5-氟尿嘧啶对神经管畸形胎鼠脑组织基因表达的影响  

Effect of 5-fluorouracil on gene expression in embryonic brain tissues of mice with neural tube defects

作　　者：王秀伟[1] 官臻[1] 朱智强[1] 牛勃[1] 王建华[1] Wang Xiuwei;Guan Zhen;Zhu Zhiqiang;Niu Bo;Wang Jianhua(Beijing Municipal Key Laboratory of Child Development and Nutriomics,Capital Institute of Pediatrics,Beijing 100020,China)

机构地区：[1]首都儿科研究所儿童发育营养组学北京市重点实验室,北京100020

出　　处：《发育医学电子杂志》2020年第3期213-219,共7页Journal of Developmental Medicine (Electronic Version)

基　　金：国家自然科学基金(81600984、81571443、81700777);北京市自然科学基金(7172038);首都儿科研究所所级基金(FX-2018-05/04)。

摘　　要：目的利用5-氟尿嘧啶(5-fluorouracil,5-FU)诱导C57BL/6J小鼠神经管畸形(neural tube defects,NTDs),观察5-FU对NTDs胎鼠脑组织基因表达的影响。方法采用无特定病原体(specific pathogen free,SPF)级C57BL/6J小鼠,孕鼠随机分为实验组和对照组。在孕7.5 d时,实验组孕鼠腹腔注射12.5 mg/kg的5-FU(叶酸相关代谢通路中关键酶胸苷酸合成酶抑制剂);对照组孕鼠腹腔注射等体积的生理盐水。孕13.5 d时通过体视显微镜及HE染色观察胎鼠畸形及神经管发育情况。在此基础上利用表达谱芯片分析NTDs胎鼠脑组织中基因表达的变化,并利用实时荧光定量PCR进行验证。统计学方法采用t检验。结果实验组小鼠NTDs的发生率为42.3%(22/52),主要为露脑畸形;对照组无一例NTDs。HE染色可见NTDs胎鼠的端脑和后脑结构紊乱。表达谱芯片筛选出差异表达基因137个,其中表达上调的基因40个,表达下调的基因97个,这些基因主要参与了多器官发育、细胞增殖及分化等生物过程以及与脑部发育相关的重要通路。选择表达差异明显且与胚胎神经发育相关的基因Avp、Hes3和Wnt1进行实时荧光定量PCR验证,结果发现所验证的基因表达情况与表达谱芯片的结果一致。结论5-FU可能通过影响胚胎神经发育过程中与神经发育相关基因Avp、Hes3及Wnt1等的表达,进而导致NTDs的发生。Objective To investigate the gene expression alteration in the brain tissue of neural tube defects(NTDs)fetal mice of C57BL/6J induced by 5-fluorouracil(5-FU).Methods Specific pathogen free(SPF)C57BL/6J pregnant mice were randomly divided into the experimental group and the control group.5-FU(12.5 mg/kg),thymidine synthase inhibitor,was injected intraperitoneally into pregnant C57BL/6J mice in the experimental group on gestational day 7.5.The pregnant mice in the control group were intraperitoneally injected with equal volume of normal saline.The malformation and neural tube closure were carefully observed by stereomicroscope and hematoxylin-eosin(HE)staining on gestational day 13.5.The expression of different genes was studied by microarray and verified by real-time quantitative polymerase chain reaction(RT-qPCR).The statistics analysis was performed with t test.Results The incidence of NTDs was 42.31%(22/52),especially exencephaly in the experimental group.There were no NTDs in the control group.The structure of telencephalon and hindbrain were abnormal in NTD mice by HE staining.The 137 genes were significantly difference in NTD mice compared with the control group,of which 40 genes were up-regulated and 97 genes were down-regulated.These genes were mainly involved in the biological processes of multicellular organismal development,cell proliferation,differentiation,and the important pathways related to brain development.Avp,Hes3 and Wnt1 genes,which were neural development related genes,were chosen and validated by RT-qPCR to show the significantly abnormal expressions in NTD brain tissues.Conclusion 5-FU might affect the expressions of the Avp,Hes3 and Wnt1 genes related to neural development during the process of embryonic brain development,and then lead to the occurrence of NTDs.

关 键 词：神经管畸形 胚胎 5-氟尿嘧啶 小鼠 基因芯片 

分 类 号：R748[医药卫生—神经病学与精神病学]