Recombinant adenovirus with human indoleamine-2,3- dioxygenase and hepatitis B virus preS was constructed and expressed in HepG2 cells

Background Indoleamine-2,3-dioxygenase （IDO） is proven to suppress hepatitis B virus （HBV） specific immune response and depletion of IDO may be a useful approach for HBV therapy. To test this concept, we constructed recombinant adenovirus with human IDO and HBV preS, which would form the basis for f...

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Bibliographic Details
Published in:Chinese medical journal 2011-10, Vol.124 (19), p.3159-3163
Main Authors: Chen, Yong-Bing, Shi, Xian-Jie, Lu, Gang, Nie, Hong-Feng, Shen, Xiao-Qing, Yu, Cong-Hui, Gong, Jian-Ping
Format: Article
Language:English
Subjects:
Adenoviridae - genetics
Cloning, Organism
Genetic Vectors
Hep G2 Cells
Hepatitis B virus - genetics
HepG2细胞
Humans
Indoleamine-Pyrrole 2,3,-Dioxygenase - genetics
Recombination, Genetic
Western印迹
人类
双加氧酶
吲哚
肝炎病毒
重组腺病毒
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Summary:Background Indoleamine-2,3-dioxygenase （IDO） is proven to suppress hepatitis B virus （HBV） specific immune response and depletion of IDO may be a useful approach for HBV therapy. To test this concept, we constructed recombinant adenovirus with human IDO and HBV preS, which would form the basis for future in vivo experiments.Methods The fragment of human IDO and HBV preS cDNA were subcloned into multiple cloning sites in an adenoviral vector system containing two cytomegalovirus （CMV） promoters. Recombination was conducted in the Escherichia coli BJ5183. The recombinant adenovirus containing hlDO gene and HBVpreS gene was packaged and amplified in 293 cells.Integration was confirmed by polymerase chain reaction as well as the quantification of viral titers. HepG2 cells were infected with the recombinant adenovirus and mRNA and protein specific for hlDO and HBVpreS was detected by RT-PCR and Western blotting respectively.Results The recombinant adenovirus was produced successfully. Its titer was 2.5x109 efu/ml. IDO and HBVpreS mRNA as well as the encoded proteins could be found in transfected HepG2 cells, but not in control HepG2 cells.Conclusion The transfer of hlDO-HBVpreS with double-promoter adenoviral vector was efficient. The recombinant adenovirus with hlDO and HBVpreS would provide the experimental basis for future studies.
ISSN:0366-6999
2542-5641
DOI:10.3760/cma.j.issn.0366-6999.2011.19.036