The Function And Mechanism Of PSTPIP2 In Alcoholic-induced Liver Injury

Background: Chronic liver disease caused by alcohol use disorders significantly increases the global burden of disease and mortality.Alcoholic liver disease(ALD)is the main cause of chronic liver disease.The occurrence of ALD is accompanied by a series of liver injuries,including fatty liver,alcoholic hepatitis(AH),liver fibrosis,cirrhosis and liver cancer,which are characterized by metabolic disorders,new lipidogenesis,steatosis and inflammatory reaction.Liver macrophages play a crucial role in alcohol-induced inflammation.When the injury occurs,they can be activated and release pro-inflammatory cytokines and chemokines,and recruit a large number of mononuclear macrophages from bone marrow to the liver,thus triggering liver inflammation.In addition,a large number of literatures have reported the possible mechanism of macrophages regulating inflammatory response,but currently there is no effective treatment for alcoholic-induced liver injury(ALI).Therefore,it is very important to elucidate the mechanism of ALI in order to reduce the alcoholic-induced liver injury and inflammatory response.Macrophage-mediated inflammation is associated with DNA methylation,and some studies have found abnormal patterns of DNA methylation in liver diseases.DNA methylation plays an important role in inhibiting gene function and the progression of liver injury.DNA methylation is usually catalyzed by DNA methyltransferases(DNMTs),including DNMT1,DNMT3 A,and DNMT3 B.Team early representative bisulfite sequencing(RRBS)results showed that PSTPIP2(chr18:77,843,840-77,843,968)in 5’-UTR region appear high methylation.PSTPIP2 is a member of the F-BAR protein family that is mainly expressed in macrophages.In macrophages,PSTPIP2 can negatively regulate the activation of macrophages and inhibit the release of inflammatory factors,thus inhibiting the process of inflammation.Therefore,in this study,we mainly study the role of methylated PSTPIP2 in macrophages’ regulation of inflammatory response and the molecular mechanism that may lead to ALI,which may provide a new therapeutic target for the treatment of ALI and reduce alcohol-induced liver injury and inflammatory response.Methods: In vivo,60 male C57BL/6J mice in adaptive breeding a week later,the mice were randomly divided into 6 groups(n = 10),respectively,the control group,model group,AAV9-empty-control group,AAV9-empty-model group,AAV9-PSTPIP2-control group and AAV9-PSTPIP2-model group.The alcoholic liver injury model of mice was established by the method of chronic alcoholic liver injury.Then,the liver tissues,blood samples and primary liver macrophages of mice were collected for subsequent experiments.First,the model of alcoholic liver injury was successfully established,and then the expression trend of PSTPIP2 was detected by western blot,real time-PCR and immunofluorescence.Furthermore,the regulation of specific overexpression of PSTPIP2 on alcohol-induced liver injury and inflammation was further investigated.In vitro,RAW264.7 cells were stimulated with Et OH and LPS,and the expression of PSTPIP2 in the model group was firstly detected.PSTPIP2 was then silenced in vitro to verify the role of PSTPIP2 in the inflammatory response induced by Et OH and LPS.Finally,western blot,MSP,Ch IP and other experiments verified that the down-regulation of PSTPIP2 was related to DNA hypermethylation in alcoholic liver injury.Results: In vivo,the expression of PSTPIP2 was down-regulated in the mouse model of alcoholic liver injury,and the overexpression of liver-specific PSTPIP2 alleviated alcohol-induced liver injury,macrophage infiltration and inflammatory response.In vitro,the silenced expression of PSTPIP2 on RAW264.7 cells aggravated the Et OH and LPS-induced inflammatory response.It was verified that PSTPIP2 plays an important role in macrophage-induced inflammatory response by regulating STAT1 and NF-κB signaling pathways.Moreover,the low expression of PSTPIP2 is associated with DNA methylation in alcoholic liver injury.Conclusion: DNMT3a-mediated PSTPIP2 methylation modulates alcohol-induced liver injury and inflammation by regulating STAT1 and NF-κB signaling pathways.