红景天苷调节肠道菌群治疗非酒精性脂肪性肝病的效应机制

The mechanism of effect of Salidroside modulating gut flora in treating nonalcoholic fatty liver disease

目的运用粪菌移植实验,围绕肠道菌群探讨红景天苷治疗非酒精性脂肪性肝病(nonalcoholic fatty liver disease,NAFLD)的作用机制。方法将40只高脂饮食12周诱导的NAFLD小鼠,随机分为对照组(Control组)、模型组(HFD组)、红景天苷组(Sal组)、模型粪菌移植组(HFD-FMT组)和红景天苷粪菌移植组(Sal-FMT组),每组8只。第7周起,Sal组小鼠以50 mg/(kg·d)的剂量灌胃给药,HFD-FMT组、SalFMT组给予10 mL/(kg·d)剂量的粪菌移植液灌胃,Control组和HFD组小鼠分别以等体积生理盐水灌胃,持续6周。观察以下指标:(1)苏木精-伊红染色(HE)观察肝组织病理变化并计算NAFLD活动度评分(NAS);(2)生化试剂盒法检测小鼠肝组织甘油三酯(TG)含量、血清转氨酶(ALT、AST)活性和血脂(TG、TC、LDL-C、HDL-C)含量;(3)16S rRNA方法分析肠道菌群多样性。结果与Control组相比,HFD组小鼠HE染色见大量脂滴空泡并伴有炎性细胞浸润现象,体重、肝重、肝组织TG含量、肝组织NAS评分、血清ALT、AST活性和血清TG、TC、LDL-C含量均显著升高(t=5.811,P<0.001;t=2.278,P=0.020;t=6.505,P<0.001;t=10.360,P<0.001;t=3.559,P=0.002;t=3.836,P<0.001;t=5.767,P<0.001;t=3.314,P=0.003;t=3.972,P<0.001);与HFD组相比,Sal组小鼠肝组织病变减轻,肝组织TG含量、肝组织NAS评分和血清ALT活性均显著降低(t=6.375,P<0.001;t=7.095,P<0.001;t=1.919,P=0.038);与HFD-FMT组相比,Sal-FMT组小鼠肝组织病变减轻,肝组织TG含量、肝组织NAS评分和血清ALT、AST活性均显著降低(t=5.071,P<0.001;t=8.775,P<0.001;t=1.935,P=0.038;t=4.725,P<0.001);与HFD组相比,HFDFMT组小鼠肝组织病变相似,体重、肝重、肝组织TG含量、肝组织NAS评分、血清ALT、AST活性和血清TG、TC、LDL-C含量差异均无统计学意义(t=0.337,P=0.741;t=0.408,P=0.690;t=0.428,P=0.675;t=0.798,P=0.438;t=0.323,P=0.752;t=1.671,P=0.117;t=0.606,P=0.555;t=0.496,P=0.627;t=0.174,P=0.864);与Sal组相比,Sal-FMT组小鼠肝组织病变相似,体重、肝重、肝组织TG含量、肝组织NAS评分、血清ALT、AST活性和血清TG、TC、LDL-C含量差异均无统计学意义(t=0.309,P=0.762;t=0.193,P=0.850;t=1.674,P=0.118;t=0.424,P=0.678;t=0.083,P=0.935;t=0.425,P=0.677;t=0.063P=0.951;t=0.873,P=0.398;t=0.388,P=0.704)。此外,16S rRNA基因肠道菌群分析显示,红景天苷能够改变肠道菌群的组成结构,调节菌群的丰度。结论红景天苷调节的肠道菌群能够有效治疗NAFLD,调节肠道菌群的组成与丰度可能是红景天苷发挥治疗效果的机制。

Objective To explore the mechanism of action of Salidroside in the treatment of nonalcoholic fatty liver disease(NAFLD)based on gut flora by using fecal microbiota transplantation(FMT).Methods Fourty NAFLD mice induced by 12 weeks of high-fat diet were randomly divided into normal group(Control group),model group(HFD group),Salidroside group(Sal group),FMT group of the model(HFD-FMT group)and FMT group of Salidroside(Sal-FMT group),with 8 mice in each group.Beginning from the 7th week,the mice in the Sal group were administered by gavage at a dose of 50 mg/(kg•d),those in the HFD-FMT group and Sal-FMT group were given fecal transplantation solution by gavage at a dose of 10 mL/(kg•d),and those in the Control group and HFD group were administered by gavage an equivalent volume of physiological saline,for 6 weeks.(1)HE was used to observe the histopathological changes of liver and calculate the NAFLD Activity Score(NAS);(2)biochemical kit method was used to detect liver triglyceride(TG)content,serum aminotransferase(ALT,AST)activities and serum lipid(TG,TC,LDL-C,HDL-C)contents;(3)16S rRNA method was used to analyze gut flora diversity.Results Compared with the Control group,HFD group mice showed massive fat vacuoles with inflammatory cell infiltration in HE staining;and the body weight,liver weight,liver TG content,liver NAS score,serum ALT and AST activities and serum levels of TG,TC and LDL-C significantly increased(t=5.811,P<0.001;t=2.278,P=0.020;t=6.505,P<0.001;t=10.360,P<0.001;t=3.559,P=0.002;t=3.836,P<0.001;t=5.767,P<0.001;t=3.314,P=0.003;t=3.972,P<0.001);compared with the HFD group,Sal group mice showed alleviated hepatic lesions,and the liver TG content,liver NAS score and serum ALT activity significantly decreased(t=6.375,P<0.001;t=7.095,P<0.001;t=1.919,P=0.038);compared with the HFD-FMT group,Sal-FMT group mice showed alleviated hepatic lesions,and the liver TG content,liver NAS score and serum ALT and AST activities significantly decreased(t=5.071,P<0.001;t=8.775,P<0.001;t=1.935,P=0.038;t=4.725,P<0.001);compared with the HFD group,HFD-FMT group mice showed similar hepatic lesions,with no statistically significant differences in body weight,liver weight,liver TG content,liver NAS score,serum ALT and AST activities and serum levels of TG,TC and LDL-C(t=0.337,P=0.741;t=0.408,P=0.690;t=0.428,P=0.675;t=0.798,P=0.438;t=0.323,P=0.752;t=1.671,P=0.117;t=0.606,P=0.555;t=0.496,P=0.627;t=0.174,P=0.864);compared with the Sal group,Sal-FMT group mice showed similar hepatic lesions,with no statistically significant differences in body weight,liver weight,liver TG content,liver NAS score,serum ALT and AST activities and serum levels of TG,TC and LDL-C(t=0.309,P=0.762;t=0.193,P=0.850;t=1.674,P=0.118;t=0.424,P=0.678;t=0.083,P=0.935;t=0.425,P=0.677;t=0.063 P=0.951;t=0.873,P=0.398;t=0.388,P=0.704).Furthermore,16S rRNA gut flora analysis revealed that Salidroside changed the composition structure of gut flora and regulated the abundance of the flora.Conclusion Gut flora regulated by Salidroside is effective in treating NAFLD,and regulating the composition and abundance of gut flora may be the mechanism of the therapeutic effect of Salidroside.