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目的研究神经干细胞(neural stem cells,NSCs)移植到慢性海人酸(kainic acid,KA)癫痫鼠海马CA3区后对大鼠癫痫发作的影响。方法用KA脑室注射制作慢性癫痫模型。将原代培养的、EGFP标记的NSCs移植到慢性癫痫鼠的海马CA3区。分别在移植后第2周、第4周、第8周和第12周连续进行7天观察大鼠癫痫发作频率和程度,在移植后第10周进行发作间期右侧海马深部脑电监测。然后取脑冰冻切片,在倒置荧光显微镜下直接观察移植细胞的存活和迁移,用免疫荧光染色观察移植细胞分化情况,Ti mm’s染色观察海马齿状回异常苔状纤维发芽。结果移植后12周仍有大量移植细胞存活(65,045.00±881.72)。NSCs在移植区以胶质细胞分化为主,在齿状回和海马各区以神经元分化为主,γ-氨基丁酸(GABA)能神经元在齿状回门区和海马CA3区分化比率较高。NSCs移植后第4周移植组癫痫鼠的发作次数与对照组相比开始减少,Ti mm’s染色计分和发作程度也有明显改善,两组发作间期脑电图尖、棘波在每个观察期的发放次数分别是3.83±4.96和27.16±21.08。结论将NSCs移植到慢性KA癫痫鼠的海马CA3区,移植细胞不仅能够长期存活、迁移到海马齿状回的各区,而且能够分化为神经元和神经胶质细胞,特别是GABA能神经元,同时还能够抑制齿状回颗粒细胞的苔状纤维发芽,从而减少癫痫发作次数,减轻癫痫发作程度。
Objective To investigate the effects of neural stem cells (NSCs) transplantation on the seizure in rats after transplanting into hippocampal CA3 region of kainic acid (KA) epileptic mice. Methods Chronic intracerebral epilepsy model was made by intracerebroventricular injection with KA. Primary cultured EGFP-labeled NSCs were transplanted into hippocampal CA3 region of chronic epileptic rats. The frequency and extent of epileptic seizures in rats were observed continuously at the second week, the fourth week, the eighth week and the twelfth week after transplantation. The right hippocampal EEG monitoring was performed 10 weeks after the transplantation. Then frozen section of brain was used to observe the survival and migration of transplanted cells directly under inverted fluorescence microscope. The differentiation of transplanted cells was observed by immunofluorescence staining. The germination of mossy fiber of hippocampal dentate gyrus was observed with Ti mm’s staining. Results A large number of transplanted cells still survived 12 weeks after transplantation (65,045.00 ± 881.72). The majority of NSCs were differentiated into glial cells in the grafted area, and mainly differentiated into neurons in the dentate gyrus and the hippocampus. The differentiation ratio of GABAergic neurons in the dentate gyrus and hippocampal CA3 regions high. The number of epileptic mice in the transplantation group began to decrease at the 4th week after transplantation compared with that in the control group, and the scores of Ti mm’s staining and the degree of attack were also significantly improved. In the two groups, Were 3.83 ± 4.96 and 27.16 ± 21.08, respectively. CONCLUSIONS: NSCs can be transplanted into hippocampal CA3 region of chronic KA-induced epileptic rats. The transplanted cells can not only survive long-term, migrate to the hippocampus dentate gyrus, but also differentiate into neurons and glial cells, especially GABAergic neurons But also to inhibit germ-shaped granulosa cells germination, thereby reducing the number of seizures and reduce the extent of seizures.