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目的研究成年大鼠帕金森病(PD)时,黑质内是否存在神经祖细胞,并进一步观察其增殖和分化的情况。方法成年大鼠纹状体内立体定位注射6_羟多巴胺(6_OHDA),并作行为学分析,筛选PD动物模型。取不同存活时间鼠脑黑质节段,用免疫组织化学染色方法,以抗巢蛋白(Nestin)抗体、抗增殖细胞核抗原(PCNA)抗体、抗Ⅲ型β_微管蛋白(Tuj1)抗体、抗酪氨酸羟化酶(TH)抗体分别显示神经祖细胞、分裂细胞、神经元前体细胞和多巴胺(DA)能神经元。光镜观察并细胞计数,统计学分析。结果PD模型大鼠右侧黑质内可见:1.Nestin反应阳性细胞:注射6_OHDA后第10 d,黑质致密部可见少量Nestin阳性细胞;第14 d时,出现大量Nestin阳性细胞;第17 d时开始减少;第21 d几乎检测不到。2.抗PCNA反应阳性细胞:注射6_OHDA后第7 d即可检测到较弱的PCNA阳性细胞;第14 d出现大量PCNA阳性细胞;至21 d开始减少、减弱;28 d时则检测到少量阳性细胞。3.抗Tuj1反应阳性细胞:注射后第10 d,开始出现少量Tuj1阳性细胞;第14 d出现大量Tuj1阳性细胞;第17 d开始减少,第21 d几乎检测不到。4.抗TH反应阳性神经元:注射6_OHDA后第7 d,黑质致密部抗TH反应阳性神经元数量显著少于对照组,且随注射时间延长而逐渐下降。结论成年大鼠纹状体内注射6_OHDA致黑质多巴胺能神经元变性、死亡,能诱导黑质内神经祖细胞在一段时期内大量增殖并向神经细胞方向分化(不包括多巴胺能神经元)。
Objective To investigate the presence or absence of neural progenitor cells in substantia nigra and to observe the proliferation and differentiation of Parkinson’s disease (PD) in adult rats. Methods Adult rats were stereotaxically injected with 6-hydroxydopamine (6_OHDA) into the striatum for behavioral analysis and PD animal models were screened. The brain substantia nigra segments of different survival time were harvested. The expression of Nestin, PCNA and Tuj1 antibodies was detected by immunohistochemical staining. Tyrosine hydroxylase (TH) antibodies show neural progenitor cells, dividing cells, neuronal precursor cells, and dopaminergic (DA) neurons, respectively. Light microscopy and cell counting, statistical analysis. RESULTS: In the right substantia nigra of PD model rats, there were: 1.Nestin-positive cells: A small amount of Nestin-positive cells were observed on the 10th day after injection of 6-OHDA. On the 14th day, a large number of Nestin-positive cells appeared; on the 17th day Began to decrease; almost undetectable on the 21st day. Anti-PCNA-positive cells: A weak PCNA positive cells could be detected on the 7th day after injection of 6-OHDA; a large number of PCNA positive cells appeared on the 14th day; they began to decrease and weakened on the 21st day; a small amount of positive cells were detected on the 28th day cell. Tuj1-positive cells: Tuj1-positive cells began to appear on the 10th day after injection; Tuj1-positive cells appeared on the 14th day; decreased on the 17th day, and almost undetectable on the 21st day. TH-positive neurons: On the 7th day after injection of 6-OHDA, the number of anti-TH positive neurons in the substantia nigra pars compacta was significantly less than that in the control group, and gradually decreased with the injection time. Conclusion Adult rat striatum injected 6_OHDA can degeneration and death of substantia nigra dopaminergic neurons, which can induce large numbers of proliferation and differentiation of neural progenitor cells (excluding dopaminergic neurons) in the substantia nigra within a certain period of time.