乙肝病毒X基因转基因树鼩模型的建立和病理学研究

目的:探索以精原细胞作为载体建立带乙肝病毒X基因的树鼩转基因动物模型的可行性.方法:首先构建带乙肝病毒pcDNA3.1- HBx基因的表达载体;然后以脂质体2000包被之.将此复合物以微量注射器注入8只雄性树鼩之睾丸组织中,1周后重复注射1次.注射后6周,使处理过的雄树鼩与雌性树鼩交配.在仔树鼩出生后1个月,取其肝组织作活检,并以多聚酶链式反应(PCR)对其DNA表达进行鉴定,观察其内是否存在乙肝病毒X基因.结果:处理的8只雄性树鼩中,6只具有生育能力,产仔树鼩11只.其中,1只仔树鼩乙肝病毒X基因阳性(阳性率9.09%).基因测序结果证实为乙肝病毒X基因.病理检查结果显示仔树鼩肝组织的改变包括:肝细胞坏死、慢性炎细胞在汇管区的浸润.结论:以精原干细胞作为载体,建立带乙肝病毒X基因的树鼩转基因动物模型是可行的.

ESTABLISHMENT OF TRANSGENE ANIMAL MODEL OF TREE SHREW WITH HBV X GENE BY USING THE SPERMATOGONIAL CELLS AS THE MEDIATING VECTOR

Objective:To investigate the possibility of using the spermatogonial cells as a vector to establish trans-gene animal model of tree shrew with HBV X gene.Methods:Expression vector with pcDNA3.1-HBx of HBV gene was constructed.Micro-volume injector was used to directly inject the recombinant of HBV pcDNA3.1-X packed by lipofetamine 2000 into the testicle tissue of 8 male tree shrews.Repeating the injection after one week.Six weeks after the respectively injections,the treated male tree shrews were paired with the females.Live biopsies were performed on the baby tree shrews after one month of their birth,and polymerase chain reaction was applied to identify the HBV X gene expression.Result:6 of the 8 treated male tree shrews showed the generational ability,and 11 baby tree shrews were born.1 of the 11 baby shrews was HBV X gene positive.The sequence was confirmed as HBV X gene.Pathological findings included necrosis of hepatic cells,chronic inflammatory cell infiltration in perivascular cuffings.Conclusion:It is practical to apply the spermatogonial cells as the vector to establish the trans-gene animal model of tree shrew with HBV X gene.