嗅成鞘细胞的移植和迁移对脑梗死大鼠神经功能恢复的作用

Effect of olfactory ensheathing cells transplantation and migration on the recovery of neurological function after cerebral infarction in rats

目的探讨对脑梗死大鼠移植嗅成鞘细胞(OEC)促进神经功能恢复的作用及OEC迁移规律。方法从2.5个月龄SD大鼠嗅球取材,利用差异贴壁方法纯化OEC。移植前,用Hoechst 33342标记OEC。按双肾双夹方法制备易卒中型肾血管性高血压大鼠模型,模型制备成功后10周,电凝大脑中动脉制作大脑中动脉梗死(MCAO)模型。MCAO大鼠63只,按随机数字法随机分为梗死侧移植组、健侧移植组及双侧移植组,每组21只大鼠。于移植术前,移植术后第2和6周行大鼠神经运动功能、感觉功能检查;免疫组化法观察脑组织神经生长相关蛋白-43(GAP-43)及神经丝蛋白(NF)的表达情况。移植后6周取移植部位脑组织行冷冻切片,荧光显微镜下观察OEC迁移情况。结果①梗死侧移植组、健侧移植组及双侧移植组移植前运动评分中位数评分均为2,移植后第2周分别为5、4、7,第6周分别为6,7、9;移植前感觉功能评价中位数3组均为6.67%,移植后第2周3组分别为20.00%、13.33%和33.33%,移植后第6周分别为26.67%、33.33%和46.67%。以上指标各时间点梗死侧移植组、健侧移植组比较差异无统计学意义;双侧移植组与其他两组比较差异均有统计学意义(P<0.05,P<0.01)。②免疫组化染色显示,各时间点梗死侧移植组、健侧移植组梗死灶边缘区GAP-43、NF阳性表达差异无统计学意义;双侧移植组与其他两组比较表达均增强,差异均有统计学意义(P<0.05,P<0.01)。③移植OEC后6周3组均观察到OEC向梗死灶及对侧的皮质迁移。结论脑梗死大鼠OEC健侧移植能够起到与患侧移植同样的促进神经功能恢复的作用,这种作用与OEC促进神经纤维再生及较强的自我迁移能力有关。

Objective To explore the effects of olfactory ensheathing cells （OEC） transplanted to the normal side of brain in rats with cerebral infarction on promoting recovery of neurological function and the migratory rule of OEC. Methods OEC were harvested from the olfactory bulb of the 2.5-month-old SD rats and were purified using differential time adherent method. They were labeled with Hoeehst 33342 before transplantation. A stroke-prone renovaseular hypertensive rat model （RHRSP） was established by the two-kidney two-clip method. Ten weeks after the successful establishment of the model, 63 rat models of middle cerebral artery occlusion （MCAO） were made by electric coagulation of middle cerebral artery （ MCA ）. The MCAO rats were randomly allocated into infarcted side transplantation, normal side transplantation, and bilateral transplantation groups （21 in each group）. Before the transplantation, 2 and 6 weeks after the transplantation, the psychomotor function and sensory function of the rats were examined. The expressions of growth-associated protein-43 （GAP-43） and neurofilament （NF） protein in brain tissues were detected by immunohistochemical method. Results ①The median motor scores before transplantation were 2 in all the three transplantation groups. At the second week after the transplantation the scores were 5, 4, and 7, respectively, and at the sixth weeks after the transplantation were 6, 7, and 9, respectively. Before transplantation, the median sensory function evaluation scores were 6.7% in all the three groups; At the second week after the transplantation, they were 20. 00% , 13.33%, and 33. 33% , respectively; at the sixth week after the transplantation, they were 26.67% , 33.33% , and 46.67% , respectively. There were no statistically significant differences of the above indexes at all time points between the infarcted side and the normal side transplantation group; but there were statistically significant differences between the bilateral transplantation group and the other two groups. ②Immunohistochemistry staining indicated that there were no statistically significant differences in the positive expressions of GAP-43 and NF in the infarction border areas at all time points between the infarcted side transplantation group and normal side transplantation group; the expressions of the bilateral transplantation group were higher than those of the other two groups, and there were statistically significant differences among them. ③Six weeks after OEC transplantation, the OEC migrated to infarction foci and contralateral cortices in all the three groups were observed. Conclusion The OEC transplanted to the normal side of brain in rats with cerebral infarction may play a same role as to the affected sides in promoting recovery of neurological functional, because the OEC can promote nerve fiber regeneration and has strong migratory ability.