基质金属蛋白酶抑制剂1和2 siRNA对CCl4诱导的肝纤维化大鼠Smad7蛋白表达的影响

Effect of tissue inhibitor of metalloproteinases 1 and 2 siRNA on the expression of Smad7 protein in hepatic fibrosis rats

摘要:

目的：研究基质金属蛋白酶抑制剂（tissue inhibitor of metalloproteinases，TIMP）1和2的siRNA对CCl4诱导的肝纤维化大鼠Smad7蛋白表达的影响。方法：采用前期构建的TIMP-1 siRNA、TIMP-2 siRNA治疗肝纤维化大鼠的肝脏组织作为研究对象，分为正常组、模型组、阴性对照组（0.25 mg/kg非特异性靶序列）、TIMP-1 siRNA 0. 25 mg/kg组和TIMP-2 siRNA 0.25 mg/kg组。Real-time PCR法检测Smad7的mRNA表达；免疫组织化学、Western blot检测Smad7的蛋白表达。结果：Real-time PCR结果显示TIMP-1 siRNA 0.25 mg/kg组（59.7±3.3）%、TIMP-2 siRNA 0.25 mg/kg组（60.2±3.4）% Smad7的mRNA表达量均较模型组（28.0±1.6）%、阴性对照组（28.6±1.7）%明显增加（P=0.000）；免疫组织化学结果显示TIMP-1 siRNA 0.25mg/kg组（8.55±0.67）、TIMP-2 siRNA 0.25 mg/kg组（8.23±0.85）Smad7的蛋白表达量均较模型组（4.25±0.53）、阴性对照组（4.19±0.55）明显增加（P=0.000）。Western blot结果显示TIMP-1 siRNA 0.25 mg/kg组（0.706±0.039）、TIMP-2 siRNA 0.25 mg/kg组（0.698±0.038）Smad7的蛋白表达量均较模型组（0.278±0.013）、阴性对照组（0.285±0.014）明显增加（P=0.000）。结论：TIMP-1 siRNA、TIMP-2 siRNA对于肝纤维化Smad7的表达有促进作用。

Abstract:

Objective：To observe the expression of Smad7 protein in carbon tetrachloride（CCl4）-induced hepatic fibrosis rats treated with TIMP-1 short interference RNA（siRNA） and TIMP-2 siRNA. Methods：The hepatic tissues collected from hepatic fibrosis rats were divided into 5 groups：normal group，model group，negative control group，TIMP-1 siRNA 0.250 mg/kg group and TIMP-2 siRNA 0.250 mg/kg group. The expression of Smad7 was detected by real-time PCR，immunohistochemistry and Western blot. Results：Real-time PCR analysis showed that mRNA expression of Smad7 was significantly higher in TIMP-1 siRNA 0.250 mg/kg group（59.7±3.3）% and TIMP-2 siRNA 0.250 mg/kg group（60.2±3.4）% than in model group（28.0±1.6）% and negative control group（28.6±1.7）%（P=0.000）. Immunohistochemistry showed that protein expression of Smad7 was significantly higher in TIMP-1 siRNA 0.250 mg/kg group（8.55±0.67） and TIMP-2 siRNA 0.250 mg/kg group（8.23±0.85） than in model group（4.25±0.53） and negative control group（4.19±0.55）（P=0.000）. Western blot showed that protein expression of Smad7 was significantly higher in TIMP-1 siRNA 0.250 mg/kg group（0.706±0.039） and TIMP-2 siRNA 0.250 mg/kg group（0.698±0.038） than in the model group（0.278±0.013） and negative control group（0.285±0.014）（P=0.000）. Conclusion：Silencing the TIMP-1 and TIMP-2 with siRNA could be involved in the improvement of the hepatic fibrosis in rats.