电针原发性高血压模型大鼠太冲、曲池穴观察microRNA-9对其下游靶点P2X7受体、炎性因子、血管紧张素Ⅱ的影响  

Novel mechanism of lowering blood pressure in SHR by electroacupuncting QuChi(LI11)and TaiChong(LR3):MicroRNA9

作　　者：岳炳南 孙娇 李晓璐 周子娴 刘鑫 刘清国[1] YUE Bingnan;SUN Jiao;LI Xiaolu;ZHOU Zixian;LIU Xin;LIU Qingguo(School of Acupuncture-Moxibustion and Tuina,Beijing University of Chinese Medicine,Beijing 100029,China)

机构地区：[1]北京中医药大学针灸推拿学院,100029

出　　处：《环球中医药》2022年第12期2342-2349,共8页Global Traditional Chinese Medicine

基　　金：国家自然科学基金(82074553)。

摘　　要：目的通过观察microRNA-9对其下游靶点P2X7受体、炎性因子、血管紧张素Ⅱ的影响,分析电针降压机制。方法选择原发性高血压模型大鼠(spontaneous hypertension rat,SHR)作为高血压动物模型,将SHR随机分为4组:假手术组、miR9抑制剂+电针组、电针组、模型组,Wistar大鼠作为空白组(共5组,每组10只)。其中,miR9抑制剂+电针组在SHR室旁核内行双极插管植入术后注射microRNA-9抑制剂(每单侧100 nL/天),假手术组、电针组注射等剂量脑脊液。术后5天进行电针干预,穴取双侧太冲、曲池。在电针前和电针后3、7、10、14天测量收缩压,连续电针治疗2周后取材,通过实时荧光定量多聚核苷酸链式反应(real-time quantitative polymerase chain reaction,RT-qPCR)对各组大鼠下丘脑microRNA-9的表达进行定量检测,利用qPCR和蛋白免疫印迹法(western blot,WB)检测各组大鼠P2X7受体的mRNA和蛋白水平,使用酶联免疫吸附法(enzyme linked immunosorbent assay,Elisa)检测下丘脑室旁核和血浆中炎性因子白介素(interleukin,IL)-6、肿瘤坏死因子(tumor necrosis factor,TNF)-α和血管紧张素Ⅱ(angiotensinⅡ,ANGⅡ)的含量。最后使用抗体Iba1进行免疫组化染色,评估下丘脑室旁核内小胶质细胞的活化程度。结果(1)与假手术组相比,电针组降低了高血压大鼠的收缩压(P<0.01),与电针组相比,miRNA9抑制剂+电针组的降压效果下降(P<0.05);(2)电针组与miRNA9抑制剂+电针组、假手术组相比,miRNA9水平明显升高(P<0.01);(3)与miRNA9抑制剂+电针组相比,电针组室旁核、血清中的IL-6含量更低(P<0.01,P<0.01),与miRNA9抑制剂+电针组相比,电针组室旁核、血清中的TNF-α含量更低(P<0.05,P<0.05);(4)与假手术组相比,电针组的ANGⅡ含量下降明显(P<0.01),但电针组与miR-9抑制剂+电针组ANGII的含量相比无统计学差异(P>0.05);(5)与假手术组相比,电针组降低了P2X7受体的蛋白及mRNA表达(P<0.01,P<0.01),与电针组相比,mObjective The research target of this study was to verify P2RX7 as one direct target of miRNA-9,which triggers the release of ANGⅡ and PICs(TNF-α,IL-6)in the PVN.The system of miRNA-9/P2RX7 might be a novel mechanism of lowering blood pressure.Methods Spontaneous Hypertension Rat(SHR)model was selected as the animal model of hypertension,and SHR was randomly divided into 4 groups:the Sham operation group,the miR9 inhibitor+EA group,the EA group,the model group,Wistar rats(WKY,Wistar Kyoto)were used as the blank group(5 groups,10 rats in each group).microRNA-9 inhibitor(100 nL/d per side)was injected after bipolar intubation in the paraventricular nucleus of SHR in the inhibitor+EA group,and the same dose of cerebrospinal fluid was injected in the sham-operation group.Electroacupuncture intervention was performed 5 days after operation,and bilateral Taichong and Quchi points were taken.Systolic blood pressure was measured before and 3,7,10,and 14 days after EA,and samples were collected after continuous EA for 2 weeks.Real-time Quantitative polymerase chain reaction(RT-PCR)was used to quantitatively detect the expression of microRNA-9 in the hypothalamus of rats in each group.The mRNA and protein levels of P2X7 receptor in each group were detected by RT-PCR and Western blotting.The inflammatory factors interleukin(IL)-6 and tumor necrosis factor(TNF)in paraventricular nucleus and plasma of hypothalamus were detected by enzyme-linked immunosorbent assay(Elisa)factor,TNF-αand Angiotensin II(ANG II).Finally,the activation of microglia in paraventricular nucleus of hypothalamus was evaluated by immunohistochemical staining with antibody Iba1.Results Compared with the sham operation group,the systolic blood pressure of hypertensive rats was decreased(P<0.01)in the electroacupuncture group,and hypotensive effect was decreased in the miRNA9 inhibitor+electroacupuncture group compared with the electroacupuncture group(P<0.05);the miRNA9 level in the electroacupuncture group was significantly increased compared with th

关 键 词：微核糖核酸 P2X7受体 电针 原发性高血压 

分 类 号：R245[医药卫生—针灸推拿学]