降脂益生菌调节胆固醇代谢改善小鼠非酒精性脂肪肝

The cholesterol-lowering probiotics improve NAFLD in mice by regulating cholesterol metabolism

目的研究一种由鼠李糖乳杆菌DM9054和植物乳杆菌86066构成的降脂益生菌组合对非酒精性脂肪性肝病(NAFLD)小鼠胆固醇代谢的影响及其可能机制。方法 24只雄性LDLR-/-小鼠随机分为对照组、模型组和益生菌干预组。高脂饮食(HFD)15周建立小鼠NAFLD模型,造模同时干预组给予鼠李糖乳杆菌DM9054联合植物乳杆菌86066灌胃,对照组和模型组给予等量生理盐水灌胃。实验过程中监测各组小鼠体重变化。实验结束后,检测小鼠血清甘油三酯(TG)、总胆固醇(TC)、低密度脂蛋白胆固醇(LDL)和高密度脂蛋白胆固醇(HDL)的水平差异。检测小鼠肝脏组织病理变化。使用Realtime PCR检测小鼠肠道内法尼脂受体(FXR)mRNA、顶端膜钠依赖的胆汁酸转运体(ASBT)mRNA、纤维生长因子15(FGF-15)mRNA和三磷酸腺苷结合盒转运体G5(ABCG-5)mRNA表达水平。Western blot检测小鼠肝脏胆固醇7α-羟化酶(CYP7A1)、FXR、三磷酸腺苷结合盒转运体G8(ABCG-8)、清道夫受体BI(SR-BI)、3-羟基-3-甲基戊二酸单酰辅酶A还原酶(HMGCR)、胆盐输出泵(ABCB-11)、纤维生长因子受体4(FGFR-4)和胆固醇调节元件结合蛋白-2(SREBP-2)蛋白表达水平。结果与模型组相比,降脂益生菌干预组小鼠体重减轻(P<0.05);小鼠血清TC、TG、LDL水平降低,HDL水平升高(P<0.05);小鼠肝脏脂肪变性和炎性细胞浸润的现象显著减少;小鼠肠道ASBT mRNA和ABCG-5mRNA表达水平明显降低(Ps<0.05),FGF-15mRNA表达水平明显升高(P<0.05),FXR mRNA表达水平差异无统计学意义(P>0.05);小鼠肝脏FGFR-4蛋白表达水平升高(P<0.05),SREBP-2和HMGCR蛋白表达水平降低(Ps<0.05),FXR、CYP7A1、SR-BI、ABCG-8和ABCB-11蛋白表达水平差异无统计学意义(Ps>0.05)。结论降脂益生菌可能通过激活FXR-FGF15通路调节胆汁酸代谢;通过下调SREBP-2表达水平,抑制HMGCR表达,减少胆固醇的生成,从而起到改善非酒精性脂肪肝的作用。

Objective To explore the effects of DM9054（Lactobacillus rhamnosus GG,LGG）combined with86066（Lactobacillus plantarum WCFS1,LP）on cholesterol metabolism in mouse models of non-alcoholic fatty liver disease（NAFLD）and analyze the possible mechanism.Methods Twenty-four SD male mice were randomly divided into three groups including control group,NAFLD model group or probiotics intervention group.The control group received normal diet.The mouse models of NAFLD was established by feeding chronic high fat diet for 15 weeks.The probiotics intervention group was given high fat diet together with cholesterol-lowering probiotics through oral gavage.General indexes of each group including body weight,levels of triglyceride（TG）,total cholesterol（TC）,low density lipoprotein cholesterol（LDL）and high density lipoprotein cholesterol（HDL）were observed.The pathological changes of liver tissue of mice were detected.The expressions of farnesoid X receptor（FXR）,apical sodium-dependent bile acid transparter（ASBT）,fibroblast grwoth factor 15（FGF15）and ATP-binding cassette sub-family G member 5（ABCG-5）at mRNA level were detected by using real-time polymerase chain reaction（Real-time PCR）.Western blot assay was used to detect the expressions of cholesterol 7 alpha-hydroxylase（CYP7 A1）,FXR,ATP-binding cassette sub-family G member 8（ABCG-8）,scavenger receptor class B type 1（SR-BI）,3-hydroxy-3-methyl-glutaryl-coenzyme A reductase（HMGCR）,ATP-binding cassette,sub-family B member 11（ABCB-11）,fibroblast growth factor receptor 4（FGFR-4）and sterol regulatory element-binding protein 2（SREBP-2）in liver tissues.Results Compared with the model group,the body weights of mice in the intervention group reduced（P〈0.05）;the serum levels of TC,TG and LDL decreased while that of HDL increased（P〈0.05）;the level of hepatic steatosis and infiltration of inflammatory cells significantly reduced;the expressions of ASBT mRNA and ABCG-5 mRNA in the intestine significantly decreased（Ps〈0.05）while that of FGF-15 mRNA significantly increased（P〈0.05）;the expression level of FXR mRNA did not show statistical difference（P〉0.05）.The expression of FGFR-4 protein in liver increased（P〈0.05）,while those of SREBP-2 and HMGCR decreased（Ps〈0.05）;the expressions of FXR,CYP7 A1,SR-BI,ABCG-8 and ABCB-11 protein did not show significant differences（Ps〉0.05）.Conclusion Probiotics intervention may regulate the bile acid metabolism by activating the FXR-FGF15 pathway and inhibiting the expression of ASBT.Treating NAFLD mice with cholesterol-lowering probiotics can reduce the expression of SREBP-2,inhibit the expression of HMGCR in liver tissue and reduce the formation of cholesterol,which contributes to the alleviation of NAFLD.