Department of Neurology, The Affiliated Hospital of Southwest Medical University, Luzhou, 646000, China.
Laboratory of Neurological Diseases and Brain Function, Luzhou, Sichuan, China.
Sci Rep. 2024 Sep 17;14(1):21654. doi: 10.1038/s41598-024-72352-9.
To investigate the spinal cord neuron apoptosis and neuroprotective mechanism of nerve growth factorganismsor (NGF) gene mediated by recombinant adenovirus (Ad-NGF) via peripheral transfection in mice with experimental autoimmune encephalomyelitis (EAE). Forty healthy female C57BL/6 mice were randomly divided into a control group, adenovirus (AdV) group, EAE group, and Ad-NGF transfection group; the control group received no treatment; the AdV group received adenovirus injection via the tail vein; the EAE and Ad-NGF transfection groups were induced with experimental autoimmune encephalomyelitis (EAE) using myelin oligodendrocyte glycoprotein 35-55 (MOG35-55), Ad-NGF transfection group received Ad-NGF injection via the tail vein, and daily neurological impairment scores were obtained. AQThe TUNEL method was employed to observe spinal neuron apoptosis in each group of mice; protein immunoblotting (western blot) and RT-PCR were used to measure NGF levels in the spinal cord tissues of each group, and western blotting was used to assess levels of cleaved caspase-3, Bax, and Bcl-2. ELISA and RT-PCR were employed to detect protein and mRNA levels of neuron-specific enolase (NSE) in spinal cord tissues, respectively. The control group and AdV mice did not develop symptoms. Compared to the EAE group, in the Ad-NGF transfection group, neurological function scores, TUNEL-positive cell counts, the ratio of NeuN + TUNEL to NeuN, levels of Bax and cleaved caspase-3 apoptotic proteins were significantly reduced, while Bcl-2 protein expression was increased. Expression levels of NGF, NGF-mRNA, NSE, and NSE-mRNA in spinal cord tissues were significantly elevated (P < 0.01). Immunofluorescence labeling revealed a significant punctate aggregation of apoptotic cells in spinal neurons of the EAE group, while the aggregation phenomenon was less pronounced in the Ad-NGF transfection group. Ad-NGF transfected by the periphery has a protective effect on spinal cord neurons in EAE mice by up-regulation NGF level, down-regulating apoptotic protein Caspase-3 in spinal cord neurons, inhibiting spinal cord neuron apoptosis and promoting NSE expression.
为了研究神经生长因子（NGF）基因通过重组腺病毒（Ad-NGF）介导的外周转染在实验性自身免疫性脑脊髓炎（EAE）小鼠脊髓神经元凋亡和神经保护机制。将 40 只健康雌性 C57BL/6 小鼠随机分为对照组、腺病毒（AdV）组、EAE 组和 Ad-NGF 转染组；对照组未进行任何处理；AdV 组通过尾静脉注射腺病毒；EAE 和 Ad-NGF 转染组用髓鞘少突胶质细胞糖蛋白 35-55（MOG35-55）诱导实验性自身免疫性脑脊髓炎，Ad-NGF 转染组通过尾静脉注射 Ad-NGF，并每天获得神经功能缺损评分。使用 TUNEL 法观察各组小鼠脊髓神经元凋亡情况；采用蛋白质免疫印迹（western blot）和 RT-PCR 法测量各组小鼠脊髓组织 NGF 水平，western blot 法检测 cleaved caspase-3、Bax 和 Bcl-2 水平。ELISA 和 RT-PCR 法分别检测脊髓组织神经元特异性烯醇化酶（NSE）的蛋白和 mRNA 水平。对照组和 AdV 组的小鼠均未出现症状。与 EAE 组相比，在 Ad-NGF 转染组，神经功能评分、TUNEL 阳性细胞计数、NeuN+TUNEL/NeuN 比值、Bax 和 cleaved caspase-3 凋亡蛋白水平均显著降低，Bcl-2 蛋白表达增加。脊髓组织 NGF、NGF-mRNA、NSE 和 NSE-mRNA 的表达水平均显著升高（P<0.01）。免疫荧光标记显示 EAE 组脊髓神经元中凋亡细胞的点状聚集明显，而 Ad-NGF 转染组的聚集现象较轻。通过外周转染的 Ad-NGF 通过上调 NGF 水平对 EAE 小鼠脊髓神经元具有保护作用，下调脊髓神经元中凋亡蛋白 Caspase-3，抑制脊髓神经元凋亡，促进 NSE 表达。
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