A Study On The Role Of ATF4 In Liver Fibrosis And Its Underlying Mechanism By Regulating Endoplasmic Reticulum Stress

Hepatic fibrosis is a reversible healing response in which the liver is stimulated by various pathogens such as drugs,viruses or ischemia and reperfusion,and is characterized by accumulation of extracellular matrix after liver injury.Liver fibrosis is the common pathology of many chronic liver diseases.Continuous progressive liver fibrosis may eventually lead to cirrhosis,liver failure,portal hypertension and even liver cancer,and eventually requires liver transplantation.Chronic viral hepatitis,toxins and drug effects,cholestasis,nutritional deficiencies,alcoholic liver disease caused by alcohol abuse and nonalcoholic fatty liver disease may lead to the occurrence of liver fibrosis,but the specific cell biology process and the potential molecular mechanism are still unclear.There are many problems that have not been cleared,therefore,it is urgent to clarify the key regulatory elements and intervention targets for liver fibrosis and related end-stage liver disease prevention and treatment.Liver cancer developed from liver fibrosis is the leading cause of liver disease related death.Due to the low early diagnosis of liver cancer and the lack of effective intervention measures,the median survival time for advanced liver cancer patients is only 6 months.The prognosis of hepatocellular carcinoma is very poor,with a 5-year recurrence rate as high as70%.Although there has been a lot of research on the progression of liver cancer,the existing classification system can not effectively define a certain subset of liver cancer and its individualized treatment due to high heterogeneity of liver cancer.Therefore,the study of hepatocellular carcinoma-related signal pathway in depth to explore the pathogenesis of hepatocellular carcinoma is of great significance,along with the screening and identification of liver cancer diagnositic markers as well as the development of new treatment.The microenvironment of chronic inflammation in the liver is considered to be one of the important causes of liver fibrosis,liver cirrhosis and even liver cancer.In the case of drug toxicity,viral infection,liver cell death and compensatory proliferation occurs repeatedly,during which chronic inflammation of the liver is often accompanied by endoplasmic reticulum stress.The endoplasmic reticulum is mainly responsible for the synthesis and secretion of protein.Under normal circumstances,the endoplasmic reticulum maintains a certain steady state.When unfolded or mistakingly folded protein accumulates too much beyond the process ability of endoplasmic reticulum,it will start the endoplasmic reticulum stress by promoting the synthesis of new proteins while reducing the general protein synthesis to cope with the stress.The three main signaling pathways for UPR response are inositol requiring?IRE?1?,PERK-eIF2?-ATF4-CHOP and ATF-6?.The UPR reaction could restore the equilibrium steady state through the autophagic degradation of protein and ER-related degradation?ERAD?to remove unfolded protein.Many studies have shown that ATF4 is a key regulator of unfolded protein response and endoplasmic reticulum stress.Its upstream eIF2?can also be phosphorylated and activated by regulatory molecules other than PERK,which can integrate viral infection,amino acid deficiency,anemia and other pathological processes in the signal pathway.ATF4 is involved in the pathogenesis of a variety of complex diseases including malignant tumors.We believe that the role of ATF4 and its regulatory target genes in liver fibrosis and liver cancer is worthy of further study.In this study,we are aimed to study the dynamic changes of ATF4 and its downstream molecules in hepatocyte injury,apoptosis,hepatic fibrosis and hepatocellular carcinoma by combined methods of ATF4 knockout mice,in vitro cell experiments and analysis of clinical specimens.We are also determined to evaluate the value of ATF4 as a molecular marker for prognosis of liver cancer.In this study,we used two liver fibrosis models--intraperitoneal injection of CCL₄ and BDL mouse model to demonstrat that liver fibrosis of ATF4 knockout mice was significantly lower than that of wild-type mice.Moreover,the mortality rate of BDL was significantly lower than the control group as ATF4 knockout reduced hepatic fibrosis.It was confirmed that the expression level of ATF4 in HCC was higher in tumor tissues compared with paratumor cells.Further mechanism study found that proliferation and apoptosis in ATF4 knockout mice are weaker than the control group.WB and RT-PCR method revealed decreased expression of ATF4will lead to the decrease of CHOP expression.Further cell proliferation and apoptosis detection demonstrated that the absence of ATF4 attenuated CHOP mediated apoptosis through PERK-eIF2?-ATF4-CHOP signal pathway.ATF4 might also play an important role in the development and progression of HCC,suggesting that it could be used as a prognosis biomarker for HCC.