Objectives: To observe the effect of electroacupuncture (EA) at "Dazhui" (GV14) and "Jizhong"(GV6) of the Governor Vessel (GV) on mitochondrial fusion and neural stem cell (NSC) proliferation and differentiation in the spinal cord of rats with spinal cord injury (SCI), so as to investigate its mechanisms underlying improvement of SCI.
Methods: SD rats were randomly divided into sham operation, model and EA groups, with 15 rats in each group. The SCI model was established by using a precision impactor. EA (20 Hz/100 Hz, 1-2 mA) was applied to GV14 and GV6 for 30 min, once daily for 14 days. The rats' hindlimb locomotor function in each group was assessed using the Basso-Beattie-Bresnahan (BBB) locomotor scale. Histopathological changes of the injured spinal cord tissue and the number of neurons were evaluated after H.E. staining and Nissl staining. The expressions of Nestin, mitochondrial fusion-related protein optic atrophy-1 (OPA1) and NSC markers sex-determining region Y-box 2 (SOX2) in the injured spinal cord tissue were detected by immunofluorescence staining. The protein and mRNA expression levels of Nestin in the spinal cord tissue were detected by quantitative real-time PCR and Western blot, separately.
Results: Compared with the sham operation group, the BBB scores after modeling, and the number of neurons were significantly decreased (P<0.001), while the mean fluorescence intensity values of Nestin, SOX2 and OPA1, and the expressions of Nestin mRNA and protein considerably increased (P<0.001, P<0.01, P<0.05) in the model group. After EA intervention and in comparison with the model group, the BBB scores at the 7th and 14th day, the number of neurons, the mean fluorescence intensity values of Nestin, SOX2 and OPA1, and the expressions of Nestin mRNA and protein were strikingly increased (P<0.05, P<0.01, P<0.001) in the EA group. H.E. staining showed swollen, ruptured and necrotic neurons of the spinal cord, with a large number of vacuoles and severe inflammatory cell infiltration after modeling, which was relatively milder in the EA group.
Conclusions: EA stimulation of GV14 and GV6 can promote the recovery of motor function in rats with SCI, which may be related to its effects in promoting mitochondrial fusion and enhancing the proliferation and differentiation of NSCs.
目的: 观察电针督脉对脊髓损伤（SCI）大鼠损伤区脊髓组织中线粒体融合及神经干细胞增殖分化的影响，探讨电针促进SCI神经修复的机制。方法: SD大鼠随机分为假手术组、模型组和电针组，每组各15只。采用精密打击器打击法构建胸10节段SCI模型。电针组大鼠予以电针“大椎”“脊中”，30 min/次，1次/d，共14 d。用Basso-Beattie-Bresnahan （BBB）运动功能评分法评估大鼠后肢的运动功能；HE染色法、尼氏染色法观察大鼠损伤区脊髓组织病理变化及神经元数量；免疫荧光染色法检测大鼠损伤区脊髓组织中神经干细胞增殖标记物巢蛋白（Nestin）、线粒体融合相关蛋白视神经萎缩蛋白1（OPA1）及神经干细胞转录因子性别决定区Y框蛋白2（SOX2）的阳性表达；实时荧光定量PCR法、Western blot法检测大鼠脊髓组织中Nestin mRNA及蛋白的表达。结果: 与同时点假手术组比较，模型组大鼠造模后BBB评分降低（P<0.001）；脊髓组织中神经元肿胀、破裂、坏死严重，尼氏小体溶解严重，神经元数量减少（P<0.001）；Nestin、OPA1、SOX2阳性表达及Nestin蛋白和mRNA相对表达量均升高（P<0.001，P<0.05，P<0.01）。与模型组比较，电针干预第7、14天时电针组大鼠BBB评分，Nestin、OPA1、SOX2阳性表达，以及Nestin蛋白和mRNA相对表达量均升高（P<0.05，P<0.01，P<0.001），脊髓神经元受损程度减轻，尼氏小体溶解程度减轻，神经元数量增加（P<0.05）。结论: 电针督脉可改善SCI大鼠运动功能，促进SCI区域神经元修复，其机制可能与电针督脉促进损伤区脊髓组织线粒体融合蛋白OPA1表达，增强神经干细胞增殖分化有关。.
Keywords: Electroacupuncture; Governor Vessel; Mitochondrial fusion; Neural stem cells; Proliferation and differentiation; Spinal cord injury.