Expression And Role Of Activin A And Its Binding Protein FS In Mouse Acute Liver Injury Induced By Con A

Activin, a member of TGF-βsuperfamily, plays important roles in cell differentiation,proliferation and apoptosis in multiple tissues. FS, an activin-binding protein, which bindswith activin with high affinity, involves in the multiple biological effect of activin. Ourresearch focused on the expressions, roles, and mechanisms of activin A and FS in liverinjury induced by concanavalin A with the methods of ELISA, PCR, immunohistochemistrytechnique, etc. Based on this study, we will provide new ideas for therapy and prevention inliver injury disease.Con A-induced acute liver injury in miceIn this experiment, ConA (15mg/kg) was injected for one time via tail vein of C57BL/6mice to prepare the liver injury models. On1d,3d,5d and7d after the administration ofConA, Serum and liver tissues of the actue liver injury model were collected. On1d afterinjection with ConA, the HE stain results showed that there were many focal necrosis amongwhich were plenty of lymphocytic infiltration. The normal hepathic lobule structure andhepatic cords disappeared and the levels of serum transaminase approached the peak. On3d,the areas of necrosis reduced, but there were still many infiltrated-lymphocytes among theinjury liver tissues with HE staining. The levels of, serum transaminases remained at arelatively high level compared with1d. On5d, the hepathic lobule structure and hepaticcords were almost recovered, but infiltrated-lymphocytes still could be found in somenecrosis areas. On7d, the necrosis liver tissues were nearly recovered to normal. The aboveexperiment indicated that we had successfully established the Con A-induced acute liverinjury model.Expressions of activin A, ActRIIA, FS in serum and liver tissue ofacute liver injury modelELISA results showed activin A levels in serum on1d after administration of Con Awas the highest, then gradually decreased and recovered to normal levels on7d.. But thelevers of FS did not change significantly on1d and3d. On5d and7d, the levels of FS wereincreased obviously and maintained at a high level. PCR results showed that expressions ofactivin A and ActRIIA mRNA in the liver tissues of mice model was in a parallel state,similar to the level of activin A in serum. The expression of FS mRNA began to increase on3d and then maintained at a high level. The immunohistochemical staining showed that the expression of activin A in normal liver tissue was only detected in a small amount in the liverparenchyma. But on1d after injected with ConA, the expression of activin A increased andbegan to decreased significantly on3d. The expression of ActRIIA was similar to activin A.Compared with the control group, the expression of FS on1d did not change significantly.On3d the expression began to increase and maintained at a high level. All of these suggestedthat activin A played a direct impact on the severity of liver injury in Con A-induced acuteliver injury model, and FS was involved in the process of impair of liver tissue after liverinjury.Effect of FS on liver injury in vivoTo investigate the role of activin A during Con A-induced acute liver injury, FS was injectedvia tail vein to neutralize the endogenous ctivin A. C57BL/6mice were randomly dividedinto four groups, the control group, the FS group, Con A group, and Con A+FS group. Themice were killed after given Con A via tail vein for1d. Levels of Serum transaminase in ConA+FS group was significantly lower than those in Con A group (p <0.01). HE stainingshowed the liver damage in Con A+FS group was decreased compared with Con A group,and the liver index in Con A+FS group was higher than that of Con A group. These resultssuggested that FS could reduce liver injury in mice.Effect of Smad3gene Knock-down on liver injuryTo investigate the specific role of activin A, Smad3gene was knocked down to blockactivin A intracelluar signal transduction. There were four groups, pGCsi-U6/Neo-shSmad3group, pGCsi-U6/Neo control group, the ConA+pGCsi-U6/Neo and ConA+pGCsi-U6/Neo-shSmad3gene Knock-down group.24h after the injection of plasmid viamouse tail vein, ConA was injected by the same method to induce mouse liver injury. Proteinlevel of activin A in serum detected by ELISA showed that ConA+pGCsi-U6/Neo-shSmad3gene silence group was significantly lower compared with ConA+pGCsi-U6/Neoexperimental group (p <0.01), while the FS protein levels were significantly increased(p<0.05). But serum transaminases ALT and aspartate aminotransferase AST levels wereequally reduced. HE staining showed that the degree of liver injury in ConA+pGCsi-U6/Neo-shSmad3group, was significantly reduced compared to ConA+pGCsi-U6/Neo group. All the results suggested that Smad3gene Knock-down couldsignificantly reduced the severity of liver injury induced by Con A by blocking the activinsignal transduction. These results indicated that activin A and activin-binding protein FS were involved inthe processes of Con A-induced liver injury and damage repair. Blocking activin A orinterfering with the signal transduction of activin can protectthe liver tissue from ConA-induced liver injury or therapy liver injury.