chronic liver injuryAcute and chronic liver injury in C57BL/6male mice were induced byintraperitoneal injection with carbon tetrachloride （CCl₄）（0.5ml/kg·BW） dissolvedin olive oil （1:19V/V）. Mice were sacrificed1,3,5and7days after the treatment.The activities of alanine aminotransferase （ALT） and aminotransferase （AST） inserum were examined and pathological changes of the liver were observed byhematoxylin and eosin staining to evaluate the liver injury extent. The above dataindicated that CCl₄-induced acute and chronic liver injury models were successfullyestablished.The expressions of activin A, ActRIIA,, Smad2and Smad3in livers ofCCl₄-induced mice acute and chronic liver injury modelsThe results of ELISA and real-time quantitative PCR in acute and chronic liverinjury models showed that activin A mRNA and protein expression levelssignificantly increased in liver tissues of CCl₄-induced liver injury model of miceafter1day injury, followed by gradually recovering to normal, which were similar tothe changes of transaminase levels in serum. The abnormal level of activin A may beinvolved in the CCl₄-induced liver injury model, reminding potential relation to liverinjury severity. The mRNA transcription levels of ActRIIA, Smad2and Smad3wereincreased in both models. ActRIIA mRNA, Smad3mRNA and activin A expressionlevel presented positive correlation, which were parallel with ALT and AST levels inserum. These results suggested that activation of activin-ActRIIA-Smad signalingpathway is involved in CCl₄-mediated acute and chronic liver injury.To determine whether the activin A mediated CCl₄-induced acute liver injury,further experiments were conducted using anti-activin A and anti-ActRIIA antibodiesto block endogenous activin A. We injected anti-activin A or anti ActRIIA antibodiesvia tail vein2hours after intraperitoneal injection of CCl₄. The results showed thatafter CCl₄injection, the serum levels of ALT and AST were significantly elevated.But after using anti-activin A antibody to combine with endogenous activin A or block the combination between activin A and its receptor with anti ActRIIA antibody,the serum levels of ALT and AST were significantly decreased. HE staining showedthat liver tissue injury was less severe in anti-activin A antibody and anti-ActRIIAantibodies blocking experiments. These results further confirmed activin A-ActRIIApathway was involved in CCl₄-induced acute and chronic liver injury in mice.In summary, the high expression of activin A may be an important potentialfactor leading to liver injury, and the activin A-ActRIIA-Smad signaling pathwaywas involved in CCl₄-induced acute and chronic liver injury. Therefore, blocking theactivin A action may be an effective method for the prevention and therapy of liverinjury.