宫内缺氧减缓小鼠普肯耶细胞发育和降低小脑肽的表达

Intrauterine hypoxia slowing the development of Purkinje cells and reducing the expression of cerebellin in mice

目的探讨宫内缺氧对出生后小鼠普肯耶细胞的发育及小脑肽表达的影响。方法20只健康成年昆明小鼠孕鼠随机分为2组:对照组与缺氧组,每组10只。自母鼠孕14 d时开始,将缺氧组的母鼠置入动物缺氧箱内,制作宫内缺氧动物模型。待母鼠分娩后,实验动物分为缺氧组和对照组,每组内有出生当日(P0)、P5、P9、P14、P21和P30这6个年龄段,每年龄段小鼠各取5只。取小脑组织做振荡切片,利用免疫荧光技术检测钙结合蛋白(calbindin)阳性普肯耶细胞的发育变化,利用小脑肽1(CBLN1)、CBLN4与calbindin双重标记的方法检测普肯耶细胞突起上小脑肽的表达。利用Western blotting对各组各时间点动物小脑组织内小脑肽蛋白表达进行半定量分析。结果与同龄对照组相比,缺氧组小脑普肯耶细胞数量减少,树突分支减少且排列混乱,皮质内CBLN1与CBLN4的表达明显降低。Western blotting结果与免疫荧光结果吻合。结论宫内缺氧导致小脑普肯耶细胞发育异常以及突触改变。

Objective To investigate the effects of the development of Purkinje cells and expression of cerebellin in postnatal mice with intrauterine hypoxia.Methods Twenty healthy adult Kunming mice were randomly divided into two groups:control group and hypoxia group,with 10 mice in each group.Mice in the hypoxia group were placed in the hypoxia chamber of the animal since the 14 th day of gestation to make an animal model of intrauterine hypoxia.After the mother gave birth,the experimental animals were divided into hypoxia group and control group.There were 6 age groups including postnatal day(P)0,P5,P9,P14,P21 and P30 in each group,and 5 mice in each age group.The cerebellum tissue was taken for vibrating sectioning.The developmental changes of calbindin-positive Purkinje cells were detected by immunofluorescence technique.The expression of cerebellin in Purkinje cell protuberances was detected by cerebellin(CBLN)1,CBLN4 and calbindin double labeling.Finally,Western blotting was used to semi-quantitatively analyze the protein expression of cerebellar peptide in cerebellum at each time point.Results Compared with the control group of the same age,the number of cerebellar Purkinje cells in the hypoxic group decreased,the dendritic branches decreased,and the arrangement was disordered,and the expression of CBLN1 and CBLN4 in the cortex were significantly reduced.Conclusion Intrauterine hypoxia leads to abnormal development of the cerebellar Purkinje cells and synaptic changes.