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

doi:10.3969/j.issn.1004-616x.2005.04.001

癌变·畸变·突变 ›› 2005, Vol. 17 ›› Issue (4): 193-197.doi: 10.3969/j.issn.1004-616x.2005.04.001

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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

黄天华;谢庆东;陈霓彤

汕头大学医学院生殖医学研究中心，广东　汕头　515041

收稿日期:2005-01-24 修回日期:2005-03-31 出版日期:2005-07-30 发布日期:2005-07-30
通讯作者: 黄天华

Replication and Expression of Sperm-Mediated Genes of

HUANG Tian-hua;XIE Qing-dong;CHEN Ni-tong

Research Center of Reproductive Medicine, Shantou

Received:2005-01-24 Revised:2005-03-31 Online:2005-07-30 Published:2005-07-30
Contact: HUANG Tian-hua

摘要/Abstract

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(Hepatitis B virus, HBV)通过精子垂直传播的可能性。方法: 将金黄地鼠精子与pBR322-HBV DNA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(Fluorescence in situ hybridization, 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： ① FISH：用全长HBV DNA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HBV DNA杂交信号；②RT-PCR：在检测样本中可见特异的HBx DNA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。

关键词: 乙肝病毒基因, 复制与表达, 垂直传播, 金黄地鼠

Abstract: 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: ① FISH HBV DNA-positive signals within each nucleus of two-cell embryo in the test sample were observed. ② RT-PCR the positive bands for HBx DNA in the tested samples were detected.③ Immunofluoresence 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.

Key words: HBV gene, replication and expression, vertical transmission, golden hamster

黄天华, 谢庆东, 陈霓彤. 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[J]. 癌变·畸变·突变, 2005, 17(4): 193-197.

HUANG Tian-hua, XIE Qing-dong, CHEN Ni-tong. Replication and Expression of Sperm-Mediated Genes of[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2005, 17(4): 193-197.

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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 Expression of Sperm-Mediated Genes of

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