降脂益生菌对非酒精性脂肪性肝病小鼠肠黏膜屏障功能和肠道菌群的影响及机制

Effects of cholesterol-lowering probiotics on intestinal barrier and gut microbiota in mice with non-alcoholic fatty liver disease and the possible mechanisms

目的探讨降脂益生菌(DM9054和86066)对高脂饮食诱导的非酒精性脂肪性肝病(nonalcoholic fatty liver disease, NAFLD)小鼠肠黏膜屏障功能及肠道菌群结构的影响及其可能机制。方法24只雄性低密度脂蛋白受体基因敲除(Ldlr^-/-)小鼠随机分为对照组、模型组和干预组。对照组给予正常饮食,模型组及干预组给予高脂饮食,干预组同时给予鼠李糖乳杆菌联合植物乳杆菌灌胃,对照组及模型组给予生理盐水灌胃,连续高脂饮食12周建立小鼠NAFLD模型。干预结束后,qPCR法检测肝脏和肠道炎症因子、肝脏胆固醇合成基因的表达水平;HE染色评估肝脏、肠道病理变化;Western blot分析肠道紧密连接蛋白及肝脏HMG-CoA还原酶(HMG-CoA reductase, HMGCR)的蛋白质表达水平;16S rDNA高通量测序法分析各组小鼠肠道菌群变化特点。结果与模型组相比较,降脂益生菌干预显著减轻了高脂饮食诱导的小鼠肝脏脂肪变性、炎症反应,降低了肝脏胆固醇的合成(P<0.05)。此外,降脂益生菌干预亦减轻肠道炎症反应,改善高脂饮食对肠黏膜的屏障功能破坏,并且使肠道菌群结构趋于对照组(P<0.05)。结论降脂益生菌可能通过降低肝脏胆固醇的合成,减轻肝脏和肠道炎症,改善高脂饮食对肠黏膜的屏障功能破坏,调节肠道菌群结构,从而起到改善小鼠NAFLD的作用。

Objective To investigate the effects of cholesterol-lowering probiotics, DM9054 combined with 86066, on the intestinal mucosal barrier and gut microbiota in mice with nonalcoholic fatty liver disease (NAFLD) induced by high-fat diet and the possible mechanisms. Methods Twenty-four male mice deficient in the low-density lipoprotein receptor gene (Ldlr^-/- mice) were randomly divided into three groups including control, NAFLD model and probiotic intervention groups. Mice in the three groups were given normal chow diet+ normal saline, high-fat diet (HFD)+ normal saline, and HFD+ cholesterol-lowering probiotics, respectively. The mouse model of NAFLD was established by feeding mice with high-fat diet (45% of calories derived from fat diet) for 12 weeks. qPCR was performed to measure the expression of liver and intestinal inflammatory genes and liver cholesterol synthesis genes. Western blot assay was used to detect the expression of intestinal tight junction proteins and HMG-CoA reductase (HMGCR). Pathological changes in tissues were evaluated by HE staining. Features of gut microbiota were analyzed by 16S rRNA gene sequencing. Results Cholesterol-lowering probiotics intervention attenuated HFD-induced hepatic steatosis, inflammatory responses and obesity and decreased the synthesis of liver cholesterol (P<0.05). Moreover, inhibited gut inflammatory responses and improved intestinal barrier function were detected in the probiotic intervention group (P<0.05). The composition of gut microbiota in mice of the probiotic intervention group was different from that of the model group, but similar to that of the control group. Conclusions Cholesterol-lowering probiotics might attenuate NAFLD in mice through reducing liver cholesterol synthesis, alleviating liver and intestinal inflammation, improving intestinal mucosal barrier function and regulating intestinal microbiota.