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目的:通过检测细胞周期相关蛋白D1(Cyclin D1)在肌萎缩侧索硬化症(ALS)转基因小鼠大脑皮层和海马中的表达变化,探讨Cyclin D1表达改变与ALS发病的关系。方法:选取成年ALS小鼠和同窝野生型小鼠,于发病早、中、晚期(95 d,108 d,122 d)取材,应用免疫荧光技术检测Cyclin D1在大脑皮层和海马的表达规律及与神经元和星形胶质细胞的共定位关系,应用RT-PCR检测Cyclin D1 mRNA表达情况,应用免疫印迹法检测蛋白表达量的改变。结果:ALS小鼠和野生型小鼠大脑皮层和海马中均可检测到Cyclin D1阳性细胞,且与神经元共表达。在发病早、中、晚期,ALS小鼠大脑皮层中Cyclin D1 mRNA和蛋白表达较野生型鼠增多(P<0.05,P<0.01,P<0.001);与同窝野生型小鼠相比,ALS小鼠海马中Cyclin D1 mRNA和蛋白在发病的早、中、晚期表达均降低(P<0.05,P<0.01,P<0.001)。Cyclin D1阳性细胞主要分布在海马CA区(包括CA1、CA2和CA3),DG区仅散在表达。结论:Cyclin D1在ALS转基因小鼠大脑皮层和海马中表达异常,表明Cyclin D1调节的细胞周期改变与ALS大脑皮层和海马区病变密切相关。
OBJECTIVE: To investigate the relationship between Cyclin D1 expression and the pathogenesis of ALS by detecting the expression of Cyclin D1 in the cerebral cortex and hippocampus of ALS transgenic mice. Methods: Adult ALS mice and wild-type littermates were selected and harvested at the early, middle and late stages (95 d, 108 d, 122 d). The expression of Cyclin D1 in the cerebral cortex and hippocampus was detected by immunofluorescence staining. With colocalization of neurons and astrocytes, the expression of Cyclin D1 mRNA was detected by RT-PCR. The protein expression was detected by Western blotting. Results: Cyclin D1 positive cells were detected in both cerebral cortex and hippocampus of ALS mice and wild-type mice, and were co-expressed with neurons. Compared with WT mice, the expression of Cyclin D1 mRNA and protein in ALS mice increased significantly (P <0.05, P <0.001, P <0.001) The expression of Cyclin D1 mRNA and protein in the hippocampus of mice decreased in the early, middle and late stages of onset (P <0.05, P <0.01, P <0.001). Cyclin D1 positive cells mainly distributed in the hippocampal CA area (including CA1, CA2 and CA3), DG area only scattered expression. CONCLUSION: Cyclin D1 is abnormally expressed in the cerebral cortex and hippocampus of ALS transgenic mice, indicating that Cyclin D1-mediated cell cycle changes are closely related to ALS cerebral cortex and hippocampal lesions.