精子携带的乙肝病毒基因在金黄地鼠胚胎中的复制与表达  

Replication and Expres sion of Sperm-Mediated Genes of Hepatitis B Virus in Two-Cell Embryo of Golden Hamster

作　　者：黄天华[1] 谢庆东[1] 陈霓彤[1] 曾曦凡[1] 吴丛梅[1] 王晓梅[1] 

机构地区：[1]汕头大学医学院生殖医学研究中心,广东汕头515041

出　　处：《癌变．畸变．突变》2005年第4期193-197,共5页Carcinogenesis,Teratogenesis & Mutagenesis

基　　金：国家自然科学基金资助项目(No.39970374)

摘　　要：背景与目的:乙型肝炎是危害人类健康的全球性疾病。本研究旨在探索乙肝病毒(HepatitisBvirus,HBV)通过精子垂直传播的可能性。方法:将金黄地鼠精子与pBR322-HBVDNA重组质粒进行共培养后,与金黄地鼠正常卵母细胞受精,取2-细胞胚分别制片和提取RNA,用荧光原位杂交(Fluorescenceinsituhybridization,FISH)、RT-PCR和免疫荧光检测技术研究精子携带的HBV基因在胚胎细胞中的复制与表达。结果:①FISH:用全长HBVDNA探针与128个2-细胞胚进行FISH,其中3个胚胎FISH结果阳性,每个胚胎的2个间期核上均有HBVDNA杂交信号;②RT-PCR:在检测样本中可见特异的HBxDNA阳性条带,阴性对照1(模板为灭菌水)和阴性对照2(未加反转录酶)均为阴性;③免疫荧光检测:2-细胞胚经血清封闭后,分成两组。一组作检测样本,另一组用PBS代替一抗(小鼠抗人HBsAg)作为阴性对照,检测样本中可见阳性免疫荧光信号,阴性对照未见信号。结论:以精子为载体,携带到卵内的HBV基因能够在胚胎细胞中复制和表达,提示HBV能够通过男性生殖细胞由父亲垂直传递给子代。BACKGROUND&AIM: Hepatitis B is a public health problem of worldwide importance.To explore possibility of hepatitis B virus(HBV)vertically transmitting via spermatozoon,to study the replication and expression of sperm-mediated HBV genes in two-cell embryos of golden hamster. MATERIAL AND METHODS: Golden hamster oocytes were fertilized with spermatozoa cocultured with pBR322-HBV DNA plasmids.The slide preparation and RNA extraction of two-cell embryos were carried out.Fluorescence in situ hybridization(FISH),RT-PCR and immunofluoresence assay were performed to detect the replication and expression of sperm-mediated HBV genes in the embryonic cells. RESULTS: ①FISH HBV DNA-positive signals within each nucleus of two-cell embryo in the test sample were observed.②RT-PCR the positive bands for HBx DNA in the tested samples were detected.③Immunofluoresence assay the HBsAg-positive signals within cytoplasm of embryonic cells in the test sample were found. CONCLUSION: Sperm-mediated HBV genes are able to replicate themselves and express their function in early mouse embryo.It suggested that the vertical transmission of HBV via male germ cells from father to offspring could exist.

关 键 词：乙肝病毒基因 复制与表达 垂直传播 金黄地鼠 

分 类 号：R394-33[医药卫生—医学遗传学] R373.21[医药卫生—基础医学]