电针干预不同时间对慢性应激抑郁模型大鼠海马组织miRNA表达的影响

Effect of Electroacupuncture for Different Time on the Expression of MiRNA in Hippocampus of Chronic Stress Depression Model Rats

目的从miRNA水平探讨电针不同干预时间抗抑郁的可能作用机制。方法50只Wistar大鼠采用随机数字表法分为正常组10只、模型组10只、氟西汀组10只和电针组20只。除正常组外,其他各组大鼠采用慢性不可预知温和应激28天方法构建慢性应激抑郁大鼠模型。在造模同时,氟西汀组大鼠予氟西汀片20 mg/(kg·d)灌胃;电针组大鼠电针“印堂”和“百会”,均干预4周。应激前与干预2、4周后各组大鼠进行糖水偏好实验、强迫游泳实验。电针组分别于干预2周、4周后选取10只大鼠取海马,正常组、模型组于干预4周后取大鼠海马,采用基因芯片微阵列分析和定量实时聚合酶链反应(qRT-PCR)检测海马组织miRNA(miR-383-5p,miR-764-5p,miR-675-3p)表达差异,采用生物信息学检测方法发现其调控的靶基因和富集的信号通路。结果干预2、4周后,模型组大鼠糖水偏好度显著低于正常组,强迫游泳不动时间显著长于正常组(P<0.01)。氟西汀组干预4周后与电针组干预2、4周后,大鼠糖水偏好度与强迫游泳不动时间均较模型组改善(P<0.05或P<0.01)。经基因芯片分析与qRT-PCR检测,与模型组比较,干预2周后电针组大鼠海马miR-675-3p表达上调(P<0.01),调控的靶基因多涉及GABA能突触、多巴胺能突触及PI3K-Akt信号通路等;干预4周后miR-383-5p、miR-764-5p表达下调(P<0.05或P<0.01),涉及MAPK信号通路、神经营养素信号通路等多条信号通路。结论电针对抑郁症的治疗可能在不同时程分别刺激调节抑郁躯体症状和心理障碍相关的不同的miRNA及靶基因相关。

Objective To explore the possible mechanism of antidepressant effects of electroacupuncture at different intervention times on the miRNA level.Methods A total of 50 Wistar rats were divided into a normal group(n=10),a model group(n=10),a fluoxetine group(n=10)and a electroacupuncture group(n=20)by random number table method.Except for the normal group,chronic unpredictable mild stress was used for 28 days to construct a chronic stress depression rat model.At the same time of modeling,rats in the fluoxetine group were given fluoxetine tablets 20 mg/(kg·d)by gavage;rats in the electroacupuncture group were given electroacupuncture with“Yintang”and“Baihui”for 4 weeks.Before stress and after 2 and 4 weeks of intervention,rats in each group were subjected to sugar water preference experiment and forced swimming experiment.In the electroacupuncture group,10 rats were selected to take the hippocampus after 2 weeks and 4 weeks of intervention respectively,and rats in the normal group and model group were taken from the hippocampus after 4 weeks of intervention.Gene chip microarray analysis and quantitative real-time polymerase chain reaction were used(QRT-PCR)to detect the expression differences of miRNA(miR-383-5p,miR-764-5p,miR-675-3p)in hippocampus,and bioinformatics detection methods was used to discover its regulated target genes and enriched signal pathways.Results After 2 and 4 weeks of intervention,the sugar water preference of the model group was significantly lower than that of the normal group,and the time of forced swimming immobility was significantly higher than that of the normal group(P<0.01).After 4 weeks of intervention in the fluoxetine group and 2 and 4 weeks in the electroacupuncture group,the rats'sugar preference and forced swimming immobility time were improved compared with the model group(P<0.05 or P<0.01).After gene chip analysis and qRT-PCR detection,compared with the model group,the expression of miR-675-3p in the hippocampus of rats in the electroacupuncture group was up-regulated(P<0.01)after 2 weeks of intervention,and the regulated target genes mostly involved GABAergic synapse,Dopaminergic synapse and PI3K-Akt signaling pathway and other signaling pathways;miR-383-5p,miR-764-5p expression was down-regulated after 4 weeks of intervention(P<0.05 or P<0.01),involving MAPK signaling pathway,neurotrophin signaling pathway etc.Conclusion The electro-acupuncture treatment of depression may stimulate different miRNAs and target genes related to the physical symptoms and psychological disorders of depression in different time courses.