The Rapeutic Effect And Mechanism Of Qingre Jiedu Liangxue Recipe On Mice With Acute-on-Chronic Liver Injury

Objective:To explore the mechanism of Qingrejiedu Liangxue Recipe on the mice models of acute-on-chronic liver injury.Methods:1.To study animal models of acute-on-chronic liver injury:Fifty male C57BL/6 mice were randomly divided into eight groups(A-D;E-H).Group A was given high-fat diet for 3 months;group B and C were given intraperitoneal injection of 10%CCL4(dissolved in olive oil)twice a week for 8 weeks and 12 weeks;group D was given biliary ligation for 2 weeks;group E was given a dose of 53%ethanol on the basis of group A;group F was given LPS 100ug/kg combined with D-gal 400mg/kg on the basis of group B;Group G was given the same dose of LPS and D-gal on the basis of Group C;Group H was given the same dose of LPS and D-gal on the basis of Group D.Serum Tbil was measured by chemical oxidation;Serum ALT?AST?IL-6?TNF-? were measured by double antibody sandwich enzyme-linked immunosorbent assay;Hepatic tissue pathological changes observed by HE and Sirius red staining.2.Study on the intervention effect of Qingre Jiedu Liangxue Recipe:Thirty-six C57BL/6 male mice were randomly divided into normal group?model group?Chinese medicine group and western medicine group.The model group was intraperitoneally injected with 10%CCL4 three times a week for 8 weeks and then given LPS 0.5mg/kg and D-gal 400mg/kg at the last time and then sacrificed at 3h?12h and 24h.The Chinese medicine group and western medicine group started receiving medical treatement after intragastric administration of 10%CCL4 for 4 weeks and then injected by LPS and D-gal after they received medical treatement for another 4 weeks and finally sacrificed at 3h.Serum ALT?AST were measured by double antibody sandwich enzyme-linked immunosorbent assay;hepatic tissue pathological changes observed by HE and Sirius red staining;and serum inflammatory factors were detected by LEGENDplex test kit;F4/80 and CDllb were observed by immunofluorescence;neutrophils expression were observed by immunohistochemistry;CD44?ICAM and HMGB1 were detected by Western blotResults:1.Compare and establish a mouse model of acute-on-chronic liver injury:Group E(HFD+Alc.)showed mild hepatic fibrosis and mild inflammatory factor infiltration and mild liver injury;group B(BDL+LPS+D-gal)showed a large number of inflammatory factors infiltrating and sub-macro necrosis.ALT,AST and Tbil were significantly increased(P<0.0001),however,mice death occurred during BDL process(2/7);group F(CCL4-8w+LPS+D-gal)and group G(CCL4-12w+LPS+D-gal)showed obvious liver fibrosis and severe hepatocyte necrosis.In the process of chronic liver injury,no death was observed in mice.Compared with group F(CCL4-8w+LPS+D-gal),the level of ALT?AST?IL-6 and TNF-? were significantly elevated in the group G(CCL4-12w+LPS+D-gal)(P<0.05).HE staining showed that liver tissue pathological damage was also more serious.2.Study on the intervention effect of Qingre Jiedu Liangxue Recipe:Compared with the normal group,ALT and AST were increased in the model group(P<0.001);GM-CSF?IFN-??IL-1??IL-6?TNF-??IL-10?MCP-1?IL-17A?IL-1? and IL-27 were increased(P<0.05);the expression of MPO?F4/80?CD11b?CD44?ICAM-1 and HMGB1 were significantly increased(P<0.05).Compared with the model group,ALT and AST of the western medicine group were significantly decreased(P<0.01);GM-CSF?IFN-??IFN-? were decreased(P<0.05).Compared with the model group,IL-17A in the serum of the Chinese medicine group was significantly increased(P<0.01),and ALT?AST?TNF-??GM-CSF?IL-10?IL-1??MCP-1 and IFN-? were decreased(P<0.05);MPO?CD44?ICAM-1?HMGB1?F4/80?CD11b were increased.Compared with the western medicine group,TNF-??IL-1??MCP-1?IL-10 and IL-6 were decreased in the Chinese medicine group(P<0.05).Conclusion:1.A mouse model of acute-on-chronic liver injury was established:10%CCL4(3 times a week for 8 weeks)+LPS(0.5 mg/kg)+D-gal(400 mg/kg).2.Qingre Jiedu Liangxue might reduce the mortality of mice of acute-on-chronic liver injury,however,it is necessary to increase the sample size for further verification.3.Qingre Jiedu Liangxue Recipe could improve liver function indexes(ALT,AST),reduced inflammatory factors(TNF-?,GM-CSF,MCP-1)and increased IL-17A.4.Qingre Jiedu Liangxue Recipe could increase the expression of F4/80 and CDllb and decrease the expression of IL-1?,ICAM-1,IL-10,CD44,MPO and HMGB1 in a mouse model of chronic acute liver injury.It is indicated that Qingre Jiedu Liangxue Recipe might improve liver function by regulating neutrophils and Kupffer cells,and its specific mechanism needs further investigation.