中国组织工程研究

中国组织工程研究 ›› 2022, Vol. 26 ›› Issue (35): 5620-5625.doi: 10.12307/2022.949

• 组织构建实验造模 experimental modeling in tissue construction • 上一篇    下一篇

电针水沟、百会穴对脑缺血再灌注损伤大鼠大脑皮质神经元凋亡的影响

刘丹妮1，2，3，孙光华1，2，3，周桂娟1，2，3，刘红雅1，2，3，周  君1，2，3，4，谭金曲1，2，3，黄夏荣1，2，3，
彭  婷1，2，3，封蔚彬1，2，3， 罗  敷1，2，3
  

  1. 南华大学衡阳医学院附属第一医院，1康复医学中心，2康复医学科，3康复医学实验室，湖南省衡阳市   421001；4四川大学华西医院康复医学中心，四川省成都市   610041
  • 收稿日期:2021-10-08 接受日期:2021-12-28 出版日期:2022-12-18 发布日期:2022-05-17
  • 通讯作者: 罗敷，硕士，医师，南华大学衡阳医学院附属第一医院，1康复医学中心，2康复医学科，3康复医学实验室，湖南省衡阳市 421001
  • 作者简介:刘丹妮，女，1996年生，湖南省衡南县人，南华大学在读硕士，主要从事脑缺血再灌注损伤的基础与临床研究。
  • 基金资助:
    湖南省自然科学基金项目(2020JJ5508)，项目负责人：孙光华；湖南省自然科学基金项目(2020JJ5512)，项目负责人：周桂娟；湖南省卫健委项目(20200474)，项目负责人：刘红雅

Effect of electroacupuncture at Shuigou and Baihui acupoints on neuronal apoptosis in cerebral cortex of rats with cerebral ischemia-reperfusion injury

Liu Danni1, 2, 3, Sun Guanghua1, 2, 3, Zhou Guijuan1, 2, 3, Liu Hongya1, 2, 3, Zhou Jun1, 2, 3, 4, Tan Jinqu1, 2, 3, Huang Xiarong1, 2, 3, Peng Ting1, 2, 3, Feng Weibin1,2,3, Luo Fu1, 2, 3   

  1. 1Rehabilitation Medicine Center, 2Department of Rehabilitation, 3Rehabilitation Laboratory, The First Affiliated Hospital of Hengyang Medical School, University of South China, Hengyang 421001, Hunan Province, China; 4Rehabilitation Medicine Center, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, China
  • Received:2021-10-08 Accepted:2021-12-28 Online:2022-12-18 Published:2022-05-17
  • Contact: Luo Fu, Master, Physician, Rehabilitation Medicine Center, The First Affiliated Hospital of Hengyang Medical School, University of South China, Hengyang 421001, Hunan Province, China; Department of Rehabilitation, The First Affiliated Hospital of Hengyang Medical School, University of South China, Hengyang 421001, Hunan Province, China; Rehabilitation Laboratory, The First Affiliated Hospital of Hengyang Medical School, University of South China, Hengyang 421001, Hunan Province, China
  • About author:Liu Danni, Master candidate, Rehabilitation Medicine Center, The First Affiliated Hospital of Hengyang Medical School, University of South China, Hengyang 421001, Hunan Province, China; Department of Rehabilitation, The First Affiliated Hospital of Hengyang Medical School, University of South China, Hengyang 421001, Hunan Province, China; Rehabilitation Laboratory, The First Affiliated Hospital of Hengyang Medical School, University of South China, Hengyang 421001, Hunan Province, China
  • Supported by:
    the Natural Science Foundation of Hunan Province, Nos. 2020JJ5508 (to SGH) and 2020JJ5512 (to ZGJ); Hunan Provincial Health Commission Project, No. 20200474 (to LHY)

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摘要：

文题释义：
脑缺血再灌注损伤：脑缺血半暗带区血流为低灌注状态，为挽救濒死的脑组织，提高患者生存概率，应尽快恢复缺血脑组织血流。当血流恢复时，可能会进一步加重再灌注脑组织损伤，导致患者功能障碍加剧，此现象称为脑缺血再灌注损伤。
细胞凋亡：是一种程序性细胞死亡，有维持器官内环境稳定的作用。细胞凋亡可以存在于正常细胞更新、免疫系统的正常发育中。过度的细胞凋亡影响多种疾病的发生发展，包括缺血性损伤疾病、神经退行性疾病、自身免疫性疾病等。目前可以通过检测凋亡相关因子半胱天冬氨酸蛋白酶3、半胱天冬氨酸蛋白酶8等，来研究细胞凋亡的水平。

背景：脑缺血再灌注损伤常见于缺血性脑卒中，严重影响患者预后，因此探索有效缓解脑缺血再灌注损伤的治疗方法十分重要。电针可有效改善缺血性脑卒中神经功能的缺损症状。
目的：探讨电针治疗对脑缺血再灌注损伤大鼠的神经保护作用及其对大脑皮质神经元凋亡的影响。
方法：将48只雄性3月龄的SD大鼠随机分为假手术组、模型组、电针组，每组16只。模型组及电针组所有大鼠进行左侧大脑中动脉脑缺血再灌注造模，缺血2 h，再灌注6 h；假手术组大鼠仅暴露并游离颈动脉。然后对电针组大鼠进行“水沟”“百会”穴的电针干预，选择疏密波，频率3 Hz/15 Hz，强度1 mA，干预20 min，1次/d，连续5 d。模型制作后第5天，使用Longa神经功能评分对所有纳入实验的大鼠进行神经功能受损程度评定；TTC染色观察脑梗死体积；ELISA法检测炎症因子水平；TUNEL法检测细胞凋亡情况；荧光定量PCR及Western blot分别检测大脑皮质区半胱天冬氨酸蛋白酶3(Caspase-3)、半胱天冬氨酸蛋白酶8(Caspase-8)mRNA及蛋白的表达水平。
结果与结论：①与假手术组相比，模型组大鼠神经功能评分明显升高(P < 0.01)；脑梗死体积明显增大(P < 0.01)；左侧大脑皮质区TUNEL阳性细胞数量明显增多(P < 0.01)；血清炎症因子白细胞介素1β、白细胞介素18、肿瘤坏死因子α表达水平升高(P < 0.05，P < 0.01，P < 0.01)；大脑皮质区Caspase-3、Caspase-8 mRNA及蛋白表达水平明显升高(P < 0.01，P < 0.01)；②与模型组相比，电针组大鼠神经功能评分降低(P < 0.05)；脑梗死体积减小(P < 0.05)；左侧大脑皮质区TUNEL阳性细胞数量明显减少(P < 0.01)；血清炎症因子白细胞介素1β、白细胞介素18、肿瘤坏死因子α表达水平均降低(P < 0.01，P < 0.05，P < 0.05)；大脑皮质区Caspase-3、Caspase-8 mRNA及蛋白表水平明显降低(P < 0.01，P < 0.01)；③提示电针治疗可能通过抑制大鼠大脑皮质神经元凋亡，缓解脑缺血再灌注损伤，发挥神经保护作用。

https://orcid.org/0000-0002-7999-716X (刘丹妮)

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

关键词: 电针, 脑缺血再灌注损伤, 大脑皮质, 凋亡, 炎症, 神经功能, 大鼠

Abstract: BACKGROUND: Cerebral ischemia-reperfusion injury is commonly seen in patients with ischemic stroke and seriously affects their prognosis, so it is important to explore effective treatments to alleviate ischemia-reperfusion injury. Electroacupuncture can effectively improve the symptoms of neurological deficits in ischemic stroke. 
OBJECTIVE: To investigate the neuroprotective effect of electroacupuncture treatment on cerebral ischemia-reperfusion injury rats and its effect on neuronal apoptosis in the cerebral cortex. 
METHODS: A total of 48 male 3-month-old Sprague-Dawley rats were randomly divided into sham-operated group, model group, and electroacupuncture group, with 16 rats in each group. Cerebral ischemia-reperfusion models were established on the left middle cerebral artery of the rats from the model and electroacupuncture groups: ischemia for 2 hours and reperfusion for 6 hours. In the sham-operated group, only the carotid artery was exposed and dissociated. Then, rats in the electroacupuncture group were subjected to electroacupuncture at Baihui and Shuigou acupoints with sparse and dense waves at 3 Hz/15 Hz, 
1 mA, for 20 minutes, once a day for 5 continuous days. On the 5th day after modeling, all rats were assessed for neurological impairment using the Longa neurological severity score. triphenyltetrazolium chloride staining was used to determine the infarct volume. Enzyme linked immunosorbent assay was used to detect the level of inflammatory factors. TUNEL was used to detect apoptosis. RT-PCR and western blot were used to detect the expression of Caspase-3 and Caspase-8 at mRNA and protein levels, respectively. 
RESULTS AND CONCLUSION: Compared with the sham-operated group, in the model group, the neurological severity scores of rats were significantly higher (P < 0.01); the brain infarct volume of rats was significantly higher (P < 0.01); the number of TUNEL-positive cells in the left cerebral cortex of rats was significantly increased (P < 0.01); the levels of serum inflammatory factors interleukin-1β, interleukin-18 and tumor necrosis factor-α were increased (P < 0.05, P < 0.01, P < 0.01); mRNA and protein expressions of Caspase-3 and Caspase-8 in the cerebral cortex of rats were significantly higher (P < 0.01, P < 0.01). Compared with the model group, in the electroacupuncture group, the neurological severity scores of rats were reduced (P < 0.05); the volume of cerebral infarction in rats was reduced (P < 0.05); the number of TUNEL-positive cells in the left cerebral cortex area of rats was significantly reduced (P < 0.01); the levels of serum interleukin-1β, interleukin-18 and tumor necrosis factor-α were reduced (P < 0.01, P < 0.05, P < 0.05); mRNA and protein expressions of Caspase-3 and Caspase-8 in the cerebral cortex area of rats were significantly reduced (P < 0.01, P < 0.01). To conclude, electroacupuncture treatment may exert neuroprotective effects by inhibiting neuronal apoptosis in the rat cerebral cortex and alleviating cerebral ischemia-reperfusion injury.

Key words: electroacupuncture, cerebral ischemia-reperfusion injury, cerebral cortex, apoptosis, inflammation, neurological function, rat

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