基于代谢组学研究蒙药苏格木勒-4对失眠大鼠的改善作用机制

Metabolomics-based study on the improvement mechanism of the Mongolian drug Sugemule-4 on insomnia rats

目的研究蒙药苏格木勒-4对失眠大鼠的代谢影响,初步探讨其改善失眠的可能机制。方法采用夹尾刺激和腹腔注射对氯苯丙氨酸溶液复合造模法建立慢性应激失眠大鼠模型。将24只雄性大鼠随机分为正常组、模型组、地西泮组(阳性对照,0.92 mg/kg)和苏格木勒-4组(5.2 g/kg),每组6只。从夹尾刺激第7天开始,苏格木勒-4组、地西泮组大鼠开始灌胃给药,正常组、模型组大鼠灌胃等体积蒸馏水,每天灌胃1次,连续14 d。利用水迷宫实验测试大鼠的学习和记忆能力,利用无创睡眠活动监测系统对大鼠24 h睡眠时间进行监测。采用超高效液相色谱串联质谱技术对大鼠血清和海马组织进行代谢组学研究。使用多元统计分析方法对各组大鼠血清和海马组织中差异代谢物进行分析,筛选各组间的差异代谢物以及代谢通路。结果与正常组比较,模型组大鼠的逃避潜伏期显著延长、穿越平台次数显著减少、平均24 h睡眠时间百分比显著降低(P<0.05)。与模型组比较,地西泮组和苏格木勒-4组大鼠上述指标水平均显著改善(P<0.05)。代谢组学研究发现,大鼠血清和海马组织中共鉴定出5-羟基吲哚乙酸、犬尿氨酸、犬尿喹啉酸、5-羟色胺、硫酸苯酚、1-羧基乙基酪氨酸、3-(4-羟基苯基)乳酸、N-乙酰基酪氨酸、酪氨酸共9个差异代谢物,主要涉及色氨酸和酪氨酸2条代谢通路。结论苏格木勒-4可以改善失眠大鼠睡眠时间和行为学表现,其机制可能与调节色氨酸、酪氨酸等氨基酸代谢途径有关。

OBJECTIVE To study the effects of the Mongolian medicine Sugemule-4 on the metabolism of insomnia rats,and to preliminarily explore its possible mechanisms for improving insomnia.METHODS The rat model of chronic stress insomnia was established by tail clipping stimulation and intraperitoneal injection of p-chlorophenyl alanine solution.Twenty-four male rats were randomly divided into the normal group,model group,diazepam group(positive control,0.92 mg/kg),and Sugemule-4 group(5.2 g/kg),with 6 rats in each group.Since the 7th day of tail clipping stimulation,the Sugemule-4 group and diazepam group began to be intragastrically administered with relevant medicine;the normal group and model group were intragastrically administered with an equal volume of distilled water,once a day,for 14 consecutive days.The learning and memory abilities of rats were tested using a water maze experiment,and the non-invasive sleep activity monitoring system was used to monitor the 24-hour sleep time of rats.A metabolomics study was conducted on rat serum and hippocampal tissue by using ultra-high-performance liquid chromatography-tandem mass spectrometry.The multivariate statistical analysis method was adopted to analyze the differential metabolites in serum and hippocampal tissue of rats,and screen for differential metabolites and metabolic pathways among those groups.RESULTS Compared with the normal group,the escape latency of rats in the model group was significantly increased,the times of crossing platforms were significantly reduced,and the percentage of average 24-hour sleep time was significantly reduced(P<0.05).Compared with the model group,the levels of the above indicators were significantly reversed in the diazepam group and Sugemule-4 group(P<0.05).Metabolomics studies found that a total of 9 differential metabolites were identified in rat serum and hippocampal tissue,including 5-hydroxyindoleacetic acid,canine urate,canine urinary quinolinic acid,5-hydroxytryptamine,phenol sulfate,1-carboxyethyltyrosine,3-(4-hydroxyphenyl)lactate,N-acetyl tyrosine,tyrosine and phenol sulfate,mainly involving 2 metabolic pathways of tryptophan and tyrosine.CONCLUSIONS Sugemule-4 can improve the sleep time and behavioral performance of insomnia rats,and its mechanism may be associated with affecting amino acid metabolic pathways such as tryptophan and tyrosine.