Construction of adenovirus vector containing hIL-12 gene and its effects on the proliferation and TGF-β expression of Hep3B cells

Objective:To construct a recombinant adenovirus expression vector carring human IL-12 gene,and investigate its effects on the proliferation and TGF-β expression of Hep3B cells.Methods:The gene of hIL-12 was amplified with PCR and the shuttle plasmid pAdTrack-IL-12 was constructed.The shuttle plasmid was transformed into Adeasier cell after linearization with Pme I for homologous recombination in E.coli BJ5183.After digested with Pac I,the recombinant vector was transfected into AD293 cells to package adenovirus pAd-IL-12.The expression of IL-12 was detected by RT-PCR and Western blot in human hepatoma cell Hep3B infected with pAd-IL-12 virus.The proliferation of Hep3B was evaluated by MTT,and the expression of TGF-β was detected with RT-PCR.Results:The recombinant adenovirus plasmid containing IL-12 was exactly constructed.Recombinant adenovirus pAd-IL-12 were successfully constructed which could infect Hep3B cell efficiently.In Hep3B infected with pAd-IL-12,high expressions of IL-12 mRNA and protein were confirmed by PCR and Western blot,and both of the proliferation and TGF-β expression of Hep3B were significantly reduced(both P0.05) compared with control group.Conclusions: The results demonstrate that recombinant adenovirus containing IL-12 were constructed successfully and IL-12 could suppress Hep3B proliferation with the decline of TGF-β gene,which provides a foundation for further study of IL-12 in tumor treatment.