Study On The Mechanism Of Immune Regulation Effect Of Jiedu Huayu Ⅱ Decoction On Rats With Acute-on-chronic Liver Failure Based On Chemokine Ligand 20

Objective:1.To observe the expression of chemokine ligand 20（Chemokine ligand20,CCL20）in acute-on-chronic liver failure（ACLF）rats and its effect on the immune regulatory network of Th17 cell differentiation.2.Jiedu Huayu II Decoction on the impact of CCL20 in ACLF and the regulation of Th17 cell differentiation immune network.Methods:ACLF rat models were established with carbon tetrachloride（CCL₄）combined with D-galactosamine（D-Gal N）and lipopolysaccharide（LPS）.They were divided into Normal group（Normal）,liver cirrhosis group（LC）,ACLF group（Model）and Jiedu Huayu DecoctionⅡgroup（JDHY）.After the model was successfully established,the degree of fibrosis and necrosis of liver tissue were observed by Masson staining and HE staining,and the survival rate of each group was also observed.The content of CCL20 in liver tissue was detected by WB,while the proportion of Th17 in peripheral blood was detected by flow cytometry.The contents of CCL20,IL-17,IL-21,TNF-α,AST,and ALT were detected by ELISA.The serum TBiL content was detected by an automatic biochemical analyzer.Results:1.Survival rate:The survival rate was 100%in the Normal group,93.3%in the LC group,46.7%in the Model group and 76.7%in the JDHY group.Compared with the Normal group,the survival rate of the Model group was significantly reduced,and the difference was statistically significant（X²=10.1953,P<0.05）.Compared with the Model group,the survival rate of rats in the JDHY group was significantly increased,and the difference was statistically significant（X²=4.5123,P<0.05）.2.Pathology:In the Normal group,HE staining showed structurally intact hepatic lobules,no formation of pseudolobular nodules,and no degeneration and necrosis of nuclei.Masson staining in LC group showed that the structural integrity of the hepatic lobule was damaged,pseudolobular nodules were formed,and fibrous tissues in and around the portal area proliferated.In the Model group,HE staining showed that the structure of hepatic lobular was damaged and unclear,and most of the hepatocytes showed obvious degeneration and necrosis,as well as karyopycnosis,fragmentation and dissolution.The hepatic lobular structures were damaged to different extents in the JDHY group,and a small amount of karyopycnosis,fragmentation and dissolution were observed.Compared with the Model group,the degeneration and necrosis of hepatocytes in the JDHY group were milder.3.Compared with the Normal group,the expression levels of CCL20 in liver tissue was significantly increased in both the LC and Model groups（P<0.05）.In addition,the expression level of CCL20 in the Model group was significantly increased（P<0.05）.Compared with the Model group,the expression level of CCL20 in the JDHY group was significantly down-regulated and the difference was statistically significant（P<0.05）.4.Compared with the Normal group,the Th17 levels in the peripheral blood of the LC group and the Model group were significantly increased,and the differences were statistically significant（P<0.05）.Compared with the LC group,the Th17 content in the peripheral blood of the Model group was significantly increased and the difference was statistically significant（P<0.05）.Compared with the Model group,the Th17 content in peripheral blood of JDHY group was significantly decreased and the difference was statistically significant（P<0.05）.Compared with the Normal group,liver IL-17,IL-21 and TNF-αcontents in the LC and Model groups were significantly increased and the differences were statistically significant（P<0.05）.Compared with the LC group,the contents of IL-17,IL-21 and TNF-αin the liver tissue of the Model group were significantly increased and the differences were statistically significant（P<0.05）.Compared with the Model group,liver IL-17,IL-21 and TNF-αcontents in the JDHY group were significantly decreased with statistical significance（P<0.05）.5.Compared with the Normal group,the levels of AST,ALT and TBiL in the LC group and the Model group were significantly increased and the differences were statistically significant（P<0.05）.Compared with the LC group,the levels of AST,ALT and TBiL in the Model group were significantly increased and the differences were statistically significant（P<0.05）.Compared with the Model group,the levels of AST,ALT and TBiL in the JDHY group were significantly decreased and the differences were statistically significant（P<0.05）.Conclusions:1.CCL20 increased significantly in ACLF,which positively promoted the inflammation and necrosis of ACLF hepatocytes.The mechanism may be that Th17 cells are recruited to stimulate Th17 to secrete cytokines such as IL-17,IL-21,TNF-α,etc.,so as to enlarge the inflammation,thus aggravating the inflammation and necrosis of ACLF hepatocytes.2.Jiedu HuayuⅡDecoction may reduce the expression of CCL20 in ACLF,decrease the aggregation of Th17,and inhibit the activity of Th17/IL-17 inflammatory axis,thus reducing the immune inflammatory injury of ACLF and protecting liver function.