Construction of recombinant adenoviral vector carrying human tissue inhibitor of metalloproteinase-1 gene and its expression in vitro.
Hepatobiliary Pancreat Dis Int
; 4(2): 259-64, 2005 May.
Article
en En
| MEDLINE
| ID: mdl-15908326
ABSTRACT
BACKGROUND:
Overexpression of human tissue inhibitors of metalloproteinase-1 (TIMP-1) may play an antitumor role in inhibiting hepatocellular carcinoma (HCC) growth and progression. The aim of the study was to construct a recombinant adenovirus vector carrying hTIMP-1 cDNA for liver gene therapy and observe its expression in vitro.METHODS:
The full-length cDNA of hTIMP-1 was positively cloned into the adenoviral shuttle vector pAdTrack-CMV, and then cotransformed into competent BJ5183 cells with the adenoviral backbone plasmid pAdEasy-1. Thus, a recombinant adenovirus AdhTIMP-1 containing full-length cDNA of hTIMP-1 was generated by homologous recombination in E. coli. AdhTIMP-1 was then packaged and amplified in adenoviral packaging cells, or human embryonic kidney 293 cells. The viral titer was checked by green fluorescent protein (GFP), and the expression of hTIMP-1 in vitro was detected by the techniques of Western blot and RT-PCR.RESULTS:
The recombinant adenovirus vector carrying hTIMP-1 was constructed and confirmed by restriction endonuclease analysis and DNA sequence analysis. The transcription of TIMP-1 mRNA in 293 cells was checked by RT-PCR and TIMP-1 protein could be detected in the culture by Western blot analysis.CONCLUSION:
The successful construction of recombinant adenoviral vector carrying human TIMP-1 and the effective expression in vitro has laid a foundation for further study of its antitumor function and may pave the way for future application in liver gene therapy.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Regulación Viral de la Expresión Génica
/
Técnicas de Transferencia de Gen
/
Inhibidor Tisular de Metaloproteinasa-3
/
Hepatocitos
Tipo de estudio:
Diagnostic_studies
/
Etiology_studies
Límite:
Humans
Idioma:
En
Revista:
Hepatobiliary Pancreat Dis Int
Asunto de la revista:
GASTROENTEROLOGIA
Año:
2005
Tipo del documento:
Article
País de afiliación:
China