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[乙型肝炎病毒携带者转基因小鼠模型的建立]

[Development of a hepatitis B virus carrier transgenic mice model].

作者信息

Caner Müge, Arat Sezen, Bircan Rifat

机构信息

Maltepe Universitesi Tip Fakültesi, Tibbi Biyoloji Anabilim Dali, Istanbul.

出版信息

Mikrobiyol Bul. 2008 Jan;42(1):71-81.

Abstract

The studies for the development of transgenic mice models which provide important profits for the studies concerning immunopathogenesis of hepatitis B virus (HBV) infections are in progress since 20 years. For this purpose different lineages bearing whole HBV genome or selected viral genes have been developed and their usage in clarifying the HBV replication and pathogenesis mechanisms have been emphasized. The aim of this study was to develop and breed a HBV carrier mice model. In the study the full HBV genome has been transferred to mouse embryos by microinjection procedure. Following transgenic manipulation, the HBV carriers among the daughter mice have been detected by molecular methods in which HBV-DNA replication and expression have been shown. The manipulations for transgene transfers have been performed in TUBITAK Marmara Research Center Transgene Laboratory, Gebze, Istanbul. The HBV-DNA carrier mice have been demonstrated by polymerase chain reaction (PCR) using the DNA samples obtained from tail tissues and also by dot-blot hybridization of the mice sera. Integrated HBV-DNA has been detected by applying in-situ hybridization to the liver tissue sections. HBV-DNA expression has been shown by reverse transcriptase PCR method with total RNA molecules that have been isolated from the liver tissues of the HBV-DNA carrier mice. HBsAg has been detected in the liver by immunohistochemical method, and HBsAg and HBeAg have additionally been demonstrated by ELISA. HBV genome, expression of the genome and the expression products have been determined in approximately 10% of the mice of which HBV-DNA have been transferred. By inbreeding heterozygote carrier mice, homozygote HBV transgenic mice line have been obtained. These HBV transgenic mice are the first lineages developed in our country. It is hopefully thought that this HBV carrier transgenic mouse model may contribute to the studies on the pathogenesis of HBV infections which are important health problems in the world as well as in Turkey.

摘要

20年来,为开发转基因小鼠模型所开展的研究一直在进行,这些研究为乙型肝炎病毒(HBV)感染的免疫发病机制研究带来了重要收益。为此,已培育出携带完整HBV基因组或选定病毒基因的不同品系,并强调了它们在阐明HBV复制和发病机制方面的用途。本研究的目的是开发并培育一种HBV携带小鼠模型。在该研究中,通过显微注射程序将完整的HBV基因组导入小鼠胚胎。转基因操作后,通过显示HBV-DNA复制和表达的分子方法在子代小鼠中检测到HBV携带者。转基因转移操作在位于伊斯坦布尔盖布泽的图比塔克斯马尔马拉研究中心转基因实验室进行。通过对从尾部组织获得的DNA样本进行聚合酶链反应(PCR)以及对小鼠血清进行斑点杂交,证实了HBV-DNA携带小鼠。通过对肝组织切片进行原位杂交检测到整合的HBV-DNA。用从HBV-DNA携带小鼠肝脏组织中分离的总RNA分子,通过逆转录酶PCR方法显示了HBV-DNA表达。通过免疫组织化学方法在肝脏中检测到HBsAg,另外通过ELISA检测到HBsAg和HBeAg。在约10%已转移HBV-DNA的小鼠中确定了HBV基因组、基因组表达及表达产物。通过对杂合子携带小鼠进行近亲繁殖,获得了纯合子HBV转基因小鼠品系。这些HBV转基因小鼠是我国培育出的首批品系。有望认为,这种HBV携带转基因小鼠模型可能有助于对HBV感染发病机制的研究,HBV感染在世界以及土耳其都是重要的健康问题。

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