探讨半夏-薏苡仁对PCPA失眠模型大鼠海马食欲素及其受体和血清细胞因子的调控作用  

Regulatory effects of Pinellia-Coix Seed on hippocampal orexin, its receptors and serum cytokines in PCPA insomnia model rats

作　　者：王亮 王学林[1,2] 王鹏 李晨 谌盈帆[1,2] 陈旸 张印[2] 李绍旦 杨明会 WANG Liang;WANG Xuelin;WANG Peng;LI Chen;CHEN Yingfan;CHEN Yang;ZHANG Yin;LI Shaodan;YANG Minghui(Chinese PLA Medical School,Beijing 100853,China;Department of Traditional Chinese Medicine,the Sixth Medical Center,Chinese PLA General Hospital,Beijing 100853,China)

机构地区：[1]解放军医学院,北京100853 [2]解放军总医院第六医学中心中医医学部,北京100089

出　　处：《解放军医学院学报》2022年第4期472-478,496,共8页Academic Journal of Chinese PLA Medical School

基　　金：国家自然科学基金资助项目(82174283);国家中医药领军人才支持计划资助项目(国中医药人教发[2018]12号)。

摘　　要：背景失眠症的发病率逐年增加,对人类身心健康造成严重危害。半夏-薏苡仁(BX-YYR)主要有效成分包括黄芩素、β-谷甾醇、豆甾醇等,但其治疗失眠症的具体作用机制尚不明确。目的 研究BX-YYR干预对氯苯丙氨酸(4-Chloro-DLphenylalanine,PCPA)失眠模型大鼠的食欲素(Orexin)及其受体、细胞相关因子的调控作用。方法 采用PCPA诱导失眠大鼠模型,将实验大鼠分为6组,每组15只,分别为正常组(N)、模型组(M)、地西泮组(D)和BX-YYR低(ZL)、中(ZM)、高(ZH)剂量组,各组按10 mL/kg进行对应灌胃处理,连续7 d。进行戊巴比妥钠翻正实验和旷场实验。HE染色观察海马结构,免疫组化观察海马B淋巴细胞瘤-2基因(B-cell lymphoma-2,Bcl-2),酶联免疫吸附试验(ELISA)检测肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-1β (interleukin-1β,IL-1β)、IL-2、IL-6和Orexin含量,RT-PCR法检测OX1R和OX2R mRNA表达;Western blot法检测OX1R和OX2R表达。结果 戊巴比妥钠翻正实验显示失眠模型大鼠睡眠潜伏期显著延长、睡眠持续时间显著缩短(P均<0.01);旷场实验中模型大鼠总活动距离减少、进入中央格距离和修饰次数增加(P均<0.01)。HE染色显示,M组海马CA1区神经元形态和结构损伤比较严重,BX-YYR各剂量组则相对减轻。免疫组化显示,与M组比较,ZH组大鼠海马Bcl-2阳性信号面积占比升高(P<0.01)。ELISA显示,与M组比较,D组和ZL、ZM、ZH组大鼠血清TNF-α、IL-1β、IL-2和IL-6显著降低(P均<0.05),D组和ZM、ZH组大鼠血清和海马Orexin水平显著降低(P均<0.01)。RT-PCR显示,与M组比较,ZL、ZM、ZH组海马OX1R mRNA表达降低(P均<0.05)。Western blot检测显示,与M组比较,ZL、ZM、ZH组大鼠海马OX1R表达降低(P均<0.01),D组和ZL组大鼠海马OX2R表达升高(P均<0.05)。结论 BX-YYR能够改善PCPA失眠模型大鼠的睡眠情况,同时调控血清TNF-α、IL-1β、IL-2、IL-6水平,其作用机制可能与调节海马Orexin、Background The incidence of insomnia is increasing, which affects human physical and mental health. The main active ingredients of Pinellia Ternata and Coix Seed(BX-YYR) include baicalein, β-sitosterol, dousterol, etc., but its specific mechanism of action is still unclear. Objective To investigate the regulatory effects of BX-YYR on Orexin and its receptors and apoptosisrelated factors in 4-Chloro-DL-phenylalanine(PCPA) insomnia model rats. Methods PCPA was applied to induce insomnia rat models, the experimental rats were divided into normal group(N), model group(M), diazepam group(D), low dose of BX-YYR(ZL), medium dose of BX-YYR(ZM), and high dose of BX-YYR(ZH) group, with 15 rats in each group. Each group was given corresponding intragastric administration at 10 mL/kg body weight for 7 consecutive days. Pentobarbital sodium correction and open field experiments were performed. HE staining was used to observe the hippocampus structure, immunohistochemistry for the hippocampal B-cell lymphoma-2 gene(Bcl-2), ELISA was used to detect the serum tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β), IL-2, IL-6, and Orexin, RT-PCR for the expression of OX1R and OX2R mRNA, and Western blotting for the protein expression of OX1R and OX2R. Results The pentobarbital sodium correction experiment significantly prolonged sleep latency and shortened sleep duration in insomnia model rats(all P<0.01). In the open field experiment, the total activity distance of the model rats decreased, the distance into the central grid and the number of modifications increased(all P<0.01). HE staining showed that the neuron morphology and structure of hippocampus CA1 area in M group were severely damaged, while the damage in Chinese medicine dose groups were relatively reduced. Immunohistochemistry showed that compared with M group, the proportion of the hippocampal Bcl2 positive signal area in ZH group increased(P<0.01). ELISA showed that compared with M group, the serum TNF-α, IL-1β, IL-2 and IL-6 in the D, ZL, ZM, ZH groups were s

关 键 词：半夏 薏苡仁 失眠动物模型 对氯苯丙氨酸 海马 食欲素 细胞因子 细胞凋亡 

分 类 号：R256[医药卫生—中医内科学]