RRC ID 12370
Author Yoshida M, Iwashita H, Otani M, Masunaga K, Inadome A.
Title Delivery of DNA into bladder via electroporation.
Journal Methods Mol. Biol.
Abstract The possibility of in vivo gene transfer into the rat bladder by electroporation (EP) was evaluated. The bladder was exposed through an abdominal midline incision in 8-week-old male rats. Plasmid DNA of marker genes, green fluorescent protein (GFP) and luciferase, and the neuronal nitric oxide synthase (nNOS) gene were then injected into the subserosal space of the bladder and EP was applied. At 72 h after gene transfer, GFP and luciferase were assayed in the isolated bladder, and immunohistochemical staining was used to detect nNOS. NOx released from isolated bladder strips was also assessed using microdialysis procedure. From the luciferase assay, 45 V, 1 Hz, 50 ms, and 8 pulses were selected as the optimum conditions for EP. Bladder specimens with GFP genes injected by EP showed numerous bright sites of GFP expression in the smooth-muscle layer. In rats with the nNOS gene injected by EP, there was marked nNOS immunoreactivity, and NOx released from bladder strips was significantly greater than that in the control groups. These results suggest that EP is a useful technique for in vivo gene transfer into rat bladder smooth muscles, and that the nNOS gene transferred by this procedure functionally expresses and contributes to NO production.
Volume 423
Pages 249-57
Published 2008
DOI 10.1007/978-1-59745-194-9_18
PMID 18370204
MeSH Animals DNA, Recombinant / administration & dosage* DNA, Recombinant / genetics Electrochemotherapy / methods* Electroporation / methods Gene Expression Gene Transfer Techniques Genes, Reporter Genetic Therapy / methods* Genetic Vectors Green Fluorescent Proteins / genetics Green Fluorescent Proteins / metabolism Immunohistochemistry Luciferases / genetics Luciferases / metabolism Male Mice Nitric Oxide / metabolism Nitric Oxide Synthase Type I / genetics Nitric Oxide Synthase Type I / metabolism Plasmids / administration & dosage Plasmids / genetics Rats Rats, Sprague-Dawley Recombinant Proteins / genetics Recombinant Proteins / metabolism Urinary Bladder / metabolism*
Resource
Human and Animal Cells