粪菌移植对小鼠抑郁样行为及海马内源性大麻素相关分子表达的影响

Effects of gut microbiota transplantation on depressive-like behaviors and expression of endocannabinoid-related molecules in the hippocampus of mice

目的观察抑郁症患者肠道菌群移植对小鼠行为及海马内源性大麻素(eCB)的影响。方法将健康受试者和抑郁症患者的肠道菌群移植给“伪无菌”小鼠,通过糖水偏好实验、旷场实验以及悬尾实验评估其抑郁样行为,并检测海马eCB相关分子的表达水平。①模型小鼠的制备:通过灌胃的方法使用“鸡尾酒”抗生素清除小鼠原有的肠道菌群,连续灌胃14 d,制备“伪无菌”小鼠模型;最后一次抗生素灌胃结束48 h后,将小鼠随机分为对照组(n=6)和抑郁组(n=10),每周一、三、五分别对各组小鼠进行粪菌移植,持续3周,制备“人源化”小鼠模型;②行为学测试:粪菌移植结束24 h后进行糖水偏好实验、旷场实验和悬尾实验以评估小鼠的抑郁样行为;③行为学测试结束24 h后处死动物,通过Western blotting法检测小鼠海马组织eCB相关分子的表达情况。结果①与对照组比较,抑郁组抑郁样行为表现显著,具体表现为旷场实验中抑郁组小鼠进入中心区次数、中心区运动距离、运动总距离、中心区停留时间比均显著降低(P<0.01),糖水偏好率显著下降(P<0.05),悬尾不动时间显著增长(P<0.01)。②Western blotting结果显示,与对照组比较,抑郁组CB1R、NAPE-PLD、DAGLα蛋白表达均显著下调,差异具有统计学意义(P<0.05)。③海马NAPE-PLD和DAGLα的表达水平与小鼠进入中心区次数、中心区运动距离、中心区停留时间比和糖水偏好率均呈正相关(均P<0.05),而且DAGLα、NAPE-PLD表达水平与悬尾不动时间均呈负相关(均P<0.05)。结论抑郁症患者肠道菌群可能通过调节海马eCB相关分子诱导小鼠产生抑郁样行为。

Objective To observe the effects of gut microbiota transplantation in patients with depression on the behaviors and endocannabinoid(eCB)in the hippocampus of mice.Methods The gut microbiota of healthy subjects and patients with depression were transplanted into“pseudo-sterile”mice,whose depressive-like behaviors were evaluated by sucrose preference test,open field test and tail suspension test,and the expression of eCB-related molecules in the hippocampus was detected.①Preparation of model mice:“cocktail”antibiotics were used to remove the original gut microbiota of mice by gavage,and the“pseudo-sterile”mouse model was prepared by gavage for 14 d.At 48 h after the last antibiotic gavage,mice were randomly divided into control group(n=6)and depression group(n=10).Fecal microbiota transplantation was performed on each group of mice on every Monday,Wednesday and Friday for 3 weeks,and the“humanized”mouse model was prepared.②Behavioral tests:sucrose preference test,open field test,and tail suspension test were performed 24 h after the fecal microbiota transplantation to evaluate depressive-like behaviors in mice.③Then mice were sacrificed 24 h after the behavioral tests,and the expression of eCB-related molecules in the hippocampus of mice was evaluated by Western blotting.Results①Compared with the control group,mice in depression group showed significant depressive-like behaviors in open field test,sucrose preference test and tail suspension test,including less times of entering the central area,decreased distance in central area,lower total distance,and reduced ratio of residence time in the central area in open field test(P<0.01),lower sucrose preference rate in sucrose preference test(P<0.05),and longer immobility time of tail suspension in tail suspension test(P<0.01).②The results of Western blotting showed that compared with the control group,the expressions of CB1R,NAPE-PLD,and DAGLαin the depression group were significantly down-regulated,with statistical significance(P<0.05).③The expressions of NAPE-PLD and DAGLαin hippocampus were positively correlated with the times of mice entering the central area,the distance in central area,the ratio of residence time in the central area,and sucrose preference rate(all P<0.05),while the expressions of DAGLαand NAPE-PLD were negatively correlated with the immobility time of tail suspension(both P<0.05).Conclusion The gut microbiota of patients with depression may induce depressive-like behaviors in mice by regulating eCB-related molecules in the hippocampus.