[Expression and effect of hCGRP recombinant adeno-associated virus in primary cultured corporal cavernosum smooth muscle cells of the rat in vitro].
OBJECTIVE
To observe the expression and effect of human calcitonin gene-related peptide (hCGRP) gene mediated by recombinant adeno-associated virus (rAAV) in primary cultured corporal cavernosum smooth muscle cells of the rat and explore the possibility of using CGRP gene for gene therapy in erectile dysfunction.


METHODS
The primary cultured corporal cavernosum smooth muscle cells of the rat were randomly divided into 4 groups and infected with recombinant virus VssHGCMV-hCGRP, VssHGCMV, VssC-MV-GFP and the untreated, respectively. CGRP-like immunoreactivity was measured by protein dot blot assay in the 24 h-culture medium, and intracellular cAMP and cGMP levels in the cultured cells were also determined using radioimmunoassay to ascertain bioactivity of transduced CGRP.


RESULTS
The exogenous gene was transferred into primary corporal cavernosum smooth muscle cells by VssHGCMV-hCGRP infection and efficiently expressed. Compared with the control group, intracellular cAMP level in the cell infected by VssHGCMV-hCGRP was significantly increased (48.7 +/- 1.1 nmol/L vs 7.8 +/- 1.4 nmol/L, P < 0.01), whereas cGMP level remained unchanged in two groups, and CGRP-like immunoreactivity was also detected in the culture medium infected by VssHGCMV- hCGRP.


CONCLUSIONS
The system of secretory expressing bioactive peptide rAAV mediated gene transfer may be used to express efficiently exogenous gene in corporal cavernosum smooth muscle cells and affect cAMP level in the corporal cavernosum smooth muscle cells of the rat.
Xing
Jun-ping
JP
Department of Urology, the First Affiliated Hospital, Xián Jiaotong University, Xián, Shaanxi 710061, China. xingjp-mr@vip.sina.com
Sun
Jian-hua
JH
Cui
Xian-feng
XF
Qiu
Shu-dong
SD
D000818:Animals; D015740:Calcitonin Gene-Related Peptide / Q000096:biosynthesis* / Q000235:genetics; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism; D000229:Dependovirus / Q000235:genetics*; D008297:Male; D009130:Muscle, Smooth / Q000166:cytology / Q000378:metabolism*; D010413:Penis / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011995:Recombination, Genetic; D014162:Transfection
D000242:Cyclic AMP; D006152:Cyclic GMP; D015740:Calcitonin Gene-Related Peptide