Insulin-like growth factor-1 gene delivery may enhance the proliferation of human corpus cavernosal smooth muscle cells

Abstract

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.