Effects of Electroacupuncture on Expression of Autophagy Related Proteins LC3-Ⅱ and Beclin1 in Rats after Cerebral Ischemia-Reperfusion
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摘要: 目的 观察电针对脑缺血再灌注损伤大鼠神经功能的改善程度及自噬相关蛋白LC3-Ⅱ、Beclin1表达的影响。方法 根据随机数字表法，将72只雄性SD大鼠随机分为对照组、模型组和电针组，每组24只。采用Longa线栓法建立改良的急性局灶性脑缺血再灌注大鼠模型，并在缺血1 h后进行再灌注。参照改良的Zea Longa 8级神经功能缺损评分法对3组大鼠脑神经功能损伤程度进行评估。电针组于造模成功后5 min和16 h进行电针干预。于再灌注后24 h取材。TTC染色法观察各组大鼠脑梗死体积；HE染色法观察大鼠脑组织病理学变化；Western blot检测大鼠右侧纹状体脑组织中LC3-Ⅱ、Beclin1蛋白表达，qPCR检测Beclin1 mRNA表达。结果 对照组无行为学改变，模型组神经功能缺损评分显著高于对照组(P < 0.05)；与模型组比较，电针组神经功能缺损评分明显低于模型组(P < 0.05)。电针组脑梗死体积较模型组明显减少(P < 0.05)。与模型组比较，电针组LC3-Ⅱ、Beclin1蛋白表达明显增加(P < 0.05)，Beclin1 mRNA的表达亦明显增加(P < 0.05)。结论 电针治疗可能通过促进自噬相关蛋白LC3-Ⅱ、Beclin1的表达，调控自噬的发生，减小大鼠脑梗死体积，改善神经功能缺损，实现对脑缺血再灌注损伤大鼠的保护效应。Abstract: OBJECTIVE To observe the improvement degree of nerve function and the effect of electroacupuncture on autophagy related proteins LC3-Ⅱ and Beclin1 expression in rats with cerebral ischemia-reperfusion injury.METHODS According to the random number table method, 72 male SD rats were randomly divided into control group, model group and electroacupuncture group, with 24 rats in each group. A modified rat model of acute focal cerebral ischemia reperfusion was established by referring to Longa line bolt method, and reperfusion was performed 1 h after ischemia. The degrees of cerebral neurological damage of the three groups of rats were evaluated by using the improved Zea Longa grade 8 neurological impairment score. Electroacupuncture intervention was performed at 5 min and 16 h after successful modeling. Materials were harvested 24 h after reperfusion. The cerebral infarction volumes of rats in each group were observed by TTC staining. The histopathological changes of rat brain were observed by HE staining. The expression of LC3-Ⅱ and Beclin1 protein in the right striatum of rats brain tissue were tested by Western blot assay, and the expression of Beclin1 mRNA was detected by qPCR.RESULTS No behavioral change was observed in the control group, and the neurological function score of the model group was significantly higher than that of the control group (P < 0.05). Compared with the model group, the score of nerve function defect in the electroacupuncture group was significantly lower than that in the model group (P < 0.05). The volume of cerebral infarction in the electroacupuncture group was significantly lower than that in the model group (P < 0.05). Compared with the model group, the expression of LC3-Ⅱ and Beclin1 protein in the electroacupuncture group significantly increased (P < 0.05), and the expression of Beclin1 mRNA significantly increased (P < 0.05).CONCLUSION Electroacupuncture therapy may regulate the occurrence of autophagy by promoting the expression of autophagy-related proteins LC3-Ⅱ and Beclin1, reduce the volume of cerebellar infarction, improve the neurological function defect, and achieve the protective effect on rats with cerebral ischemia-reperfusion injury.
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Key words:
- electroacupuncture /
- cerebral ischemia-reperfusion /
- autophagy /
- LC3-Ⅱ /
- Beclin1
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图 5 3组大鼠右侧脑组织中LC3-Ⅱ、Beclin1蛋白表达量
注：与对照组比较，*P < 0.05，**P < 0.01；与模型组比较，#P < 0.05。x ±s，n=6。
表 1 qPCR引物序列
名称 长度/bp 序列(5'→3') Beclin1 143 F: CGAGTGTCAGAACTACAAACGCT R: CTCCTCCTCCAAGGCCAACT GAPDH 121 F: GGCAAGTTCAACGGCACAG R: CGCCAGTAGACTCCACGACAT 
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