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目的检测DDX3和酪蛋白激酶1ε(CK1ε)在肌萎缩侧索硬化症(ALS)转基因鼠海马中的表达变化,揭示DDX3和CK1ε的表达改变与ALS发病的关系。方法 33只ALS转基因鼠和33只野生型鼠分别于发病不同时期取材,应用RT-PCR和Western blotting技术检测海马组织中DDX3和CK1ε的表达变化;通过免疫荧光双标染色技术观察DDX3和CK1ε阳性细胞的分布特点及其与神经元、星形胶质细胞等的共表达情况。结果与野生型鼠相比,ALS转基因鼠海马中DDX3 mRNA和蛋白在发病早期变化不明显,中期和晚期表达均明显降低;CK1εmRNA和蛋白则在发病早期、中期和晚期表达均降低。免疫荧光双标结果显示,在ALS鼠和野生型鼠海马齿状回区和CA区均可检测到DDX3和CK1ε阳性细胞。DDX3和CK1ε主要表达在神经元,在星形胶质细胞未见表达。与野生型鼠比较,ALS转基因鼠海马中DDX3和CK1ε免疫反应性明显降低。结论 DDX3和CK1εmRNA和蛋白在ALS转基因鼠海马中表达均降低,表明DDX3和CK1ε表达异常与ALS海马区病变密切相关。
Objective To detect the expression of DDX3 and casein kinase 1ε (CK1ε) in the hippocampus of amyotrophic lateral sclerosis (ALS) transgenic mice and to reveal the relationship between the changes of DDX3 and CK1ε and the pathogenesis of ALS. METHODS: Thirty-three ALS transgenic mice and 33 wild-type mice were harvested at different time points respectively. The expression of DDX3 and CK1ε in hippocampus were detected by RT-PCR and Western blotting. DDX3 and CK1ε were detected by immunofluorescence double staining The distribution of cells and their co-expression with neurons, astrocytes and so on. Results Compared with wild-type mice, the expression of DDX3 mRNA and protein in the hippocampus of ALS transgenic mice did not change significantly in the early stage of disease, but both of them were significantly lower in the middle and late stages of the ALS transgenic mice. The expression of CK1ε mRNA and protein decreased in the early, middle and late stages of the disease. Immunofluorescence double-labeled results showed that in both ALS and wild-type mice hippocampal dentate gyrus and CA area can be detected DDX3 and CK1ε-positive cells. DDX3 and CK1ε are mainly expressed in neurons and are not expressed in astrocytes. The immunoreactivity of DDX3 and CK1ε in hippocampus of ALS transgenic mice was significantly lower than that of wild-type mice. Conclusion The expression of DDX3 and CK1ε mRNA and protein in the hippocampus of ALS transgenic mice decreased, indicating that the abnormal expression of DDX3 and CK1ε is closely related to the ALS hippocampal lesions.