Grp75 Protects The Liver By Oxidative Damage By Improving Mitochondrial Function

Liver fibrosis is a result of chronic hepatocellular damage due to a variety of liver diseases including viral hepatitis B, hepatitis C and alcoholic and non-alcoholic fatty liver diseases. Thus, the chronic process of liver fibrosis poses a major problem of health worldwide. Accordingly, prevention and treatment of fibrosis is expected to reduce the occurrence of liver cancer. However, there is currently no specific therapy. Increased deposition of extracellular matrix and reduced matrix degradation are the major characteristics of liver fibrosis. So curing the primary disease is an ideal approach but in many liver diseases.Heat shock protein (Hsp) family maintaining the intracellular homeostasis and proteins interaction integrity of the cell exists in almost all of the prokaryotes and eukaryotes. In the presence of heat shock proteins, DNA and protein molecules in cells execute their boilogical functions strictly according to biological theorem. Glucose regulated protein75(Grp75), a member of the heat shock protein family, has been shown to be involed in many important cellular processes, such as fuctioning as a molecular chaperon, aiding new translated polypeptide correctly folding, stabilizing cytoskeleton, involving in DNA replication and transcription as well as taking part in the regulation of intracellular signal transduction. More and more studies showed that Grp75can protect cells that under different kinds of stresses including glucose deprivation from injuring thorough several different pathways.Grp75is constitutive expression molecular chaperons which exist in many organs, such as heart, brain, lung, spleen.Grp75can be up-regulated by stress to protect cell and inhibit apoptosis process. However Grp75can not constitutive express in liver.So we transfer Grp75gene into cells and rat to up-regulate its expression, through its antioxidant effect and inhibition of hepatocyte apoptosis in treatment of liver fibrosis, we can provide a basis for future clinical gene therapy of hepatic fibrosis.There are different animal models of liver fibrosis including hepatotoxin-induced injury (CCL4, dimethylnitrosamine and thioacetamide), nutritional derangement (alcohol and high fat low choline diet), and cholestasis induced by bile duct ligation. The CCL4-injection model was chosen in our study mainly because it has high reproducibility and consistency and is widely accepted by researchers.In this study, we observed that Grp75inhibit oxidative damage factor induced liver injury both in vitro and in vivo.We analyzed the MTT,AST,ALT level to indentify the protection effect of Grp75from H2O2induced cell damage in HL-7702cells. The mitochondrial function were determined during H2O2induction because Grp75lodges predominantly in mitochondria, and mitochondria could be a conceivable target for GRP75to exert the protective effect.We examined the intracellular ATP level of hl-7702cells, to see whether the over expression of Grp75could release the ATP depletion during H2O2treatment. Our data showed that over expression of Grp75affect the alteration of intracellular ATP level upon H2O2treatment. HL-7702cells over expressing Grp75showed moderate cell damage than control cells, while the ATP level also show significant discrepancy. The similar condition was seen in the effect of over expression of GRP75on the mitochondrial potential of HL-7702cells. H2O2induced depolarization of mitochondrial potential progressively and over expression of Grp75prevent the collapse of mitochondrial potential upon H2O2exposure.It is widely accepted that ROS is produced predominantly in mitochondria as a by-product of oxidative phosphorylation. Our data demonstrated that overexpression of Grp75in HL-7702cells inhibited the ROS accumulation significantly upon H2O2exposure.It has been reported that the releasing of Cytochrome C may induce cell apoptosis. We examined Cytochrome C in cytoplasm to indentify whether Grp75can regulate the releasing of Cytochrome C. Our date showed that the Cytochrome C expression in overexpression of Grp75in HL-7702cells was obviously less than control cells upon H2O2exposure.Grp75ameliorated CCL4-induced hepatic inflammation in rats as evidenced by improved histological findings and reduced ALT and AST levels. We also analyzed the ATP level and Cytochrome C, to indentify the mechanism of inhibition in CCL4induction. The date showed the similar trend with cell culture.In conclusion, Grp75reduced hepatic inflammation induced by CCL4. It prevents the oxidative damage both in vitro and in vivo though ameliorating mitochondria function such as motivate ATP level, inhibit the ROS accumulation and inhibit the releasing of Cytochrome C. This study may contribute to the treatment of liver fibrosis.