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目的研究头皮位点药物注射对HIE新生大鼠内源性神经干细胞增殖分化及神经再生抑制因子Nogo-A mRNA表达的影响。方法新生7日龄SD大鼠80只。随机分成4组:假手术组、模型组、位点注射9 g.L-1盐水组及位点注射V itB1、V itB12、组,每组各20只。采用结扎左侧颈总动脉,并吸入2 h氮氧混合气体制作新生大鼠HIE模型。位点注射9 g.L-1盐水组和位点注射V itB1、V itB12组在模型建立第14天,分别在大鼠的左侧额顶叶皮下注射等量9 g.L-1盐水或V itB1、V itB12混合稀释液,1次.d-1,共20 d;假手术组和模型组不予处理。各组于干预后7 d处死动物,40 g.L-1多聚甲醛心脏灌注后断头取脑组织,分别做连续冠状切片。用免疫荧光方法测定各组海马神经干细胞标志物巢蛋白(Nestin)表达,免疫组织化学方法检测其海马胶质纤维酸性蛋白(GFAP)和神经元特异性烯醇化酶(NSE)的表达,原位杂交法测定其海马神经再生抑制因子Nogo-A mRNA的表达水平。结果 1.位点注射可增加海马Nestin的表达,以注射V itB1、V itB12组表达最高,假手术组表达最少。2.海马NSE免疫组织化学显示假手术组和位点注射V itB1、V itB12组表达较多,模型组海马表达最少(P<0.05)。3.海马GFAP免疫组织化学显示假手术组表达最少,模型组最多,位点注射9 g.L-1盐水和位点注射V itB1、V itB12可抑制GFAP的表达,其中以位点注射V itB1、V itB12组抑制作用最为明显(P<0.05)。4.假手术组Nogo-A mRNA表达最少,模型组表达最高,位点注射9 g.L-1盐水组和位点注射V itB1、V itB12组表达水平均低于模型组,其中位点注射V itB1、V itB12组更为明显(P<0.05)。结论头皮位点药物注射可激发内源性神经干细胞产生,促进其增殖分化,同时可抑制神经再生抑制因子Nogo-A的表达,从而促进大脑神经的修复。
Objective To study the effects of scalp site injection on the proliferation and differentiation of endogenous neural stem cells and the expression of Nerve regeneration inhibitory factor Nogo-A mRNA in HIE neonatal rats. Methods Eighty newborn SD rats aged 7 days were enrolled. The rats were randomly divided into 4 groups: sham operation group, model group, site injection of 9 g.L-1 saline group and site injection of VitB1, VitB12, group, 20 rats in each group. The neonatal rat model of HIE was established by ligating the left common carotid artery and inhaling nitrogen and oxygen mixture for 2 h. The rats in the 9 gL-1 saline group and the VitB1 and VitB12 groups were injected subcutaneously with the same dose of 9 gL-1 saline or VitB1, V itB12 mixed diluent, once .d-1, a total of 20 d; sham-operated group and model group untreated. Animals in each group were sacrificed at 7 days after the intervention, and brains were removed after cardioplegia with 40 g. L-1 paraformaldehyde and consecutive coronal sections were made respectively. Immunofluorescence method was used to detect the expression of Nestin, a marker of hippocampal neural stem cells in each group. The expression of glial fibrillary acidic protein (GFAP) and neuron specific enolase (NSE) in hippocampus was detected by immunohistochemistry. The expression of Nogo-A mRNA in hippocampus was detected by hybridization. Site injection can increase the expression of Nestin in hippocampus, in order to inject VitB1, VitB12 group had the highest expression and sham operation group had the least expression. The hippocampal NSE immunohistochemistry showed that there were more expression of VitB1 and VitB12 in the sham operation group and the site of injection, the least in the hippocampus of the model group (P <0.05). Immunohistochemistry of GFAP in the hippocampus showed that the expression of GFAP in the sham-operation group was the lowest, the model group was the most, the site injection of 9 gL-1 saline and VitB1 and VitB12 inhibited the expression of GFAP, itB12 group the most obvious inhibition (P <0.05). The expression of Nogo-A mRNA was the lowest in sham-operation group, the expression was the highest in model group, the expression level of VitoB1 and VitoB12 in group injected with 9 gL-1 saline and VitoB12 was lower than that in model group, , V itB12 group was more obvious (P <0.05). Conclusions Scalp site injection can stimulate the production of endogenous neural stem cells, promote their proliferation and differentiation, and inhibit the expression of nerve regeneration inhibitor Nogo-A, thereby promoting the repair of cerebral nerves.