柞蚕雄蛾液对酒精肝纤维化小鼠的预防作用及其机制研究

Preventive Effect and Mechanism of Male China Oak Silkworm Liquid on the Alcoholic Liver Fibrosis in Mice

目的:观察柞蚕雄蛾液对酒精诱导的小鼠肝纤维化的预防作用并探讨其可能的机制。方法:选择40只SPF级C57BL/6J小鼠,随机分为对照组、模型组、柞蚕雄蛾液低剂量组、柞蚕雄蛾液高剂量组,每组各10只。除对照组外,其他各组均通过酒精灌胃建立酒精性肝纤维化模型。柞蚕雄蛾液低剂量组、柞蚕雄蛾液高剂量组每天先予柞蚕雄蛾液灌胃,两小时后予以酒精灌胃。16周后,所有小鼠禁食12 h后通过眼球取血后处死,取出肝脏。采用HE和Masson染色观察肝组织病理改变并进行肝纤维化分级;检测血清中ALT及AST的含量,免疫组化法检测肝组织转化生长因子β1(TGF-β1)、基质金属蛋白酶抑制剂-1(TIMP-1)、结缔组织生长因子(CTGF)的表达,Western blot法测定肝组织中Smad2/3的蛋白表达。结果:柞蚕雄蛾液低剂量组镜下只见少量点状坏死和炎细胞浸润,肝组织损伤比模型组明显减轻。与对照组比较,模型组TGF-β1、TIMP-1、CTGF、Smad2/3的表达均明显增高(P<0.001),血清ALT及AST水平显著升高(P<0.001);与模型组比较,柞蚕雄蛾低剂量组TGF-β1、TIMP-1、CTGF、Smad2/3表达明显降低(P<0.001),血清ALT及AST水平显著下降(P<0.001)。结论:柞蚕雄蛾液对酒精诱导的小鼠肝纤维化具有显著的预防作用,低剂量柞蚕雄蛾液较高剂量柞蚕雄蛾液可能更有效,其机制可能与调控TGF-β1/Smad2/3信号转导通路,下调TIMP及CTGF抑制HSC的活化与增殖有关。

Objective： To observe the preventive effect of male China oak silkworm liquid on the alcoholic liver fibrosis in mice and explore its possible mechanism. Methods： 40 SPF C57BL/6J mice were randomly selected and divided into the control group, model group, low-dose group of male China oak silkworm liquid and high-dose group of male China oak silkworm liquid, 10 mice in each group. Except the control group, the other groups were gavage with alcohol to establish the model of alcoholic liver fibrosis. The low-dose group of male China oak silkworm liquid and the high-dose group of male China oak silkworm liquid were gavage with male China oak silkworm liquid firstly every day, and then alcohol two hours later.16 weeks later, all the mice were put to death 12 hour after fasting diet to get blood by eye sampling and remove the liver. HE and Masson staining were used to observe the pathological change in liver tissue and the liver fibrosis grading; The levels of AST and ALT in serum were detected, immunohistochemical method was used to detect the expression of transforming growth factor-β1（TGF-β1）, tissue inhibitor of metalloproteinases-1（TIMP-1） and connectivet issue growth factor（CTGF） in liver tissue, Western blot method was used to measure the Smad2/3 protein expression. Results： Less punctate necrosis and infiltration of inflammatory cells were observed in the low-dose group of male China oak silkworm liquid and the liver tissue injury was remarkably lower than that of the model group. Compared with the control group, the expressions of TGF-β1, TIMP-1, CTGF,Smad2/3 were significantly increased in the model group（P〈0.001）, the serum ALT and AST levels were significantly increased（P〈0.001）; compared with the model group, the expressions of TGF-β1, TIMP-1, CTGF, Smad2/3 were significantly lower in the low-dose group of male China oak silkworm liquid（P〈0.001）, the serum ALT and AST levels were significantly decreased（P〈0.001）. Conclusion：The male China oak silkworm liquid played a significant role in the prevention of the alcohol-induced liver fibrosis in mice, low-dose of male China oak silkworm liquid might be more effectively than the high-dose. The mechanism might be related to the regulation of TGF-β1/Smad2/3 signaling pathway, down regulation of TIMP and CTGF in the inhibition of HSC activation and proliferation.