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目的探讨基质金属蛋白酶抑制剂1(TIMP-1)在大鼠酒精性肝病发生发展中的动态变化及表达。方法应用灌胃的方法制备大鼠酒精性肝病的动物模型,肝组织作HE染色及Masson三色胶原染色,Masson染色结果用病理图象分析仪分析,测定胶原纤维面积百分比。应用免疫组织化学技术检测TIMP-1在酒精性肝病中的表达,其结果用病理图象分析仪分析。应用流式细胞术检测酒精性肝病不同阶段TIMP-1 FI值的动态变化。结果肝脏组织病理学改变,对照组HE染色可见正常中央静脉及放射状排列的肝板。模型组4wk末肝细胞轻度脂胁变性及点状坏死。8wk末肝细胞中重度脂肪变性,炎性细胞浸润加重,Masson染色终末静脉周围胶原纤维增因加。12wk末及16wk末肝细胞坏死增多,炎性细胞浸润成团,Masson染色终末静脉增厚,胶原纤维向窦周隙延伸,窦周纤维组织较前增加,造模成功。免疫组化研究表明,TIMP-1主要位于血管内皮细胞、窦内皮细胞、肝窦细胞及静脉周围肝细胞的胞质。病理图象分析结果示对照组4,8,12,16wk TIMP-1面积比分别为4.8±0.7,4.8±0.6,5.2±0.8,5.1±0.7,模型组四个阶段分别为9.8±1.4,16.7±2.4,21.3±3.4,25.3±3.7,明显高于对照组(P<0.05)。流式细胞术检测示对照组TIMP-1 FI值四个阶段分别为0.4±0.1,0.5±0.2,0.4±0.1,0.5±0.1,模型组分别为0.6±0.1,0.8±0.1,1.0±0.1,1.2±0.2,明显高于对照组(P<0.05),并且随造模时间延长其含量进行性增加。结论在大鼠酒精性肝病发生发展中,TIMP-1水平进行性增加,提示其可能参与酒精性肝病的进展。
Objective To investigate the dynamic changes and expression of TIMP-1 in the development of alcoholic liver disease in rats. Methods The rat model of alcoholic liver disease was prepared by intragastric administration. The liver tissue was stained with HE and Masson trichrome staining. The masson staining was analyzed by pathological image analyzer to determine the percentage of collagen fiber area. The expression of TIMP-1 in alcoholic liver disease was detected by immunohistochemistry. The results were analyzed by pathology image analyzer. Flow cytometry was used to detect the dynamic change of TIMP-1 FI in different stages of alcoholic liver disease. Results The histopathological changes of liver tissue showed normal central veins and radial arrangement of liver plate in control group. At the end of 4wk, the model group was mildly lipidated and necrosis of hepatocytes. Severe steatosis, inflammatory cell infiltration aggravated in the end of 8wk liver cells, and the increase of collagen fibers around the terminal vein of Masson staining. At the end of 12wk and the end of 16wk, necrosis of hepatocytes increased, inflammatory cells infiltrated into the mass, Masson-stained terminal venous thickened, collagen fibers extended to the peroneal space and the fibrous tissue around the sinus increased earlier, and the model was successful. Immunohistochemical studies showed that TIMP-1 mainly located in the cytoplasm of vascular endothelial cells, sinus endothelial cells, hepatic sinusoidal cells and perivenous hepatocytes. Pathological image analysis showed that the area ratio of 4, 8, 12, and 16wk TIMP-1 in the control group were 4.8 ± 0.7, 4.8 ± 0.6, 5.2 ± 0.8 and 5.1 ± 0.7, respectively. The four stages in the model group were 9.8 ± 1.4 and 16.7 ± 2.4,21.3 ± 3.4,25.3 ± 3.7, significantly higher than that of the control group (P <0.05). Flow cytometry showed that the TIMP-1 FI value in the control group was 0.4 ± 0.1, 0.5 ± 0.2, 0.4 ± 0.1 and 0.5 ± 0.1 in four stages and 0.6 ± 0.1, 0.8 ± 0.1 and 1.0 ± 0.1 in the model group, 1.2 ± 0.2, which was significantly higher than that of the control group (P <0.05), and its content increased progressively with the time of modeling. Conclusions In the development of alcoholic liver disease in rats, the progressive increase of TIMP-1 level suggests that it may be involved in the progression of alcoholic liver disease.