夹脊电针通过miR-128-3p/ATG12轴干预大鼠脊髓损伤后神经功能的机制研究  

Mechanism study of electroacupuncture at Jiaji(EX-B2)intervening in neurological function after spinal cord injury via miR-128-3p/ATG12 axis in rats

作　　者：吴海波 钟前波 邓寒冰 冯伦冬 WU Haibo;ZHONG Qianbo;DENG Hanbing;FENG Lundong(The Third People’s Hospital of Hainan Province,Haikou 571100,China)

机构地区：[1]海口市第三人民医院,海口571100

出　　处：《上海针灸杂志》2024年第8期898-905,共8页Shanghai Journal of Acupuncture and Moxibustion

摘　　要：目的分析夹脊电针对大鼠脊髓损伤后神经功能的影响,并探讨其可能机制。方法将40只Wistar大鼠随机分为假手术组、模型组、夹脊电针组以及夹脊电针+anta-miR-128-3p组,每组10只。假手术组与模型组不做处理,夹脊电针组给予电针干预(每日1次,每次20 min,连续2周),夹脊电针+anta-miR-128-3p组在末次电针刺激后,尾静脉注射anta-miR-128-3p。干预结束后通过行为学(Basso Beattie Bresnahan,BBB)评分评估大鼠运动功能;苏木素-伊红染色与原位末端标记法染色观察大鼠脊髓病理改变以及凋亡情况;免疫荧光法测定脊髓组织神经元特异性核蛋白(neuron-specific nuclear protein,NeuN)阳性表达;qRT-PCR法检测脊髓组织miR-128-3p、自噬相关蛋白12(autophagy-related protein 12,ATG12)mRNA水平;Western blot法检测ATG12蛋白表达;双荧光素酶报告基因验证miR-128-3p与ATG12关系。结果假手术组脊髓结构完整,排列整齐;模型组大鼠脊髓结构紊乱,神经元数量减少,存在大量炎性浸润;夹脊电针组脊髓结构紊乱现象有所减轻,炎性浸润减少;与夹脊电针组比较,夹脊电针+anta-miR-128-3p组脊髓损伤加剧;与假手术组比较,模型组大鼠BBB评分、miR-128-3p水平、NeuN阳性神经元数量降低(P<0.05),凋亡率、ATG12 mRNA水平与ATG12蛋白表达升高(P<0.05);夹脊电针组大鼠BBB评分、miR-128-3p水平、NeuN阳性神经元数量高于模型组(P<0.05),凋亡率、ATG12 mRNA水平与ATG12蛋白表达低于模型组(P<0.05);与夹脊电针组比较,夹脊电针+anta-miR-128-3p组大鼠BBB评分、miR-128-3p水平、NeuN阳性神经元数量降低(P<0.05),凋亡率、ATG12 mRNA水平与ATG12蛋白表达升高(P<0.05);miR-128-3p与ATG12靶向结合。结论夹脊电针可能通过调控miR-128-3p/ATG12轴,改善脊髓损伤大鼠神经功能损伤。Objective To observe the effect of electroacupuncture at Jiaji(EX-B2)on neurological function after spinal cord injury in rats and explore the plausible mechanism.Method Forty Wistar rats were randomized into a sham operation group,a model group,a Jiaji electroacupuncture group,and a Jiaji electroacupuncture+anta-miR-128-3p group,with 10 rats in each group.The sham operation and model groups did not receive any intervention.The Jiaji electroacupuncture group received electroacupuncture intervention(once daily,20 min each time for 2 consecutive weeks).The Jiaji electroacupuncture+anta-miR-128-3p group received tail vein injection of anta-miR-128-3p following the final electroacupuncture stimulation.When the intervention finished,the behavioral score[Basso Beattie Bresnahan(BBB)]was used to assess the rat’s motor function;hematoxylin-eosin(HE)staining and TdT-mediated dUTP nick end labeling(TUNEL)staining were used to observe the pathological changes and apoptosis in the spinal cord of rats;the positive expression of NeuN in the spinal cord tissues was detected using the immunofluorescence assay;the levels of miR-128-3p and autophagy-related protein 12(ATG12)mRNA in the spinal cord tissues were detected using the quantitative real-time polymerase chain reaction(qRT-PCR);the expression of ATG12 protein was detected using Western blot;the relationship between miR-128-3p and ATG12 was verified by dual luciferase reporter gene assay.Result The spinal cord had a complete structure with neat arrangements in the sham operation group;in the model group,the structure of the spinal cord was disordered,and the number of neurons decreased with massive inflammatory infiltration;the spinal cord structural disorder improved with less inflammatory infiltration in the Jiaji electroacupuncture group;compared to the Jiaji electroacupuncture group,spinal cord damage worsened in the Jiaji electroacupuncture+anta-miR-128-3p group.Compared to the sham operation group,the BBB score,the level of miR-128-3p,and the number of NeuN positive

关 键 词：穴 夹脊 电针 脊髓损伤 神经功能 miR-128-3p 自噬相关蛋白12 大鼠 

分 类 号：R2-03[医药卫生—中医学]