周围神经缺血后相关神经元凋亡及神经生长因子动态变化的研究

Study on the apoptosis of related neuron and the changes of nerve growth factor after peripheral nerve ischemia in rats

目的探讨大鼠坐骨神经缺血后相关脊髓节段和背根节神经元的凋亡及神经生长因子(NGF)表达的变化,以期为周围神经缺血性神经病的机制及临床治疗提供理论依据。方法结扎大鼠单侧髂总动脉制作坐骨神经缺血模型,分别于术后3、5、7、14、21、28d用4%多聚甲醛灌流固定,取腰髓L4～6节段和双侧第4～6腰脊神经节固定,分别进行免疫组织化学及TUNEL染色检测神经元的NGF和凋亡表达,同体对照。结果坐骨神经缺血的不同时间段L4～6脊髓前角运动神经元及背根节神经元可见凋亡神经元细胞,背根节神经元凋亡率术后3d时显著性升高,术后7d时达到峰值后缓慢下降,后期接近对照侧水平。腰髓前角运动神经元在术后5d时开始有显著性提高,14d时达到最大值后逐渐下降,后期较对照侧仍显著性增高。腰髓前角运动神经元及背根节神经元NGF的表达5d时阳性率较对照侧开始显著降低,7d时达到最低点后逐渐复原,至术后28d时与对照侧无明显差异。结论周围神经缺血性损伤导致相关神经元胞体中NGF含量减少,与神经元凋亡相关。缺血性损伤对背根节感觉神经元的影响较脊髓运动神经明显。

Objective To observe the apoptosis of related neuron and the changes of expression of nerve growth factor （NGF） in lumbar segment and dorsal root ganglion （DRG） neurons after peripheral nerve ischemia in rats, and to investigate the relationship between neuron apoptosis and neurotrophic factors （NTFs） in order to provide theoretical basis for the study on peripheral neuropathy in basic medicine and clinical medicine. Methods The animal models of peripheral nerve ischemia in Wistar rats were made by ligating experimental side common iliac artery. The rats were sacrificed on the 3rd, 5th, 7th, 14th, 21st and 28th day after operation respectively and were perfused with 4% paraformaldegyde. The forth to sixth lumbar spinal cord segment and the forth to sixth lumbar DRG were harvested and fixed with 4% paraformaldehyde ,then were embedded in paraffin and sectioned. TUNEL and immunohetochanity technique were used to detect the expression of NGF and apoptosis of neurons. Results Apoptosis of neurons was found in the forth to sixth lumbar segment and the dorsal root ganglion neurons at different time points after operation. The expression of apoptosis in dorsal root ganglion neurons of experimental side began to increase after 3 days and reached the peak on the 7th day and then recovered gradually and reached normal criteria after 4 weeks. The expression in spinal cord of experimental side began to increase on the 5th day, reached the peak on the 14th day. The positive rate decreased with the time going on, but the expression of apoptosis was still obviously higher than that of control side after 4 weeks. The expression of NGF on the experimental side was obviously lower than that of control side on the 5th day and peaked on the 7th day. After that the positive rate decreased gradually with time going on. The difference was not significant at the 28th day. Conclusion In the progress of peripheral nerve ischemia, the expression of NGF in Lumbar spinal cord motoneurons and dorsal root ganglion neurons decrease, which is related with neuronal apoptosis. The effect of ischemia injury on DRG neurons is more obvious than that on motoneurons in spinal cord.