Korean J Androl.
2001 Aug;19(2):89-97.
Development of New Gene Therapy for Erectile Dysfunction Using Antisense cDNA for Type V Phosphodiesterase Gene
- Affiliations
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- 1Department of Urology, Seoul National University College of Medicine, Seoul, Korea. swkim@sun.ac.kr
Abstract
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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.
