Lan Wen-Bin, Lin Jian-Hua, Chen Xuan-Wei, Wu Chao-Yang, Zhong Guang-Xian, Zhang Li-Qun, Lin Wen-Ping, Liu Wei-Nan, Li Xiang, Lin Jin-Luan
The First Clinical Medical College of Fujian Medical University, Fuzhou, Fujian, China.
The First Clinical Medical College of Fujian Medical University, Fuzhou, Fujian, China.
Brain Res. 2014 May 8;1562:100-8. doi: 10.1016/j.brainres.2014.03.020. Epub 2014 Mar 24.
The current study was performed to evaluate the mechanisms and therapeutic effects of overexpressing neuroglobin (Ngb) on spinal cord injury (SCI). Adeno-associated virus (AAV) was injected in the T12 section 7 days before SCI. Animals were randomly divided into four groups: a sham group, a vehicle group, an AAV-EGFP group and an AAV-Ngb group. Recovery of hind limb locomotor function was determined during the 3-week post operation period by the Basso, Beattie and Bresnahan locomotor rating scale. At 24 h after SCI and at the end of the study, the segments of spinal cord, centered with the lesion site were harvested for histopathological analysis. Immunofluorescence was performed using antibodies to recognize neuN in the lesion sections. At 24 h after SCI, the spinal cord tissue samples were removed to analyze tissue concentrations of superoxide dismutase (SOD) and malondialdehyde (MDA). Apoptotic cells were assessed using a terminal deoxynucleotidyl transferase, dUTP nick end labeling (TUNEL) kit. The expression of bcl-2, bax, cytochrome c, and cleaved caspase-3, were determined by Western blot assay and immunostaining analysis. The results showed that animals overexpressing Ngb had significantly greater recovery of locomotor function, less neuronal loss and fewer apoptotic cells. In addition, overexpressing Ngb significantly increased bcl-2 expression and SOD level, decreased bax expression, attenuated the release of cytochrome c from mitochondria to the cytosol fraction, and reduced the activity of caspase-3 and MDA level after SCI. These findings suggest, that overexpressing Ngb can significantly improve the recovery of locomotor function. This neuroprotective effect may be associated with the inhibition of neural apoptosis via the mitochondrial pathway.
本研究旨在评估过表达神经球蛋白（Ngb）对脊髓损伤（SCI）的作用机制及治疗效果。在脊髓损伤前7天，将腺相关病毒（AAV）注射到T12节段。动物被随机分为四组：假手术组、载体组、AAV-EGFP组和AAV-Ngb组。在术后3周内，通过Basso、Beattie和Bresnahan运动评分量表测定后肢运动功能的恢复情况。在脊髓损伤后24小时和研究结束时，采集以损伤部位为中心的脊髓节段进行组织病理学分析。在损伤切片中使用抗神经核蛋白（neuN）抗体进行免疫荧光检测。在脊髓损伤后24小时，取出脊髓组织样本分析超氧化物歧化酶（SOD）和丙二醛（MDA）的组织浓度。使用末端脱氧核苷酸转移酶dUTP缺口末端标记（TUNEL）试剂盒评估凋亡细胞。通过蛋白质免疫印迹法和免疫染色分析测定bcl-2、bax、细胞色素c和裂解的半胱天冬酶-3的表达。结果表明，过表达Ngb的动物运动功能恢复明显更好，神经元损失更少，凋亡细胞也更少。此外，过表达Ngb可显著增加bcl-2表达和SOD水平，降低bax表达，减弱细胞色素c从线粒体向细胞质部分的释放，并降低脊髓损伤后半胱天冬酶-3的活性和MDA水平。这些发现表明，过表达Ngb可显著改善运动功能的恢复。这种神经保护作用可能与通过线粒体途径抑制神经细胞凋亡有关。
