乙硫氨酸对神经管畸形胎鼠脑组织Fos和Cyp2e1基因表达的影响  

Influence of ethionine on Fos and Cyp2e1 genes in brain tissue of mice embryos with neural tube defects

作　　者：张丽[1] 张钰璇 王秀伟[3] 李建婷 赵娟[4] 孙雨晴 刘娟娟 张丽慧 曹睿[2] 解军[2] ZHANG Li;ZHANG Yu-xuan;WANG Xiu-wei;LI Jian-ting;ZHAO Juan;SUN Yu-qing;LIU Juan-juan;ZHANG Li-hui;CAO Rui;XIE Jun(Hepatobiliary and Pancreatic Surgery and Liver Transplant Center,the First Hospital of Shanxi Medical University,Taiyuan,Shanxi 030000,China;Department of Biochemistry and Molecular Biology,Shanxi Medical University,Taiyuan,Shanxi 030000,China;Department of Translational Medicine Research,Capital Institute of Pediatrics,Beijing 100000,China;Department of Mental Health,the First Hospital of Shanci Medical University,Taiyuan,Shanxi 030000,China;Department of Science and Technology,the First Hospital of Shanxi Medical University,Taiyuan,Shanxi 030000,China;Department of Anesthesiology,the First Hospital of Shanxi Medical University,Taiyuan,Shanxi 030000,China)

机构地区：[1]山西医科大学第一医院肝胆胰外科及肝脏移植中心,山西太原030000 [2]山西医科大学生物化学与分子生物学教研室,山西太原030000 [3]首都儿科研究所转化医学研究室,北京100000 [4]山西医科大学第一医院精神卫生科,山西太原030000 [5]山西医科大学第一医院科技处,山西太原030000 [6]山西医科大学第一医院麻醉科,山西太原030000

出　　处：《中华实用诊断与治疗杂志》2023年第4期325-331,共7页Journal of Chinese Practical Diagnosis and Therapy

基　　金：国家自然科学基金(82201319);山西省基础研究计划(20210302124291,20210302123347);山西省卫生健康委科研课题计划(2020090);山西医科大学博士启动基金项目(XD2005,SD2004)。

摘　　要：目的采用乙硫氨酸诱导小鼠胚胎神经管畸形(neural tube defects,NTDs),观察乙硫氨酸对NTDs小鼠胚胎脑组织基因表达的影响。方法SPF级C57BL/6J孕鼠80只,随机分为模型组42只和对照组38只。孕7.5 d时,模型组孕鼠腹腔注射乙硫氨酸500 mg/kg,对照组腹腔注射等剂量磷酸盐缓冲液。孕10.5 d时体视显微镜下分离胚胎并行HE染色,观察胚胎及神经管发育情况。取模型组NTDs胚胎脑组织和对照组正常胚胎脑组织进行转录组测序,并对差异表达基因进行GO功能注释分析、KEGG通路富集分析、蛋白质互作网络分析。采用实时荧光定量PCR法检测差异表达基因mRNA相对表达量,采用Western blot法检测差异表达基因的蛋白相对表达量,对差异表达基因进行验证。结果模型组吸收胎、生长缓慢、NTDs发生率(10.3%、19.5%、56.0%)均高于对照组(2.4%、4.5%、0)(P<0.05),正常胚胎发生率(12.6%)低于对照组(93.1%)(χ^(2)=353.690,P<0.001),其他畸形发生率(1.6%)与对照组(0)比较差异无统计学意义(χ^(2)=4.144,P=0.113)。对照组胚胎神经管发育良好,管腔规则;模型组NTDs胚胎发育迟缓,端脑和后脑形态异常,结构紊乱。与对照组正常胚胎比较,模型组NTDs胚胎差异表达基因119个,其中表达上调基因30个,表达下调基因89个;差异表达基因主要参与多器官发育、细胞增殖及分化、氧转运等功能,主要富集于代谢通路;蛋白质互作网络分析细胞代谢相关差异表达基因前10位为Cyp4a10、Cyp4a12a、Cyp4b1、Cyp2e1、Cyp2j5、Ugt2b34、Ugt2b38、Fos、Hoxa5、Hoxa7。模型组NTDs胚胎脑组织Cyp4a10、Cyp4a12a、Cyp4b1、Cyp2e1、Cyp2j5、Ugt2b34、Ugt2b38 mRNA相对表达量均低于对照组正常胚胎(P<0.05),Fos、Hoxa5、Hoxa7 mRNA相对表达量均高于对照组正常胚胎(P<0.05),其中Cyp2e1、Fos mRNA相对表达量差异最大。模型组NTDs胚胎脑组织Cyp2e1蛋白相对表达量(0.187±0.030)低于对照组正常胚胎(1.978±0.125)(t=13.970,Objective To induce neural tube defects(NTDs)in the brain tissue of mice embryos with ethionine and to observe the influence of ethionine on gene expression in the brain tissue of mice embryos with NTDs.Methods Eighty SPF C57BL/6Jpregnant mice were randomly divided into model group(n=42)and control group(n=38).By day 7.5of gestation,model group was intraperitoneally injected with ethionine 500mg/kg,while control group was intraperitoneally injected with equivalent volume of PBS.By day 10.5of gestation,the embryos were isolated under stereomicroscope and were stained with HE to observe the development of embryos and neural tubes.The brain tissues of NTDs embryos in model group and the brain tissues of normal embryos in control group were taken for transcriptome sequencing,and differentially expressed genes were analyzed for GO functional annotation,KEGG pathway enrichment,and protein interaction network analysis.The relative expressions of the mRNA and protein of differentially expressed genes were detected by real-time fluorescence quantitative PCR and Western blot,and the differentially expressed genes were verified.Results The incidences of embryo resorption,slow growth and NTDs were higher in model group(10.3%,19.5%,56.0%)than those in control group(2.4%,4.5%,0)(P<0.05),the incidence of normal embryos was lower in model group(12.6%)than that in control group(93.1%)(χ^(2)=353.690,P<0.001),and the incidence of other malformations showed no significant difference between two groups(1.6%vs.0)(χ^(2)=4.144,P=0.113).In control group,the embryonic neural tubes developed well with regular lumens.In model group,the embryos of NTDs developed slowly,with abnormal morphology and structural disorder in the telencephalon and hindbrain.Compared with normal embryos in control group,there were 119differentially expressed genes in NTDs embryos in model group,including 30up-regulated genes and 89down-regulated genes.The differentially expressed genes were mainly involved in multiple organ development,cell proliferation and diff

关 键 词：神经管畸形 乙硫氨酸 小鼠胚胎 转录组测序 

分 类 号：R714.5[医药卫生—妇产科学]