The Effects Of Astragalus On Liver Fibrosis Induced By Partial Bile Duct Obstruction

Objective:To establish a new model of hepatic fibrosis induced by partial bile duct obstruction in rats.Methods:(1) Thirty-six SD rats were randomly divided into6groups, normal control (N), sham (S), partial bile duct obstruction (PBDO) groups. And then PBDO group were further divided into four groups, PBDO1, PBDO2, PBDO3and PBDO4, according to the duration after PBDO.(2)PBDO rats, anesthetized with10%chloral hydrate (300mg/kg) by intraperitoneal injection, were made a longitudinal incision along abdominal midline. The choledoch was found and freed from inferior vena cava, then were ligated with one infusion needle of0.55mm diameter. Diameter of choledoch was narrowed to0.55mm by the coil after drawing out the needle. N group did nothing. The choledoch of S group was only freed without ligation. Every animal was raised in one cage under the same environment after surgery.(3) We collected blood (4～5ml) from each rat at first, second, third and fourth week after PBDO and then centrifuged them for10minutes on3000r/min. The serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), direct bilirubin (DBIL), alkaline phosphatase (ALP) And serum total protein (TP) were obseved by automatic biochemical analyzer. The serum levels of hyaluronic acid (HA), laminin (LN), III procollagen peptide (PⅢ P), IV collagen (CIV) were detected by time-resolved fluorescence immunoassay method.(4) Right lobe tissues of liver were fixed by10%formalin and then paraffin-embedded after gradient dehydration. The hepatic fibrosis was assessed by H&E and Van Gieson staining. The pathological changes in liver tissue and fibrous proliferation were graded according to "viral hepatitis prevention and treatment programs" published in2000.Result:(1)Rats of PBDO group had less eating,activity and drinking than N group, but2days later increased. Some rats’ears appeared bile-stained24hours later and most of rats’ears appeared yellow during3days after PBDO. The jaundice increased gradually and lasted for about2days then disappeared within7days.(2) The proximal diameter of common bile duct in rats of PBDO1group dilated from6to10mm1week later, filled with yellow-green bile, and distal segment of the ligated bile duct appeared yellow. The chyme in duodenum looks golden brown.(3) There was no statistically significant difference among all groups of ALT、AST、TBIL and DBIL. TP level of PBDO group rats decreased significantly at first week after PBDO. HA level of PBDO2group increased compared with N group, while LN, PⅢP and CIV levels increased significantly at3rd or4th week when compared with groups N, S and there was statistically significant difference (P<0.05).(4) All slides of liver tissues were observed under microscopy after HE-stain and VG-stain. Liver capsule is thicker at first week after PBDO with proliferation of cholangiole epithelial cell.Edema and degeneration of liver cells were also seen as focal necrosis of liver cells.In some areas, neutrophils and lymphocyte infiltration can be seen and the number of bile capillary increased with marked hyperplasia of biliary epithelial cells.The scope of portal area became wider. Fibrous tissue increased around vascellum of portal area and biliary ducts. Severe vacuolar degeneration of liver cells at3rd and4th week and more collagen fiber proliferation were also seen. VG-stains of liver tissue showed proliferation of cholangiole epithelial cells, wider portal area and fibrous tissue hyperplasia.It was S1or S2stage in pathology. The architecture of hepatic lobules was normal at first week after PBDO. The number of cholangiole was significantly increased with more small veins and collagen fibers. Fibrous tissues extended along the edge of the hepatic lobule, showing S1to S3stages in pathology. At3th and4th weeks, the collagen fibers proliferated significantly surrounding the hepatic lobule and fibrous septa made lobular structural disorder, but no cirrhosis, showing S3stage in pathology.Conclusion By using choledoch partial obstruction, liver fibrosis induced by partial biliary obstruction was successfully estabolished. Objective:To observe the effects of astragalus on liver fibrosis and MMP-2, TIMP-2alternations.Method:Forty-two SD rats were randomly divided into7groups, normal control (N), sham (S), astragalus (A), partial bile duct obstruction (PBDO) groups.PBDO groups were further divided into four groups, PBDO1, PBDO2, PBDO3,and PBD04according to the duration after PBDO.Liver fibrosis model induced by partial bile duct obstruction in rats were established. Rats of group A were given astragalus(800mg/kg) by intraperitoneal injection from third week to fourth week. Liver function,serum level of CIV, HA, LN, PIIIP were detected. HE and VG staining sections were assessed for liver fibrosis. Immunohistochemistry was used to detect MMP-2, TIMP-2expression in liver tissues.Results:The liver fibrosis grade of group A is lower than that of other PBDO groups in liver function when compared with other groups. There was significant difference in Serum level of CⅣ, HA, LN, PIIIP compared with PBDO4group, P<0.05. MMP-2and TIMP-2expression in PBDO groups liver tissue gradually enhancement with duration after PBDO. A group level of TIMP-2is lower compared with PBDO4group (P<0.05), but there is no significant difference about MMP-2.Conclusion:The experiment showed that astragalus might inhibit liver fibrosis progression due to bile duct obstruction by inhibiting TIMP-2but enhancing the function of MMP-2.