To Study The Anti-fibrosis Mechanisms Of Qizhu Granules Based On The Microscale Biomechanical Properties Of Hepatic Sinusoidal Endothelial Cells

Liver fibrosis is a common disease in liver and biliary system diseases,and is the inevitable stage of the development of most chronic liver diseases.At present,the medical community generally believes that early liver fibrosis can be reversed,but how to reverse,has not yet formed a general consensus.Many pathological changes in the early stages of liver fibrosis,the pathological changes of sinusoidal endothelial cell(SECs)drawing attention increasingly,its characteristic cytologic behavior change including the change of "fenestration" size and number,and the formation of the basement membrane.once SECs undergo capillarization,the process of liver fibrosis is irreversible.It is indisputable fact that at present.For the treatment of liver fibrosis,modern medicine can only give etiological treatment.As to liver fibrosis itself,there is no positive effect of drugs or treatment.Traditional Chinese medicine(TCM)has a long history of experience in the treatment of liver fibrosis,and has formed a complete understanding of its pathogenesis.In particular,early intervention for chronic liver disease under the guidance of the idea of "preventive treatment of disease"can reduce the risk of liver fibrosis in patients in the future to a certain extent.Qizhu granule is professor naili-Yao's work,which combined his years of experience and consensus in TCM community about liver fibrosis.This research "Based on the microscale biomechanical properties of hepatic sinus endothelial cells to research Qizhu granule against liver fibrosis mechanism"(NO:81774282)is funded by National Natural Science Fund,with the early results from our team to research the mechanism of Qizhu granule against liver fibrosis.1.Meta-analysis based on the clinical efficacy of the method of YiQi and HuoXue in the treatment of liver fibrosisObjective:To evaluate the clinical efficacy of YiQi and HuoXue in the treatment of liver fibrosis by Meta-analysis.Methods:1.PubMed,China Knowledge Infrastructure Project and Wanfang Data Knowledge Service Platform were used as retrieval databases,and the retrieval time span was from self-built database to December 31,2020;The search words "liver fibrosis" and"hepatic fibrosis" were used as the main line,and then "effect" or "efficacy" was used as the key words to exclude each entry."Liver fibrosis”was used as the search term,and then "clinical observation" and "therapeutic effect analysis" were used as the key words to exclude one by one.The above databases have no language restrictions.2.Literatures of randomized controlled clinical studies was included,and outcome indexes such as four items of liver fibrosis,instantaneous liver elastography,liver and spleen morphology,portal and splenic vein width,adverse reactions,safety and other indicators were used as assessment indexes.Results:A total of 93 qualified literatures were included in this study,86 of which took four items of liver fibrosis as clinical evaluation indexes.The comparison between the two groups showed that the clinical efficacy of the Chinese herb group was better than that of the control group,with statistical difference[MD=51.15,95%CI(56.95,45.35),P<0.00001].In 15 literatures,the liver hardness value was used as the clinical assessment index,and the comparison between the two groups showed that the Chinese herb group had a better efficacy in improving the liver hardness value,with statistically significant differences[MD=3.52,95%CI(4.78,2.26),P<0.00001].The portal vein diameter was used as the clinical assessment index in 33 literature studies,and the comparison between the two groups showed that the traditional Chinese herb group had better efficacy in improving the portal vein diameter,with statistical difference[MD=1.20,95%CI(2.14,0.26),P=0.01].In 31 literature studies,spleen thickness was used as the clinical assessment index,and the comparison between the two groups showed that the TCM group had better efficacy in improving spleen thickness,with statistical difference[MD=6.83,95%CI(9.54,4.12),P<0.00001].13 literature studies took the diameter of splenic vein as the clinical assessment index,and the comparison between the two groups showed that the Chinese herb group had a better efficacy in improving the diameter of splenic vein,with statistically significant differences[MD=1.77,95%CI(3.04,0.49),P=0.007].Conclusion:Compared with western medicines,Chinese herbal prescriptions with the main effect of YiQi and HuoXue show better clinical efficacy in the treatment of liver fibrosis.2.To study the anti-fibrosis mechanisms of Qizhu granules based on the microscale biomechanical properties of hepatic sinusoidal endothelial cellsObjectives:(1)To study the regulatory effects of Qizhu granules on liver inflammatory factors,liver fibrosis indexes and pathological histology in liver fibrosis.(2)The effects of Qizhu granules on the eNOS and NO in hepatic sinusoidal endothelial cells were determined by qRT-PCR Western blot and immunofluorescence.(3)Based on the theory of multiphase and multilevel porous media,the mechanical characteristics of hepatic sinusoid endothelial cells under the intervention of Qizhu granules were determined.Methods:(1)Carbon tetrachloride was used as the inducer of liver fibrosis to construct the rat model of liver fibrosis.Mealwhile,Qizhu granules were given for intervention.The effects of Qizhu granules on the biochemical and hipathology of liver fibrosis rats were studied by ELISA,HE and Masson staining.(2)In situ collagenase perfusion+in vitro digestion+gradient density separation were used to extract hepatic sinusoidal endothelial cells.(3)10%hepatic fibrosis rat serum+5%fetal bovine serum was used as the external injury factor to treat normal sinusoid endothelial cells cultured in vitro,and the injury model of hepatic sinusoid endothelial cells was established by simulating the biochemical environment in vivo.The cells were divided into groups according to normal control group,injury group,normal rat serum injury group,low,medium and high concentration of drug-containing serum of Qizhu granules.(4)The expressions of eNOS and NO in cells were detected by qRT-PCR,Western blot and immunofluorescence.(5)Based on the theory of multiphase and multilevel porous media,atomic force microscopy was used to characterize the mechanical properties of hepatic sinusoidal endothelial cells under the intervention of Qizhu granules.Results:(1)Qizhu granules can improve the biochemical indexes and pathological morphology of liver in rats with liver fibrosis.(2)According to cell morphology and immunofluorescence identification,the extracted cells were rat hepatic sinusoid endothelial cells with high purity.(3)Detection of eNOSmRNA?eNOS and NO by qRT-PCR,Western blot and immunofluorescence showed that Qizhu granules containing serum could up-regulate the expression of eNOS and NO.(4)After the intervention of Qizhu granules containing drug serum,the cytoskeletal modulus and viscosity of hepatic sinusoidal endothelial cells were increased,but the diffusion coefficient did not change significantly.Conclusions:(1)Qizhu granules can inhibit liver inflammation and restore normal liver tissue morphology.(2)Qizhu granules can up-regulate the expression of eNOSmRNA?eNOS and NO,thus improving microcirculation,which is one of the important mechanisms of Qizhu granules against liver fibrosis.(3)Qizhu granules can improve the mechanical properties of sinusoidal endothelial cells and maintain the normal mechanical properties of sinusoidal endothelial cells,which may be another important mechanism of its anti-fibrosis.