BMP-7/Smad6信号通路在酒精性肝损伤大鼠模型中的调节作用

Function of BMP-7/Smad6 signaling pathway in the rat model of alcoholic liver disease

目的探讨BMP-7/Smad6信号传导通路在酒精性肝病发病机制中的作用并检测其在酒精性肝病大鼠模型中的基因水平和蛋白水平。方法以自由摄取白酒加高脂饲料诱导酒精性肝纤维化大鼠模型,并比较普通饲料与高脂饲料对酒精性肝病的影响。实验结束后取各实验组肝组织观察病理学变化,用生化检测仪检测大鼠血浆ALT、AST变化。用实时RCR检测肝的骨形成蛋白7(BMP-7)和Smad6 mRNA水平变化,并用Western Blot检测蛋白水平变化。结果肝脏组织形态学改变显示正常饲料组肝组织出现脂肪肝表现,高脂饲料组脂肪变性程度加重并出现了肝炎及胶原纤维增生,而对照组大鼠肝脏组织结构正常。生化分析仪检测ALT、AST结果显示:至实验结束时,实验组比对照组转氨酶活性明显升高(P<0.05)。BMP和Smad6这两个基因的蛋白丰度与它们各自的mRNA表达水平呈正相关,与对照组比较两个实验组BMP-7表达水平降低,并与肝脏病理损伤程度呈负相关,两个实验组Smad6表达水平较对照组升高,但与肝脏病理变化严重程度无相关性。结论饮用白酒水溶液结合高脂饲料的方法可以成功建立酒精性肝损伤的大鼠模型,BMP-7/Smad6信号通路的失活促进了肝纤维化的形成。

Objective To investigate the function of BMP-7/Smad6 signaling pathway in alcoholic liver disease rat model and detect the two genes in both mRNA and protein levels. Methods Rats were fed with alcohol and high fat diet to establish alcoholic liver disease model. In the end of the experiment,liver of rats were dissected for histological studies whereas plasma obtained from rats was taken for biochemical testing to detect the levels of alanine aminotransferase（ ALT） and aspartate aminotransferase（ AST）. mRNA was extracted from the liver to perform real-time PCR in order to detect the mRNA level of bone morphogenetic proteins 7（ BMP-7） and Smad6. Western blot was performed to determine the protein level of BMP-7 and Smad6. Results Livers from the experimental group fed with alcohol demonstrated histopathological changes such as fatty change,inflammation as well as fibrosis; however,no pathological changes were observed in the livers of rats in normal control group. Biochemical testing results showed that levels of AST and ALT were higher in experimental group compared to normal control group. The results of PCR and western blot showed that the mRNA levels of BMP-7 and Smad6 positively correlated to the corresponding protein levels. BMP-7 level of normal control group was higher than that of experimental group,and the level was negatively correlated to the severity of hepatic pathological changes of liver. Smad6 levels were higher in experimental group compared to the normal control group,however,there was no correlation between the level of Smad6 and hepatic pathological changes. Conclusion Feeding high fat diet with alcohol dilution on rats is a convenient,low cost method in establishing animal model of alcoholic liver disease. Inactivation of BMP-7 /Smad6 pathway contributes to the development of alcoholic liver fibrosis.