卵母细胞携带的HBV DNA在小鼠早期胚胎中的复制与表达  

Replication and Expression of Oocyte-mediated HBV DNA in Early Mouse Embryo

作　　者：陈桂兰[1] 黄天华[1] 谢庆东[1] 王晓梅[1] 熊小芳[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3期179-182,共4页Carcinogenesis,Teratogenesis & Mutagenesis

基　　金：国家自然科学基金资助(No.39970374)。

摘　　要：背景与目的:研究卵母细胞携带的乙肝病毒(HepatitisBvirus,HBV)DNA能否在小鼠早期胚胎中复制与表达,为HBV经卵垂直传播提供科学证据。材料与方法:手术暴露昆明小鼠卵巢,将pBR322_HBV重组质粒DNA注射到昆明小鼠卵巢内,1个动情周期(4d)后进行超排卵并与正常雄性小鼠合笼,收集二细胞胚,用荧光原位杂交(Fluorescenceinsituhybridization,FISH)检测HBV在早期胚胎内的整合和复制;用逆转录聚含酶链反应(Reversetranscriptase_polymerasechainreaction,RT_PCR)和免疫荧光试验检测HBV基因在早期胚胎的表达。结果:FISH在小鼠二细胞胚细胞核上检测到HBVDNA阳性杂交信号,RT_PCR和免疫荧光试验在二细胞胚内检测到HBxmRNA和HBsAg。结论:HBVDNA进入雌性生殖细胞后,可以整合到宿主基因组内,通过自然受精随卵母细胞进入胚胎,并能在胚胎中复制与表达。提示HBV有可能通过雌性生殖细胞进行垂直传递。BACKGROUND&AIM: To study whether hepatitis B virus DNA carried by oocyte can express and replicate in mouse embryo,then provide evidence for that HBV can transmit vertically via oocytes. MATERIAL AND METHODS: Ovaries of young adult female mice were injected with HBV DNA.These females were superovulated after an estrual cycle and serially mated to males.Two-cell embryos were collected for Fluorescence in situ hybridization(FISH)to detect the integration of HBV DNA into nuclei and its replication with cell division in embryonic development.Reverse transcriptase polymerase chain reaction(RT-PCR)and immunofluorescence assay were performed to observe the expression of the HBV gene in two-cell stage. RESULTS: FISH demonstrated that the each nucleus in some two-cell embryos presented positive signals for HBx DNA.RT-PCR showed the specific bands of cDNA of HBx DNA in some two-cell embryos.Immunofluoresence assay presented the positive signals for HBsAg in some two-cell embryos. CONCLUSION: HBV DNA integrated into oocyte genome can be brought into embryo by fertilization with normal spermatozoon.The replication and expression of HBV genes in embryo provide the direct evidence for that HBV DNA are able to transmit vertically to next generation via female germ line.

关 键 词：乙型肝炎病毒DNA 复制与表达 卵母细胞 小鼠胚胎 

分 类 号：Q345.21[生物学—遗传学] R512.62[医药卫生—内科学]