Efficient Expression Of The Interleukin-12 Gene Mediated By A Hydrodynamics-based Procedure In Mice

Interleukin-12 (IL-12) is a cytokine with powerful antitumor effects. The anti-tumor mechanism involves inducing production of IFN-γand NO, enhancing the cellular immune function and inhibiting tumor angiogenesis. IL-12 gene transfer mediated by viral vectors is highly effective and results in the retardation of tumor growth or eradication of the tumor. Unfortunately, the production, amplification and purification of viral vectors is laborious. Moreover, the antiviral immunity induced by viral vectors not only results in side effects, but also precludes re-administration of the vector. The naked DNA of non-viral vectors is an attractive alternative because of its advantages including safety, simplicity, and absent or low immunogenicity. However, the low transfection efficiency limits its application.In our study, we used a novel and effective non-viral vector gene transfer technique, the hydrodynamics-based procedure for naked DNA to investigate the kinetic expression and distribution of palsmid DNA encoding the IL-12 gene. The effect of repeated injection of plasmid DNA on production of IL-12, IFN-γand NO were also studied. The aim of this research was to evaluate the transfer efficiency for possible application of the method to IL-12 gene transfer in cancer therapy.Methods: Varying doses of pCMV-mIL-12 plasmid were injected into Balb/c mice by a hydrodynamics-based procedure. Thereafter, the blood and organs were collected at specific time interval. The kinetic and plasmid DNA dose-dependent expression of IL-12 and IFN-γin serum were determined by ELISA. NO production was measured using a commercial kit. The distribution and expression of pCMV-mIL-12 were assayed by measuring DNA, RNA and protein levels using PCR, RT-PCR and immunohistochemical staining. To investigate the expression of some cytokines after plasmid re-administration, the same amount of pCMV-mIL-12 was repeatedly injected into the mice 7 days, 14 days and 30 days respectively after the initial injection. The levels of IL-12,...