CHOP Activates Macrophages And Promotes Liver Fibrosis In Schistosoma Japonicum-Infected Mice

Background:Schistosomiasis,an ancient parasitic disease,threatens the health of an estimated 200 million people in more than 74 countries.Liver fibrosis is a main symptom of schistosomiasis japonica in chronic stage,which persists and eventually leads to complications such as cirrhosis,hepatic portal hypertension and even death.Praziquantel is currently the first-line drug for the treatment of schistosomiasis.However,the exact mechanism of action of praziquantel is not clear and drug resistance has been reported.Therefore,preventing,alleviating and even reversing liver fibrosis caused by Schistosoma japonicum(S.japonicum),improving the quality of life of patients,and reducing the mortality rate of patients are still urgent problems that need to be solved urgently.Liver macrophages are the main source cells of schistosomiasis granuloma,soluble egg antigen(SEA)induces alternative activation macrophages,called M2 macrophages.In the acute phase,M2 macrophages prevent acute inflammatory lesions induced by eggs via reducing the production of pro-inflammatory factors.However,in the chronic phase,M2 macrophages secrete profibrotic cytokines,leading to liver fibrosis.Numerous studies have confirmed that M2 macrophages play a crucial role in the formation of chronic liver granuloma and hepatic fibrosis.CHOP(CCAAT/enhancer-binding homologous protein)is a specific transcription factor for endoplasmic reticulum stress(ER stress),which is closely related to the formation of fibrosis.The reduction of infiltration of M2 macrophages was observed in the CHOP-/-animal fibrosis model,which was reflected in the decreased differentiation of M2 macrophages,suggesting that macrophages may change their phenotype by regulating their activation direction through CHOP and further participate in the disease process of tissue fibrosis.However,the role of CHOP in liver fibrosis caused by S.japonicum has not been reported.In this study,CHOP was used as the research target.C57BL/6 mice infected with S.japonicum cercariae were used to induce mice hepatic fibrosis model,which was to confirm that CHOP was involved in the polarization of macrophages induced by S.japonicum,to explore the effect of CHOP on liver fibrosis and its mechanism,and to provide a new potential target for the development of drugs to prevent liver fibrosis.Objective:To investigate the role of CHOP protein in macrophage polarization induced by S.japonicum and its effect on liver fibrosis and its mechanism,and to provide a new potential target for drugs to prevent liver fibrosis.Methods:Animal model of schistosomiasis japonica liver fibrosis was established.:eight-week-old SPF C57BL/6 mice with half the number of male and female were randomly divided into control group and infected group.Mice in the infected group were infected with S.japonicum cercariae through abdominal skin.Mice were euthanatized at 6 and 10 weeks after infection,and liver and serum samples were collected.(1)Paraffin sections were prepared for pathological HE staining and MASSON collagen fiber staining,and the average area of egg granuloma and collagen fiber expression were analyzed.(2)Changes of CHOP and Fibronectin protein expression in liver of mice at 6 and 10 weeks after S.japonicum infection were detected by immunohistochemical assay.(3)The expression of CHOP in M2 macrophages in liver of mice at 6 and 10 weeks after S.japonicum infection was observed by immunofluorescence assay.(4)Liver protein was extracted,and CHOP,KLF4,STAT6 and pSTAT6 proteins in the liver of mice at 6 and 10 weeks after S.japonicum infection were detected by Western Blot.(5)Liver RNA was extracted,and the expression of CHOP,STAT6,KLF4,IL-13Ra and IL-4Ral in the liver at 6 and 10 weeks after S.japonicum infection were detected by RT-qPCR.(6)The serum of mice was collected and the expression of IL-13 and TNF-α protein content in serum of mice at 6 and 10 weeks after S.japonicum infection were detected by ELISA.Results:Compared with the control group,mice infected with S.japonicum cercariae after 6 and 10 weeks:(1)Liver/body weight ratio(LBWR,LBWR(%)=wet weight of mouse liver/body weight x 100%)was increased significantly(P<0.05,P<0.01).(2)Liver pathological changes showed that the average area of egg granuloma in liver tissue was significantly increased(P<0.01),and the collagen deposition in the liver increased in a time-dependent manner(P<0.01).(3)The expression of collagen fibers in liver tissue increased(P<0.01).(4)The expression of CHOP mRNA and protein levels increased in a time-dependent manner(P<0.01,).(5)The macrophage infiltration in the liver of mice showed a time-dependent manner trend,showing a significant increase in the number of CHOP+F4/80+macrophages,mainly CHOP+CD206+ macrophages.(6)The expression of KLF4 protein in mice liver was decreased(P<0.01),pSTAT6 protein expression increased(P<0.01),IL-13Rα1 mRNA expression at 6 and 10 weeks after infection increased(P<0.05,P<0.01),IL4Ra mRNA expression in the 10 weeks after infection increased(P<0.01),6 weeks after infection has no obvious change.(7)The peotein content of TNF-α at 6 and 10 weeks after infection increased(P<0.01).The peotein content of IL-13 in the 6 weeks after infection increased(P<0.01),10 weeks after infection has no obvious change.Conclusion:(1)CHOP promoted liver fibrosis caused by S.japonicum.(2)CHOP may mediate liver fibrosis induced by S.japonicum by promoting selective activation of macrophages.