兔慢性肝损害急性肝衰竭动物模型的建立

Objective To establish an ideal animal model of acute-on-chronic liver failure (ACLF) in New Zealand white rabbits in order to provide a large animal model for further researches.Methods Totally 75 New Zealand rabbits were randomly divided into experimental group (n =70) and control group (n = 5 ). Rabbits in the experimental group were injected with CCl4 into the abdominal cavity twice every week and the doses of CCl4 were modified according to the index of liver function and the body weight, whereas those in the control group were treated with the same volume of saline. At the 10th week,48 New Zealand rabbits with hepatic fibrosis were randomly assigned to 4 groups and injected with CCl4 as before, D-Gal at a dose of 0. 65 g/kg body weight (BW), 0. 70 g/kg BW and 0. 75 g/kg BW, respectively. By observing and comparing the general state, survival time, biochemical indexes, and the histopathology, a method of establishing a stable animal model of acute hepatic failure was found. Results As compared with those in control group, the levels of ALT, AST, GGT, HA, LN and PC-Ⅲ in the experiment group were increased significantly, while the level of ALB was decreased at the end of 10 weeks. Typical features of hepatic fibrosis and the formation of pseudo-lobules were observed at the end of 10 weeks. After treatment with D-Gal, all rabbits in group Ⅰ survived with minimal changes in liver function tests. In group Ⅱ , there was a temporary hepatic injury, but no hepatic coma. Four of the 12 rabbits died (33. 3% ). In group Ⅲ , biochemical indexes changed obviously 12 h after the administration and hepatic injury reached its peak after 48 h. Ten of 12 rabbits were died of severe hepatic failure with a survival time of ( 53. 00 ± 25. 69) h. Histology of liver section revealed massive necrosis in nodules. In group Ⅳ , hepatic injury occurred early and severely. All the rabbits died of severe hepatic failure with a survival time of (32. 70 ± 17. 46) h. Conclusion The experimental model of ACLF could be established by injected with D-Gal in New Zealand rabbits with hepatic fibrosis, induced by CCl4 intraperitoneal injection for 10 weeks.The one induced by 0. 70 g/kg of D-galactosamine was more stable and showed similar clinical pathophysiological changes in human beings. So it can be a good experimental platform for studies of ACLF.     

兔慢性肝损害急性肝衰竭动物模型的建立

目的 建立新西兰兔的慢加急性肝衰竭(ACLF)动物模型.方法 将75只新西兰兔随机分为实验组(n=70)与对照组(n=5),实验组采用四氯化碳(CCl4)腹腔注射建立兔代偿性肝纤维化模型,每周2次,通过体质量及谷丙转氨酶(ALT)/谷草转氨酶(AST)改变调整药物剂量,共10周,获得48只肝纤维化新西兰兔,在此基础上将动物随机分为4组(n=12),Ⅰ组:继续腹腔注射CCl4,Ⅱ组:静脉注射D-氨基半乳糖(D-Gal)0.65 g/kg,Ⅲ组:静脉注射D-Gal 0.70 g/kg,Ⅳ组:静脉注射D-氨基半乳糖0.75 g/kg,观察各组动物的一般情况、生化指标及病理改变.结果 与对照组比较,肝纤维化实验组兔10周时ALT、AST、ALB、γ-谷氨酰转肽酶(GGT)、透明质酸(HA)、层粘连蛋白(LN)、Ⅲ型前胶原(PC-Ⅲ)差异均有统计学意义(P＜0.05),可观察到肝纤维化的病理表现,出现典型的假小叶,在肝纤维化基础给予D-Gal后,Ⅰ组动物全部存活,生化指标轻度改变;Ⅱ组动物死亡率为33.3%(4/12),生化指标仅出现一过性的改变;Ⅲ组动物死亡率为83.3%(10/12),平均存活时间为(53.00±25.69)h,给药后12 h生化指标及临床表现出现改变,48 h达到高峰,病理显示肝脏大块坏死;Ⅳ组动物均死于肝衰竭,平均存活时间为(32.70±17.46)h,肝损害出现时间早,损伤剧烈.结论 对CCl4诱导的肝纤维化新西兰兔给予D-Gal急性攻击可建立ACLF模型.其中给予D-Gal 0.70 g/kg的模型稳定性好,能较大程度上的模拟临床上ACLF的病理生理过程.

Abstract：

Objective To establish an ideal animal model of acute-on-chronic liver failure (ACLF) in New Zealand white rabbits in order to provide a large animal model for further researches.Methods Totally 75 New Zealand rabbits were randomly divided into experimental group (n =70) and control group (n = 5 ). Rabbits in the experimental group were injected with CCl4 into the abdominal cavity twice every week and the doses of CCl4 were modified according to the index of liver function and the body weight, whereas those in the control group were treated with the same volume of saline. At the 10th week,48 New Zealand rabbits with hepatic fibrosis were randomly assigned to 4 groups and injected with CCl4 as before, D-Gal at a dose of 0. 65 g/kg body weight (BW), 0. 70 g/kg BW and 0. 75 g/kg BW, respectively. By observing and comparing the general state, survival time, biochemical indexes, and the histopathology, a method of establishing a stable animal model of acute hepatic failure was found. Results As compared with those in control group, the levels of ALT, AST, GGT, HA, LN and PC-Ⅲ in the experiment group were increased significantly, while the level of ALB was decreased at the end of 10 weeks. Typical features of hepatic fibrosis and the formation of pseudo-lobules were observed at the end of 10 weeks. After treatment with D-Gal, all rabbits in group Ⅰ survived with minimal changes in liver function tests. In group Ⅱ , there was a temporary hepatic injury, but no hepatic coma. Four of the 12 rabbits died (33. 3% ). In group Ⅲ , biochemical indexes changed obviously 12 h after the administration and hepatic injury reached its peak after 48 h. Ten of 12 rabbits were died of severe hepatic failure with a survival time of ( 53. 00 ± 25. 69) h. Histology of liver section revealed massive necrosis in nodules. In group Ⅳ , hepatic injury occurred early and severely. All the rabbits died of severe hepatic failure with a survival time of (32. 70 ± 17. 46) h. Conclusion The experimental model of ACLF could be established by injected with D-Gal in New Zealand rabbits with hepatic fibrosis, induced by CCl4 intraperitoneal injection for 10 weeks.The one induced by 0. 70 g/kg of D-galactosamine was more stable and showed similar clinical pathophysiological changes in human beings. So it can be a good experimental platform for studies of ACLF.     

脾切除对肝纤维化大鼠肝脏PDGF-B表达及血清PDGF-BB水平的影响

Objective To explore the effects of splenectomy on hepatic fibrosis and on the expression of PDGF-B in the liver and PDGF-BB in the serum of rats with hepatic fibrosis. Methods By hypodermic injection CCl4, we established 65 rat models with hepatic fibrosis, splenectomies were performed in the three groups at different phases: before hypodermic injection CCl4 (A group), five weeks after hypodermic injection CCl4 (B group), and ten weeks hypodermic injection CCl4 (C group). The control groups were established at the same time, with samples of the livers and serum of the rats taken in different phases. The expressions of PDGF in the liver were detected by immunohistochemistry technique and the degree of hepatic fibrosis was detected by HE staining. The serum levels of PDGF-BB were analyzed by ELISA technique. Results Absorbance values of PDGF-B in the experimental group were significantly lower than the control groups (P＜0. 05). Serum levels of PDGF-BB of the rats after splenectomy were significantly lower than those in the control groups (P＜0.05). HE and Masson's staining showed that the progression of Hepatic fibrosis was slow in the A group. Hepatic pathologic state was significantly relieved in the B group and the inflammation and fibrosis was relieved in the C group. Conclusion Earlier period splenectomy could delay the proceeding of experimental hepatic fibrosis. After splenectomy the decline in the level of PDGF may be one of the mechanisms causing the delay.     

自体骨髓移植治疗股骨头无菌性坏死

Objective To investigate the effectiveness and mechanism of transplantation of autol-ogous bone marrow in treating femoral head necrosis. Methods Sixty New Zealand rabbits were randomly divided into groups A,B,and C after establishment of the models of femoral head necrosis. The left femoral head served as the control one,and the right as the experimental group. The mitoxantrone (0.1 mg/kg) with the help of DSA was injected into the femoral head of group A. One ml autologous bone marrow was injected into the femoral head of group B. In group C,mitoxantrone (0. 1 mg/kg) was injected,and 72 h later,1 ml autologous bone marrow was injected. Four months later,all rabbits were killed,and the femoral heads were removed and observed histologically and electron microscopically. Results The number of necrotic osteoclasts in groups A and B showed no significant difference ( P > 0.05 ), ande in group C, the number of necrotic femoral heads at the left and fight sides was 40. 60±4.11 and 21.33±2.16 respec-tively ( P < 0.05 ). At the experimental side of group C, the structure of majority bone cells was clear and intact,and necrosis was occasionally seen. Conclusion At the cellular level, local chemotherapy and au-tologous hematopoietic stem cell transplantation had certain effectiveness for aseptic necrosis of the femoral head.     

自体骨髓移植治疗股骨头无菌性坏死

Objective To investigate the effectiveness and mechanism of transplantation of autol-ogous bone marrow in treating femoral head necrosis. Methods Sixty New Zealand rabbits were randomly divided into groups A,B,and C after establishment of the models of femoral head necrosis. The left femoral head served as the control one,and the right as the experimental group. The mitoxantrone (0.1 mg/kg) with the help of DSA was injected into the femoral head of group A. One ml autologous bone marrow was injected into the femoral head of group B. In group C,mitoxantrone (0. 1 mg/kg) was injected,and 72 h later,1 ml autologous bone marrow was injected. Four months later,all rabbits were killed,and the femoral heads were removed and observed histologically and electron microscopically. Results The number of necrotic osteoclasts in groups A and B showed no significant difference ( P > 0.05 ), ande in group C, the number of necrotic femoral heads at the left and fight sides was 40. 60±4.11 and 21.33±2.16 respec-tively ( P < 0.05 ). At the experimental side of group C, the structure of majority bone cells was clear and intact,and necrosis was occasionally seen. Conclusion At the cellular level, local chemotherapy and au-tologous hematopoietic stem cell transplantation had certain effectiveness for aseptic necrosis of the femoral head.     

自体骨髓移植治疗股骨头无菌性坏死

Objective To investigate the effectiveness and mechanism of transplantation of autol-ogous bone marrow in treating femoral head necrosis. Methods Sixty New Zealand rabbits were randomly divided into groups A,B,and C after establishment of the models of femoral head necrosis. The left femoral head served as the control one,and the right as the experimental group. The mitoxantrone (0.1 mg/kg) with the help of DSA was injected into the femoral head of group A. One ml autologous bone marrow was injected into the femoral head of group B. In group C,mitoxantrone (0. 1 mg/kg) was injected,and 72 h later,1 ml autologous bone marrow was injected. Four months later,all rabbits were killed,and the femoral heads were removed and observed histologically and electron microscopically. Results The number of necrotic osteoclasts in groups A and B showed no significant difference ( P > 0.05 ), ande in group C, the number of necrotic femoral heads at the left and fight sides was 40. 60±4.11 and 21.33±2.16 respec-tively ( P < 0.05 ). At the experimental side of group C, the structure of majority bone cells was clear and intact,and necrosis was occasionally seen. Conclusion At the cellular level, local chemotherapy and au-tologous hematopoietic stem cell transplantation had certain effectiveness for aseptic necrosis of the femoral head.     

自体骨髓移植治疗股骨头无菌性坏死

Objective To investigate the effectiveness and mechanism of transplantation of autol-ogous bone marrow in treating femoral head necrosis. Methods Sixty New Zealand rabbits were randomly divided into groups A,B,and C after establishment of the models of femoral head necrosis. The left femoral head served as the control one,and the right as the experimental group. The mitoxantrone (0.1 mg/kg) with the help of DSA was injected into the femoral head of group A. One ml autologous bone marrow was injected into the femoral head of group B. In group C,mitoxantrone (0. 1 mg/kg) was injected,and 72 h later,1 ml autologous bone marrow was injected. Four months later,all rabbits were killed,and the femoral heads were removed and observed histologically and electron microscopically. Results The number of necrotic osteoclasts in groups A and B showed no significant difference ( P > 0.05 ), ande in group C, the number of necrotic femoral heads at the left and fight sides was 40. 60±4.11 and 21.33±2.16 respec-tively ( P < 0.05 ). At the experimental side of group C, the structure of majority bone cells was clear and intact,and necrosis was occasionally seen. Conclusion At the cellular level, local chemotherapy and au-tologous hematopoietic stem cell transplantation had certain effectiveness for aseptic necrosis of the femoral head.     

自体骨髓移植治疗股骨头无菌性坏死

Objective To investigate the effectiveness and mechanism of transplantation of autol-ogous bone marrow in treating femoral head necrosis. Methods Sixty New Zealand rabbits were randomly divided into groups A,B,and C after establishment of the models of femoral head necrosis. The left femoral head served as the control one,and the right as the experimental group. The mitoxantrone (0.1 mg/kg) with the help of DSA was injected into the femoral head of group A. One ml autologous bone marrow was injected into the femoral head of group B. In group C,mitoxantrone (0. 1 mg/kg) was injected,and 72 h later,1 ml autologous bone marrow was injected. Four months later,all rabbits were killed,and the femoral heads were removed and observed histologically and electron microscopically. Results The number of necrotic osteoclasts in groups A and B showed no significant difference ( P > 0.05 ), ande in group C, the number of necrotic femoral heads at the left and fight sides was 40. 60±4.11 and 21.33±2.16 respec-tively ( P < 0.05 ). At the experimental side of group C, the structure of majority bone cells was clear and intact,and necrosis was occasionally seen. Conclusion At the cellular level, local chemotherapy and au-tologous hematopoietic stem cell transplantation had certain effectiveness for aseptic necrosis of the femoral head.     

自体骨髓移植治疗股骨头无菌性坏死

Objective To investigate the effectiveness and mechanism of transplantation of autol-ogous bone marrow in treating femoral head necrosis. Methods Sixty New Zealand rabbits were randomly divided into groups A,B,and C after establishment of the models of femoral head necrosis. The left femoral head served as the control one,and the right as the experimental group. The mitoxantrone (0.1 mg/kg) with the help of DSA was injected into the femoral head of group A. One ml autologous bone marrow was injected into the femoral head of group B. In group C,mitoxantrone (0. 1 mg/kg) was injected,and 72 h later,1 ml autologous bone marrow was injected. Four months later,all rabbits were killed,and the femoral heads were removed and observed histologically and electron microscopically. Results The number of necrotic osteoclasts in groups A and B showed no significant difference ( P > 0.05 ), ande in group C, the number of necrotic femoral heads at the left and fight sides was 40. 60±4.11 and 21.33±2.16 respec-tively ( P < 0.05 ). At the experimental side of group C, the structure of majority bone cells was clear and intact,and necrosis was occasionally seen. Conclusion At the cellular level, local chemotherapy and au-tologous hematopoietic stem cell transplantation had certain effectiveness for aseptic necrosis of the femoral head.