基于16S rDNA测序研究当归芍药散调控肠道菌群改善SAMP8小鼠认知能力的作用机制

Mechanisms of Danggui Shaoyaosan in Improving Cognitive Ability in SAMP8 Mice by Regulating Gut Microbiota via 16S rDNA Sequencing

目的:基于16S rDNA测序研究当归芍药散(DSS)对阿尔茨海默病(AD)模型小鼠SAMP8肠道菌群的影响。方法:按随机数字表法,将24只7月龄SAMP8小鼠分为DSS低、中、高剂量组(14.4、28.8、57.6 g·kg^(-1)·d^(-1))及模型组,另取6只同月龄SAMR1小鼠,为正常组。连续给药8周后,采用16S rDNA高通量测序技术检测小鼠粪便肠道菌群的变化;Morris水迷宫实验评价小鼠定位航行与空间探索能力;尼氏染色观察海马CA1区神经元病理变化;酶联免疫吸附测定法(ELISA)检测海马β淀粉样蛋白(Aβ)和Tau蛋白过度磷酸化(p-Tau)蛋白含量。结果:与正常组比较,模型组小鼠肠道菌群α多样性降低、β多样性明显改变;逃避潜伏期延长(P<0.05),穿越平台象限次数和平台象限停留时间明显减少(P<0.05);CA1区神经元尼氏小体数量明显减少(P<0.05);Aβ和p-Tau蛋白水平明显升高(P<0.05)。DSS干预后,各组小鼠肠道菌群α多样性均增加,中、高剂量组β多样性与正常组相似、以中剂量组效果最佳;在门水平上,厚壁菌门丰度明显增加(P<0.05),拟杆菌门、变形菌门丰度明显降低(P<0.05);在属水平上,乳酸杆菌等菌属丰度明显增加(P<0.05),拟杆菌、幽门螺杆菌、理研菌、副杆状菌、萨特氏菌、黏液性菌等菌属丰度明显降低(P<0.05);小鼠逃避潜伏期明显缩短(P<0.05)、穿越平台象限次数和平台象限停留时间明显增加(P<0.05);尼氏小体数量明显增加(P<0.05),Aβ和p-Tau蛋白水平明显降低(P<0.05);Pearson相关性分析表明黏液性菌、拟杆菌和萨特氏菌菌属丰度与SAMP8小鼠认知能力呈负相关,乳酸杆菌和Butyricimonas菌属丰度与SAMP8小鼠认知能力呈正相关。结论:DSS可改善SAMP8小鼠认知能力,其作用机制可能与调节肠道菌群多样性、群落组成有关。

Objective:To analyze the effects of Danggui Shaoyaosan(DSS)on the gut microbiota of the Alzheimer’s disease(AD)model in SAMP8 mice based on 16S rDNA sequencing.Method:Twenty-four SAMP8 mice aged seven months were randomly divided into low-,medium-,and high-dose DSS groups(14.4,28.8,57.6 g·kg^(-1)·d^(-1))and a model group according to a random number table,with six rats in each group.Six SAMR1 mice of the same age were assigned to the normal group.After intragastric administration for eight consecutive weeks,16S rDNA sequencing was performed to detect the gut microbiota of feces in mice.Morris water maze was employed to assess the directional navigation and space exploration ability of mice.Nissl staining was performed to observe the pathological changes of neurons in the hippocampal CA1 area.Enzymelinked immunosorbent assay(ELISA)was adopted to measure the protein content of hippocampal amyloid β-protein(Aβ)and hyperphosphorylated Tau(p-Tau).Result:Compared with the normal group,the model group presented a decliningαdiversity(P<0.05),markedly altered β diversity,prolonged escape latency(P<0.05),reduced number of platform crossings and cumulative duration in the targeted quadrant(P<0.05),decreased neurons and Nissl bodies in the CA1 hippocampal area,and up-regulated Aβ and p-Tau expression(P<0.05).However,DSS intervention enhanced theαdiversity,and medium-and high-dose DSS,especially the medium-dose DSS,could result inαdiversity similar to the control group.Moreover,at the phylum level,the abundance of Firmicutes increased(P<0.05),while the abundance of Bacteroidetes and Proteobacteria decreased(P<0.05).At the genus level,the abundance of Lactobacillus and other genera increased(P<0.05),while the abundance of Bacteroides,Helicobacterium,Rikenella,Parabacteroides,Sutterella,and Mucilaginibacter decreased(P<0.05).The DSS groups also showed shortened escape latency(P<0.05),increased number of platform crossings and cumulative duration in the targeted quadrant(P<0.05),increased Nissl bodies(P<0.05),and reduced Aβ and p-Tau content(P<0.05).Pearson correlation analysis showed that the abundance of Mucilaginibacter,Bacteroides,and Sutterella was negatively correlated with the cognitive ability of SAMP8 mice,while the abundance of Lactobacillus and Butyricimonas was positively correlated with the cognitive ability of SAMP8 mice.Conclusion:DSS can improve the cognitive ability of SAMP8 mice,and its mechanism may be related to the regulation of gut microbiota diversity and community composition.