【摘要】 目的探讨C型尼曼-皮克病(NPC)小鼠模型神经元病变时,维生素D依赖型钙结合蛋白D28k(calbindin D28k)和MC6蛋白异常表达与神经元细胞骨架病变之间的相关性。方法利用免疫组化、免疫荧光标记以及免疫印迹等方法检测NPC1小鼠和野生型对照组小鼠(各8只)脑组织不同部位神经元变性过程中calbindin D28k及MC6蛋白表达,细胞骨架蛋白抗有丝分裂活化蛋白激酶-2(MAP2)和神经丝(neurofilament,NF)的病理变化。结果NPC1小鼠4周龄时,calbindin D28k表达为0·68±0·32,稍微高于野生型小鼠(0·53±0·20,P=0·665),以后5~8周龄的连续变化0·71±0·33,1·22±0·73均低于对照组的1·20±0·47和2·28±1·42(P=0·34,0·045)。NPC1鼠小脑浦肯野神经元发生变性早期,calbindin D28k蛋白表达增高,晚期表达降低。小脑白质、脑干、基底节和部分大脑中出现异常calbindin D28k蛋白颗粒和MC6蛋白。异常的MC6蛋白与calbindin D28k高度共表达。相同部位的神经元NF也明显病理改变。浦肯野神经元则没有发现MC6以及MAP2、NF病理改变。结论calbindin D28k在NPC1小鼠神经元变性早期可能有短暂的神经元保护作用。随着病程发展calbindin D28k表达异常,并与MC6蛋白密切相关。两者均与神经元细胞骨架结构破坏明显相关。浦肯野细胞变性与MC6和MAP2、NF关系不大。由此推测calbindin D28k参与NPC1小鼠模型神经元细胞骨架病变机制;同时,浦肯野神经元变性、死亡的途径与其他的神经元可能不同。更多还原

【Abstract】 Objective To study the expressions of calbindin D28k and MC6 protein, and the relation′s to cytoskeletal pathology in brain tissues of Niemann-Pick disease type C (NPC) mice. Methods The expressions and distributions of calbindin D28k and MC6 protein in brain tissues of NPC mice and wild-type mice were investigated by using immunohistochemical methods and Western-blots. Results During the earlier pathologic stage, the expression of calbindin D28k in neuronal cells of NPC-1 mice brain was increased stronger than that in the wild mice. When the atrophy of Purkinje cells was more easily observed, the expression of calbindin D28k (0.71±0.33, 1.22±0.73) became much weaker than that in the wild mice (1.20±0.47 and 2.28±1.42(P=0.340,0.045)). And in the brains of NPC-1 mice, especially in the white matter of the cerebellum, brainstem and basal ganglia, a lot of positive calbindin D28k and MC6 protein granules dyed by DAB were obviously present. In contrast, the similar granules were hardly seen in the wild mice. The distribution of pathologic protein (MC6) in the NPC mice was nearly colocalized with calbindin D28k staining. Both MC6 protein and positive granules of calbindin D28k were present in the cytoskeleton which was pathologically changed. The immunoreactivity of calbindin D28k in Purkinje cells of NPC brains in 8 weeks old mice was decreased, and the majority of Purkinje cells were gone. Conclusions The expression of calbindin D28k in NPC mice brains suggests that calbindin D28k might be involved in degeneration and death of neuronal cells. The detailed roles of calbindin D28k in NPC pathogenesis might still remain to be elucidated. The abnormal distribution of calbindin D28k should be physically colocalized with MC6 protein immunoreactivity in brain of NPC mice. Both of them should have a close relation to the neuronal cytoskeletal (MAP2, neurofilament) pathology. Unexpectedly, the similar changes were not observed in the degenerating Purkinje cells, we presume that a different way leading to Purkinje cell degeneration might be present.更多还原