Study On The Correlation Among The Expression Of Gremlin1,VEGFR2,CD31 And VWF In CCL4-induced Mouse Liver Fibrosis

Objective: It can be said that liver fibrosis is the basis of complications such as portal hypertension,ascites,and hepatic encephalopathy in the late stage of liver disease.Therefore,research on the mechanism of liver fibrosis had important clinical significance for the treatment and prevention of liver diseaseses.Gremlin1 played an important role in the occurrence of a variety of fibrotic diseases.Some research found that Gremlin1 was an important liver fibrosis promoting factor,and one of its mechanisms is achieved by antagonizing the BMP signaling pathway.Recently,it has been discovered that Gremlin1 was also a novel VEGFR2 agonist,which was related to angiogenesis-related diseases.Vascular endothelial growth factor receptor-2(VEGFR2)was a major angiogenic factor receptor expressing tyrosine kinase receptor in endothelial cells,and regulated angiogenesis and remodeling.Current studies had found that liver fibrosis is related to hepatic sinusoidal capillaries and intrahepatic angiogenesis.In the process of liver fibrosis,the endothelial window of the liver sinusoids gradually decreases or disappears,and the basement membrane under the endothelium forms,similar to continuous capillaries.Under normal circumstances,liver tissues hardly express continuous endothelial cell markers such as platelet endothelial cell adhesion factor(CD31)and factor Ⅷ-related antigen(v WF).Therefore,our study intended to explore the expression of Gremlin1,VEGFR2,CD31 and v WF in mouse liver fibrosis models,and their correlation mechanism of action,in order to provide new targets for clinical treatment of liver cirrhosis.Methods:(1)Establish model: 30 male C57 mice were randomly divided into normal control group and CCL4 model group,15 in each group.The CCL4 model group was given intragastrically 35% CCL4 olive oil solution(10ml/kg)twice a week for 8 weeks;the normal control group was given intraperitoneal injection of olive oil solution(10ml/kg)twice a week for 8 weeks.(2)Observe the degree of liver fibrosis with Sirius red staining;(3)Immunohistochemical staining analyzed the expression and distribution of Gremlin1,VEGFR2,CD31 and v WF in liver fibrotic tissues;(4)Western Blotting analyzed the expression and distribution of Gremlin1,VEGFR2,CD31 and v WF in liver fibrotic tissues;(5)Graph Pad Prism 7,Image Plus Pro statistical software was used for data analysis and mapping.Unpaired t-test was used between the two samples.The Pearson correlation coefficient was used to evaluate the degree of association between the two samples.For data analysis,P <0.05 means the difference was statistically significant,P <0.01 means the difference was statistically significant.Results:(1)The results of immunohistochemistry showed that the expression levels of Gremlin1,VEGFR2,CD31 and v WF in mouse liver fibrosis tissue were higher than those in the normal control group(P<0.0001,P<0.05).(2)In CCL4 mouse liver fibrosis,the expression level of Gremlin1 was positively correlated with VEGFR2,CD31 and v WF(P<0.0001).(3)In CCL4 mouse liver fibrosis,the expression level of VEGFR2 was positively correlated with CD31 and v WF(P<0.01).(4)Western Blotting results showed that the expression levels of Gremlin1,VEGFR2,CD31 and v WF in mouse liver fibrosis tissues were higher than those in the normal control group(P<0.05,P<0.01).Conclusion:The expressions of Gremlin1,VEGFR2,CD31 and v WF in mouse liver fibrosis tissues are positively correlated,suggesting that Gremlin1,VEGFR2,CD31 and v WF may be involved in the occurrence and development of liver fibrosis.