Development of New Gene Therapy for Erectile Dysfunction Using Antisense cDNA for Type V Phosphodiesterase Gene / 대한남성과학회지

ABSTRACT

PURPOSE: We examined the feasibility of gene therapy for erectile dysfunction using cultured human corpus cavernosal smooth muscle cells. MATERIALS AND METHODS: The vector construct was designed to contain a fusion gene of enhanced green fluorescent protein (gfp) and beta-galactosidase (lacZ) which was under control of CMV promoter. Cells within the second passage were transfected with the vector DNA only and vector DNA containing a part of cDNA in an antisense orientation for human type V phosphodiesterase (PDE) gene using lipofection. Reporter gene expressions were investigated by fluorescence microscopy and X-gal staining at 24-hour interval. Effects of gene transfer of type V PDE antisense cDNA were investigated after 48 hours of gene transfection using the RT-PCR for type V PDE gene and measurement of intracellular cGMP level treated by sodium nitroprusside (SNP), NO-donor, of various concentrations. RESULTS: Expressions of gfp and lacZ were observed for upto 72 hours after gene transfection. Results from RT-PCR analysis also confirmed the gene expression at the transcriptional level. Type V PDE mRNA expression was significantly inhibited and magnitude of cGMP increase was significantly enhanced by gene transfer of antisense cDNA for type V PDE gene compared with non-transfectant control cells. CONCLUSIONS: Our results demonstrate that the liposome-mediated gene transfer was shown to be effective in corpus cavernosal smooth muscle cells. Gene transfer of antisense cDNA for type V PDE gene effectively inhibited the expression of type V PDE gene at the transcriptional and translational levels, suggesting that this newly developed gene transfer system may be a potential gene therapy in the treatment of erectile dysfunction.