目的:探讨氧调控性神经生长因子（nerve growth factor，NGF）基因修饰神经干细胞（neural stem cells，NSCs）移植治疗急性脊髓损伤（spinal cord injury，SCI）的可行性并观察脊髓损伤后的功能修复情况。方法:以腺相关病毒（adeno-associated virus，AAV）为载体构建基因修饰神经干细胞，制备SCI动物模型3 d后将氧调控性神经生长因子基因修饰的神经干细胞移植到脊髓损伤部位作为AAV-5HRE-NGF-NSCs组（NGF组）；另设GFP修饰的神经干细胞组（AAV-5HRE-GFP-NSCs组，GFP组）；假手术组（空白组）；SCI组（对照组）。在移植后第1、3、7、10、14、21、28、35、42、60 d共10个时间点采用Basso-Beattie-Bresnahan（BBB）运动功能评分，斜板试验和脚步印迹检测大鼠后肢运动功能的恢复情况。通过盒式磁带录像（video cassette recorder，VCR）图像及定量测定大鼠离地高度，错误脚步及后肢轮替动作检验大鼠的后肢支持力及灵活度。通过观察脊髓直观图粗测大鼠脊髓损伤程度。通过尼氏染色、HE染色和免疫荧光方法评价脊髓损伤区的神经元修复及形态学变化情况。通过CM-DiI追踪移植神经干细胞并用免疫荧光的方法分析干细胞的分化情况。结果:基因修饰神经干细胞移植60 d后，NGF组大鼠的BBB，斜板测试和脚步印迹试验的功能评分均高于SCI组和GFP组，差异有统计学意义（n P<0.05）；通过VCR图像分析，NGF组大鼠的后肢支持力与活动灵活度优于SCI组和GFP组，差异有统计学意义（n P<0.05）；通过脊髓直观图分析，各组大鼠脊柱肉眼观对比图示NGF组脊柱未呈现明显萎缩和颜色加深，损伤程度低于SCI组和GFP组；通过尼氏染色、HE染色和免疫荧光方法检测，相比于SCI组与GFP组，NGF组在移植部位NeuN呈明显阳性，同时在形态学水平可见明显再生的神经结构，且SCI空洞面积减小，神经元和尼氏小体增多，差异具有统计学意义（n P<0.05）。通过CM-DiI追踪神经干细胞，用NeuN标记神经元，用GFAP标记星形胶质细胞，发现神经干细胞可以有效分化为神经元和星形胶质细胞，GFP组神经干细胞更多向星形胶质细胞分化，NGF组神经干细胞更多向神经元分化。n 结论:通通过腺相关病毒介导氧调控性NGF基因修饰神经干细胞移植治疗SCI，一方面神经干细胞分化为神经干细胞和星形胶质细胞可以填补损伤空洞；另一方面神经干细胞充当NGF基因治疗的载体，对邻近受损神经细胞发挥保护作用，减少神经元细胞的死亡，这有望为急性脊髓损伤的治疗提供新思路，同时给NGF蛋白药物的研发做出新尝试。“,”Objective:To investigate the feasibility of transplantation of neural stem cells (NSCs) modified by hypoxia-regulated nerve growth factor (NGF) gene to treat acute spinal cord injury (SCI) and observe the functional repair after SCI.Methods:Adeno-associated virus (AAV) was used as the vector to construct gene-modified NSCs. Three days after SCI attack on the animal model, the NSCs modified by hypoxia-regulated NGF were transplanted to the site of SCI as the NGF group. The GFP-modified neural stem cell group (GFP group), sham group, SCI group were set up. Hindlimb motor function was assessed by Basso-Beattie-Bresnahan (BBB) Locomotor Rating Scale, inclined plane tests and footprint analysis at 10 time points on day 1, 3, 7, 10, 14, 21, 28, 35, 42 and 60 after transplantation. The video cassette recorder (VCR) image and quantitative measurement of the height of the rat from the ground, the foot error and plantar steps were used to test the hindlimb support and flexibility of the rats. The degree of spinal cord injury in rats was roughly measured by observing the visual map of the spinal cord. The neuronal repair and morphological changes in SCI area were evaluated by Nissl staining, HE staining and immunofluorescence. CM-DiI was used to trace neural stem cells and to analyze the differentiation of NSCs by immunofluorescence.Results:Two months after transplantation of genetically modified NSCs, the BBB, inclined plane tests and footprint Analytical scores of NGF group rats were higher than those of SCI group and GFP group (n P<0.05); Through VCR image analysis, the hindlimb support and mobility of the rats in the NGF group were better than those in the SCI group and GFP group, and the difference was statistically significant (n P<0.05). Visual analysis showed that the spinal cord of the rats in each group was visually compared to the NGF group, and the spine did not show significant atrophy and color deepening, and the degree of injury was lower than that of the SCI group and GFP group; Through Nissl staining, HE staining and immunofluorescence detection, obviously positive in NeuN at the transplant site was noted at NGF group, and evidently regenerated neural structure can be seen at the morphological level. The cavity in SCI was obviously reduced, neurons and Nissl bodies were distinctly increased (n P<0.05). CM-DiI was used to track NSCs, NeuN was used to mark neurons, and GFAP was used to mark astrocytes. It was found that neural stem cells could differentiate into neurons and astrocytes. Neural stem cells in GFP group were more differentiated into astrocytes, and neural stem cells in NGF group were more differentiated into neurons.n Conclusion:NSC transplantation with oxygen-regulated NGF gene mediated by adeno-associated virus can treat SCI, NSCs can differentiate into neural stem cells and astrocytes to fill the damaged cavity, NSCs secrete NGF as the carrier, playing the protective role on adjacent damaged nerve cells and reducing the death of neurons, which is expected to provide new ideas for the treatment of acute spinal cord injury, and at the same time make new attempts for the development of NGF protein drugs.