Insulin-like Growth Factor-1 Gene Delivery May Enhance the Proliferation of Human Corpus Cavernosal Smooth Muscle Cells
Received 17 August 2009; accepted 8 December 2009. published online 08 March 2010. Corrected Proof
Objectives
To investigate the effects of IGF-1 gene transfer in human corpus cavernosal smooth muscle cells (HCCSMCs) ex vivo. Insulin-like growth factor-1 (IGF-1) promotes the proliferation of penile cavernous smooth muscle cells in the rats.
Methods
A plasmid expressing human IGF-1 was constructed by subcloning hIGF-1 into pcDNA3.1 vector. HCCSMCs were harvested from 3 impotent patients and cultured in vitro. The cultured smooth muscle cells were identified by immunofluorescent staining. RNA was extracted from HCCSMCs, and the gene expression of IGF-1 was determined by reverse transcription-polymerase chain reaction. Cell growth was examined by use of a novel cell proliferation assay based on the bioreduction of the fluorescent dye Alamar blue. The subcloned product was transfected into HCCSMCs. Western blotting and immunoassay were performed 2 days after transfection to evaluate the transfection efficiency.
Results
Endogenous IGF-1 mRNA expression was detected by reverse transcription-polymerase chain reaction analysis of total RNA extracted from cultured HCCSMCs. Increased proliferation of HCCSMCs in vitro was observed with exogenous treatment with IGF-1 (100 ng/mL) in a dose-dependent manner. Exogenous IGF-1 gene transfer to cultured HCCSMCs enhanced IGF-1 protein expression compared with the control, and the expression level peaked at 4 days after transfection and decreased slowly thereafter. Secretion of IGF-1 from transfected HCCSMCs induced cellular proliferation.
Conclusions
IGF-1 gene transfer into HCCSMCs enhanced cellular proliferation, which was mediated by secretion of IGF-1. Our results suggest that IGF-1 gene therapy may be applied to corpus cavernosum regeneration.
aDepartment of Urology, Chonnam National University Medical School, Sexual Medicine Research Center, Chonnam National University, Gwangju, Republic of Korea
bDepartment of Biomedical Science, Chonnam National University Medical School, Sexual Medicine Research Center, Chonnam National University, Gwangju, Republic of Korea
cDepartment of Sexual Medicine Research Center, Chonnam National University Medical School, Sexual Medicine Research Center, Chonnam National University, Gwangju, Republic of Korea
Reprint requests: Kwangsung Park, M.D., Ph.D., Department of Urology, Chonnam National University, Medical School, Sexual Medicine Research Center, Chonnam National University, 8 Hak-dong, Dong-Gu, Gwangju 501–757, Republic of Korea
ABSTRACT