肠道菌群及5-HT在大气PM_(2.5)暴露致小鼠抑郁焦虑样行为中的作用

Role of intestinal flora and 5-HT in depression-and anxiety-like behaviors in mice exposed to PM_(2.5)

[背景]研究表明,PM_(2.5)暴露与中枢神经系统疾病密切相关,可引起认知功能障碍,也会改变肠道菌群的组成,但关于肠道菌群在PM_(2.5)所致小鼠抑郁焦虑样行为中的作用研究很少。[目的]观察大气PM_(2.5)暴露对小鼠抑郁焦虑样行为及肠道菌群组成的影响,探讨肠道菌群调控5-羟色胺(5-HT)在大气PM_(2.5)暴露致小鼠抑郁焦虑样行为中的作用。[方法]8周龄雄性SPF级C57BL/6J小鼠随机分为对照组(NS组)、益生菌组(LGG组)、PM_(2.5)组(PM组)、联合暴露组(PML组),每组6只。PM组和PML组小鼠采用细颗粒物实时染毒系统进行PM_(2.5)暴露,浓度为室外的8倍,每天6 h,每周6 d,共7周。LGG组和PML组小鼠灌胃鼠李糖乳杆菌,NS组和PM组灌胃等量生理盐水。高架十字迷宫实验和旷场实验评价小鼠焦虑和抑郁状态。利用16S rRNA高通量测序技术测定小鼠粪便中肠道菌群丰度、多样性和结构组成。ELISA法检测血清和海马组织中5-HT水平。Spearman相关分析肠道差异菌群与海马组织5-HT水平、小鼠抑郁焦虑样行为指标的相关关系。[结果]PM组小鼠开臂次数百分比的M(P25,P75)为0.0%(0.0%,33.3%),低于NS组[47.7%(25.0%,50.8%)]和PML组[46.9%(40.0%,50.0%)],差异有统计学意义(P<0.05)。PM组运动总距离和中央区停留时间()分别为(2.01±0.90)m、(10.31±1.99)s,均低于NS组[(3.80±0.89)m,(14.47±3.07)s];PML组运动总距离为(2.73±1.12)m,低于NS组和LGG组[(4.21±1.08)m],差异均有统计学意义(P<0.05)。与NS组相比,PM组Simpson指数升高(P<0.05);与LGG组相比,PML组的Simpson指数下降(P<0.05)。Beta多样性分析结果表明各组小鼠之间肠道菌群组成存在差异。与NS组和LGG组相比,PM组及PML组丹毒丝菌科、杜氏杆菌属细菌丰度升高,普雷沃氏菌科_UCG-001属丰度降低,差异有统计学意义(P<0.05)。PM组小鼠海马组织5-HT水平为(135.02±10.31)μg·g^(-1),低于NS组[(178.77±43.15)μg·g^(-1)]和LGG组[(224.85±22.98)μg·g^(-1)];PML组小鼠5-HT水平[(161.27±15.81)μg·g^(-1)]低于LGG组(P<0.05)。海马组织中5-HT水平与普雷沃氏菌科_UCG-001属的相对丰度呈正相关(r=0.6090,P=0.012),开臂次数百分比与杜氏杆菌属的相对丰度呈负相关(r=−0.4630,P=0.023)。[结论]大气PM_(2.5)暴露可引起小鼠抑郁焦虑样行为,并与肠道菌群的多样性及相对丰度改变和5-HT水平降低有关,添加益生菌后抑郁焦虑样行为得以改善。

BackgroundSome studies have shown that PM_(2.5) exposure is closely related to central nervous system diseases that lead to cognitive dysfunction and change the composition of intestinal flora.However,there are few studies on the role of intestinal flora in PM_(2.5)-induced depression-and anxiety-like behaviors in mice.ObjectiveTo observe the effects of PM_(2.5) exposure on depression-and anxiety-like behaviors and the composition of intestinal flora in mice,and to explore the role of intestinal flora in regulating 5-hydroxytryptamine(5-HT)in depression-and anxiety-like behaviors in mice exposed to PM_(2.5).MethodsEight-week-old male SPF C57BL/6J mice were randomly divided into control group(NS group),probiotic group(LGG group),PM_(2.5) group(PM group),and combined exposure group(PML group),6 mice in each group.Mice in the PM group and the PML group were exposed to PM_(2.5) in a dynamic exposure cabinet for 6 h per day,6 d a week for 7 consecutive weeks,and the PM_(2.5) concentrations were approximately 8 times higher than the outdoor concentration.The LGG group and the PML group were orally administered with Lactobacillus rhamnosus while the NS group and the PM group were orally administered with the same amount of saline.Elevated plus maze test and open field test were used to detect depression and anxiety in mice.Fecal samples of mice were collected to evaluate intestinal flora abundance,diversity,and structure between groups using high-throughput sequencing of 16S rRNA.ELISA was employed to detect the levels of 5-HT in serum and hippocampus.Spearman correlation was used to analyze the correlations of differential intestinal flora with 5-HT level in hippocampus and depression-and anxiety-like behavior indicators in mice.ResultsThe percentage of open-arm entry[M(P25,P75)]in the PM group was 0.0%(0.0%,33.3%),lower than those in the NS group[47.7%(25.0%,50.8%)]and the PML group[46.9%(40.0%,50.0%)],and the differences were statistically significant(P<0.05).The total travelled distance and the time spent in central area()in the PM group were(2.01±0.90)m and(10.31±1.99)s respectively,shorter than those of the NS group[(3.80±0.89)m,(14.47±3.07)s],the total travelled distance in the PML group[(2.73±1.12)m]was shorter than those of the NS group and the LGG group[(4.21±1.08)m],and the differences were statistically significant(P<0.05).Compared to the NS group,the Simpson index of the PM group significantly increased(P<0.05).Compared to the LGG group,the Simpson index of the PML group significantly decreased(P<0.05).The results of Beta diversity analysis showed that there were differences in the composition of intestinal flora among the four groups of mice.Compared with the NS group and the LGG group,the abundances of Erysipelotrichaceae and Dubosiella in the PM group and the PML group increased,while the abundances of Prevotellaceae_UCG-001 decreased,and the differences were statistically significant(P<0.05).In hippocampus,the level of 5-HT in the PM group[(135.02±10.31)μg·g^(-1)]was lower than those in the NS group[(178.77±43.15)μg·g^(-1)]and the LGG group[(224.85±22.98)μg·g^(-1)],and the level of 5-HT in the PML group[(161.27±15.81)μg·g^(-1)]was lower than that in the LGG group(P<0.05).5-HT level in hippocampus was significantly positively correlated with the relative abundance of Prevotellaceae_UCG-001(r=0.6090,P=0.012).The percentage of open-arm entry was significantly negatively correlated with the relative abundance of Dubosiella(r=−0.4630,P=0.023).ConclusionAtmospheric PM_(2.5) exposure may cause depression-and anxiety-like behaviors in mice.The observed behavior dysfunction may be associated with the changes in diversity and relative abundance of intestinal flora as well as the decrease of 5-HT level.Such depression-and anxiety-like behaviors are alleviated after adding probiotics.