五灵胶囊对肝纤维化大鼠肝组织TGF-β/Smad及Ras/ERK信号通路蛋白的影响

Effects of Wu Ling capsules on TGF-β/Smad and Ras/ERK signal pathway protein in liver tissue of liver fibrosis rats

目的探讨五灵胶囊对肝纤维化大鼠肝组织TGF-β/Smad及Ras/ERK信号通路蛋白的影响,阐述五灵胶囊治疗肝纤维化的作用机制。方法SD大鼠以高脂低蛋白饲料饲养,饮用10%乙醇,并腹腔注射4.1mol/LCCl4制备肝纤维化模型,给予五灵胶囊进行治疗1个月。Western blot检测肝组织中α-SMA、Ras、ERK、p-ERK、Smad2/3、p-Smad2/3、Smad7、TβRⅠ、TβRⅡ;RT-PCR法检测肝组织中COL-Ⅰ、COL-Ⅲ、TGF-β1mRNA;酶法测定血清ALT、AST;ELISA测定血清COL-Ⅰ、COL-Ⅲ含量;肝组织masson和HE染色进行肝细胞变性程度与胶原纤维增生程度评分。结果五灵胶囊可显著降低血清AST、ALT水平,下调肝组织中Rasp21、α-SMA、p-ERK、TβRⅠ蛋白与TGF-β1、COL-Ⅰ、COL-ⅢmRNA表达(P<0.01或0.05),增加p-Smad2/3蛋白表达(P<0.01),使肝细胞变性程度与胶原纤维增生程度显著降低。结论五灵胶囊治疗肝纤维化的作用机制是显著降低肝纤维化大鼠组织肝细胞变性和胶原纤维增生的程度、抑制TGF-β/Smad及Ras/ERK信号通路TβRⅠ、Rasp21、p-ERK蛋白表达及TGF-β1过量分泌。

Objective To investigate the influence of Wu Ling capsules （WL） on TGF-β/Smad and Ras/ERK signal pathway protein in liver tissue of liver fibrosis rats, and to expound the effect and mechanism of WL. Methods The rats liver fibrosis models were produced by subcutaneous injection of 4.1 mol/L CC14, drinking 10% alcohol water solution, feeding hyperliposis-low protein diet for 6 weeks. After 6 weeks, the rats liver fibrosis models were treated with WL 3.86 g/kg dosage by irrigate every day for 5 weeks. The expressions of α-SMA, Ras, ERK, p-ERK, Smad2/3, p- Smad2/3, Smad7, TI3R Ⅰ, TI3R Ⅱ were detected by Western blot. COL- Ⅰ , COL-Ⅲ , TGF-β1 mRNA were examined by RT-PCR. The levels of AST, ALT were detected by enzyme methods and the contents of COL- Ⅰ , COL-Ⅲ in serum were detected by ELISA. The staging of liver fibrosis was assessed by HE staining and the degree of collagen fibril hyperplasia was assessed through Masson. Results Wu Ling capsules might significantly decrease the levels of AST and ALT in serum. The expression levels of Ras p21 ,α-SMA, p-ERK, TI3R I and TGF-β1 , COL-Ⅰ , COL-Ⅲ mRNA were markedly down-regulated in liver fibrotic rats （P 〈 0.01 or 0.05）. The expression level of p-Smad2/3 was markedly augmentation. The extent of liver cells denaturalization and the degree of collagen fibril hyperplasia were saliently lower. Conclusion Wu Ling capsules could markedly depress the degree of liver cells denaturalization. Collagen fibril hyperplasia may be correlated with which inhibit the expression of TGF-β/Smad and Ras/ERK signal pathway protein as well as the excessive secrete of TGF-β1.