动物造模,肝纤维化模型 z-bio 2021-12-09 1473

　　(1) Replication method Adult rats, open the abdominal cavity after anesthesia, find and release the common bile duct in the duodenal descending mesentery, pass two silk threads under it, and cut a "V"-shaped incision into the liver. Use 1 ml of tissue adhesive (TH adhesive) or 0.3 ml/kg body weight of N-butyl-2-cyanoacrylate into the proximal end of the syringe, ligate twice and cut the common bile duct. Store it regularly for 8 weeks after surgery. During this period, observe the general activities and physical signs of the animals, and dynamically collect blood from the orbit to alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), And prepare the conjugate serum. Bilirubin (DBIL). ), alkaline phosphatase (ALP), γ-glutamyl transpeptidase (GGT), hyaluronic acid (HA) and laminin (LN) content detection, complete the model, harvest the liver to prepare hydroxyproline homogenate Hyp content and sampling. Morphological examination of liver tissue was performed according to the requirements of optical microscope and electron microscope.

　　(2) Characteristics of the model One week after the operation, the model animal developed jaundice. The levels of serum ALT, AST, TBIL, DBIL, ALP, and GGT increased at 2, 4, and 8 weeks, and their levels gradually increased with the increase of serum HA and LN and the increase of the biliary atresia time of liver tissue Hyp. Observation under an optical microscope revealed that the structure of the liver lobules was destroyed 1 to 3 weeks after the modeling, and the liver cells were of different sizes, some of them were balloon-shaped, and there was obvious inflammatory cell infiltration in the portal vein area and the bile duct. I will expand, cholestasis, proliferation and biliary epithelial cell proliferation, a large number of fibroblasts appear, extending to the lobules and forming fibrous cords. Most of the phylum area has a large amount of type I and type III collagen growth. In 8 weeks, the proliferative bile ducts occupies most of the liver lobules, and the bile ducts with the remaining small hepatocytes are divided into islands, a large amount of collagen is deposited along the proliferated bile ducts, and the fibrous interval is thinner. Electron microscope observation revealed that most of the hepatocytes in the model group had irregular nuclei, wrinkles or ruptured nuclear membranes, some nucleoli disappeared, and the organelles in the cytoplasm were significantly reduced and changed. The rupture products of the organelles were visible, some of which showed roughness, and the organelles showed Cystic swelling occurs, a lot of ribosomes are lost, and some mitochondrial swelling disappears. The cell membrane is incompletely ruptured; the villi on the surface of the hepatocyte sinusoids are shed, the connections between the cells are loosened, and the gaps are enlarged; the bile duct cells proliferate, the cavity is enlarged, and the microvilli of the lumen is deformed, edema or missing; a large number of collagen fiber ducts and molds are visible around the bile Appear in the cavity, exposing the capillaries around the sinuses.

　　(3) Comparative medicine Extrahepatic bile duct obstruction can cause the obstructed area above the bile duct to expand, cholestasis, increase the pressure of the bile duct, and may cause the expansion and rupture of the intrahepatic bile duct. When blood vessels in the liver are compressed by dilated bile ducts and bile extravasation, liver cells undergo ischemia and necrosis. The fibrous tissue extends to the bile duct, surrounds the liver lobules, spreads around the liver cells, and causes fibrosis, hyperplasia, and even bile duct development. Liver fiber. The animal model of liver fibrosis induced by retrograde injection of tissue adhesive (TH adhesive) and common bile duct ligation has the unique characteristics of biliary liver fibrosis. The main characteristics of the model: the degree of inflammation in the model animals is small, the formation of liver fibrosis is faster, and the spontaneous reversal rate is low. Specifically, the necrosis of hepatocytes after the modeling operation is unclear. The serum ALT level, which reflects the degree of liver cell damage, only increased rapidly and rapidly on the first day after surgery, but then decreased rapidly. One to two weeks after modeling, the bile ducts near the entrance area of the liver tissue were hyperplastic with fibrous tissue deposits around them; two weeks later, the proliferative fibrous tissue around the bile ducts surrounded the liver lobules and began to separate, which also reflected the degree of liver fibrosis . The content of LN and HA in serum and tissues can be increased or maintained at a higher level, and the content of Hpy in liver tissue can rise to three times the normal level within 4 weeks. In 4-8 weeks of modeling, compared with the hyperplastic bile duct, a large amount of collagen is deposited around the diffuse hyperplastic bile duct, and the hepatocytes are significantly reduced. The indicators TBIL, DBIL, ALP and GGT reflecting cholestasis are still high. The LN and HA indicators of serum tissue are still very high. After 16 weeks, the liver tissue still showed obvious fibrosis with thickened fibrous septum. Compared with animal models of liver fibrosis caused by simple bile duct ligation or bile duct ligation and resection, the latter two model animals are more prone to bile duct recanalization, which leads to longer bile duct atresia, which is very difficult. The characteristics of liver fibrosis can be reduced or disappeared in a short period of time. The reason why the model can maintain the liver fibrosis state of the model animal for a long time is that the injection of TH adhesive or N-butyl N-2-cyanoacrylate into the bile duct during the modeling operation is an important technique. This method technically guarantees the complete occlusion of the bile duct, especially the intrahepatic bile duct. Considering the simple modeling method of the model, shorter modeling period, longer maintenance time, obvious pathological characteristics and dynamic use of non-invasive serological indicators, many scholars have chosen liver fiber analysis. This model is more suitable for studying the formation mechanism of liver fibrosis and evaluating the efficacy of drugs in treating fibrosis.