井穴刺络放血调控Nrf2/ARE通路改善脑卒中大鼠认知功能的机制研究*

作者：蓝相洲1，李铃佳2，李叶舟1，薛 珂1，陈 苗1，文 志1，陈 青1

单位：1.湖南中医药大学第一附属医院，湖南 长沙 410007； 2.湖南中医药大学护理学院，湖南 长沙 410208

引用：引用：蓝相洲，李铃佳，李叶舟，薛珂，陈苗，文志，陈青.井穴刺络放血调控Nrf2/ARE通路改善脑卒中大鼠认知功能的机制研究[J].中医药导报,2025,31(9):9-16,31.

DOI：10.13862/j.cn43-1446/r.2025.09.002

PDF： 下载PDF

摘要：

目的：探讨井穴刺络放血通过激活核因子E2相关因子2（Nrf2）/抗氧化反应原件（ARE）信号通路对脑卒中大鼠认知功能及炎症反应的影响。方法：将70只SD大鼠随机分为空白组、假手术组、模型组、井穴刺络放血组、Nrf2抑制剂（ML385）组、井穴刺络放血+ML385组、Nrf2激动剂（Resveratrol）组。空白组不进行处理，假手术组给予假手术进行对照，模型组、井穴刺络放血组、ML385组、井穴刺络放血+ML385组、Resveratrol组均采用大脑中动脉闭塞法（MCAO）制作永久性缺血性脑卒中大鼠模型。术后井穴刺络放血组大鼠予井穴刺络放血治疗3 d（1次/d，左右两侧交替进行）；ML385组大鼠予ML385[30 mg/（kg·d）]腹腔注射7 d；井穴刺络放血+ML385组大鼠予以井穴刺络放血治疗和ML385[30 mg/kg·d）]腹腔注射；Resveratrol组大鼠予Resveratrol[40 mg/（kg·d）]腹腔注射7 d。造模后第1天及取材前1 d使用改良神经功能缺损评分法（mNss）进行神经功能评分；治疗后采用Y迷宫和新异物识别实验评价大鼠的认知功能障碍情况；采用2,3,5-三苯基氯化四氮唑（TTC）染色评估大鼠脑梗死面积；采用酶联免疫吸附试验（ELISA）法检测大鼠血清白细胞介素-6（IL-6）、白细胞介素-1β（IL-1β）、肿瘤坏死因子-α（TNF-α）、丙二醛（MDA）、微量还原型谷胱甘肽（GSH）含量和超氧化物歧化酶（SOD）活力；Real-time PCR检测大鼠海马组织Nrf2 mRNA、血红素氧合酶-1（HO-1） mRNA、NAD（P）H醌脱氢酶1（NQO1） mRNA表达水平；Western blotting检测大鼠海马组织Nrf2、HO-1、NQO1蛋白表达水平。结果：井穴刺络放血组、Resveratrol组大鼠mNss评分均低于模型组（P＜0.05），大鼠自发交替百分比高于模型组（P＜0.01或P＜0.05）；Resveratrol组大鼠认知指数高于模型组（P＜0.05）；井穴刺络放血组、井穴刺络放血+ML385组、Resveratrol组大鼠梗死面积均小于模型组（P＜0.05）；ML385组大鼠血清IL-6、IL-1β、TNF-α含量高于模型组（P＜0.01）；井穴刺络放血组、井穴刺络放血+ML385组和Resveratrol组大鼠血清IL-6、IL-1β、TNF-α含量均低于模型组（P＜0.01）；井穴刺络放血组、Resveratrol组大鼠血清MDA、GSH含量和SOD活力高于模型组（P＜0.01）；ML385组、井穴刺络放血+ML385组大鼠血清MDA、GSH含量和SOD活力均低于模型组（P＜0.01）。HE染色结果显示井穴刺络放血组、井穴刺络放血+ML385组、Resveratrol组大鼠神经元损伤程度减轻，而ML385组大鼠神经元损伤程度加重；井穴刺络放血组、Resveratrol组大鼠海马组织Nrf2 mRNA、HO-1 mRNA、NQO1 mRNA相对表达量及Nrf2、HO-1、NQO1蛋白相对表达量均高于模型组（P＜0.01）。结论：井穴刺络放血能通过激活Nrf2/ARE通路改善脑卒中大鼠认知功能障碍。其作用机制为上调HO-1/NQO1表达以增强抗氧化能力，同时抑制炎症因子释放，从而减轻大鼠的氧化应激与神经炎症，缩小脑梗死面积。

关键词：脑卒中；井穴刺络；认知功能障碍；核因子E2相关因子2/抗氧化反应原件信号通路；大鼠

Abstract：

Objective: To investigate the effect of Jing-well points acupuncture on improving cognitive dysfunction in rats with stroke by activating the Nrf2/ARE signaling pathway and its impact on the inflammatory response. Methods: Totally 70 SD rats were randomly divided into blank group, sham-operation group, model group, Jing-Well points acupuncture group, ML385 (Nrf2 inhibitor) group, Jing-Well points acupuncture+ML385 group, and Resveratrol (Nrf2 agonist) group. The blank group received no treatment, the sham-operation group underwent a sham operation, and the other groups were subjected to permanent ischemic stroke via middle cerebral artery occlusion (MCAO). The Jing-Well points acupuncture group was treated with Jing-Well points acupuncture for 3 days (once daily, alternating sides). The ML385 group received intraperitoneal injections of ML385 [30 mg/(kg·d)] for 7 days. The Jing-Well points acupuncture+ML385 group received Jing-Well points acupuncture and intraperitoneal injections of ML385 [30 mg/(kg·d)]. The Resveratrol group received intraperitoneal injections of Resveratrol [40 mg/(kg·d)] for 7 days. Neurological function was scored using the modified neurological severity score (mNss) on the first day after modeling and the day before tissue collection. Cognitive function was assessed using Y-maze and novel object recognition tests. Brain infarct area was evaluated by 2,3,5-triphenyltetrazolium chloride (TTC) staining. Serum levels of interleukin-6 (IL-6), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), model driven architecture (MDA), glutathione (GSH) and superoxide dismutase (SOD) activity were measured by enzyme-linked immunosorbent assay (ELISA). Real-time PCR was used to detect levels of Nrf2 mRNA, HO-1 mRNA and NQO1 mRNA in the hippocampus, and Western blotting was used to assess protein levels of Nrf2, HO-1, and NQO1. Results: The Jing-Well points acupuncture group and Resveratrol group showed lower mNss than model group (P<0.05), while higher spontaneous alternation percentages than model group (P<0.01 or P<0.05). The Resveratrol group showed higher cognitive index than model group (P<0.05). The Jing-Well points acupuncture group, Jing-Well points acupuncture + ML385 group, and Resveratrol group exhibited smaller brain infarct areas than model group (P<0.05). The ML385 group showed higher serum levels of IL-6, IL-1β and TNF-α than model group (P<0.01); The Jing-Well points acupuncture group, Jing-Well points acupuncture + ML385 group, and Resveratrol group showed lower levels of IL-6, IL-1β and TNF-α than model group (P<0.01). The Jing-Well points acupuncture group and Resveratrol group showed higher serum MDA, GSH levels and SOD activity than model group (P<0.01), while the ML385 group and Jing-Well points acupuncture + ML385 group showed lower serum MDA, GSH levels and SOD activity than model group (P<0.01). HE staining revealed reduced neuronal damage in the Jing-Well points acupuncture group, Jing-Well points acupuncture + ML385 group, and Resveratrol group, while the ML385 group showed aggravated damage. The Jing-Well points acupuncture group and Resveratrol group exhibited higher Nrf2 mRNA, HO-1 mRNA, and NQO1 mRNA and protein levels of Nrf2, HO-1 and NQO1 than model group (P<0.01). Conclusion: Jing-Well points acupuncture can improve cognitive dysfunction in stroke rats by activating the Nrf2/ARE pathway. It upregulates HO-1/NQO1 to enhance antioxidant capacity, suppresses inflammatory factors, alleviates oxidative stress and neuroinflammation, and reduces infarct size.

Key words：stroke; Jing-Well points acupuncture; cognitive impairment; nuclear factor erythroid 2-related factor 2/antioxidant response element signaling pathway; rat

发布时间：2026-01-08

点击量：44