Human endothelial nitric oxide synthase gene delivery promotes angiogenesis in a rat model of hindlimb ischemia.
OBJECTIVE
Endothelium-derived NO has been shown to mediate the mitogenic effect of vascular endothelial growth factor on cultured microvascular endothelium. To evaluate the role of endothelial NO synthase (eNOS) in angiogenesis in the ischemic hindlimb, we engineered an adenovirus containing human eNOS cDNA.


RESULTS
After gene transfer, expression of eNOS in cultured cells was detected by increased intracellular cGMP and nitrate/nitrite levels and NO synthase activity. Adenovirus containing either the eNOS or luciferase gene was injected into the adductor muscle of rat hindlimbs immediately after femoral artery removal. Human eNOS protein was detected throughout the course of the experiment by immunostaining. Significant increases in blood perfusion were monitored by laser Doppler imaging from 2 to 4 weeks after gene delivery in the ischemic hindlimb of rats receiving eNOS compared with control rats receiving the reporter gene. An increase in regional blood flow was also detected after eNOS gene transfer by a fluorescent microsphere assay. eNOS gene delivery in the ischemic hindlimb resulted in significant increases in intracellular cGMP levels and in capillary density identified by anti-CD-31 immunostaining. Angiogenesis was further confirmed in mice after eNOS gene transfer by increased hemoglobin content in Matrigel implants.


CONCLUSIONS
Taken together, these results indicate that eNOS enhances angiogenesis and raises the potential of eNOS gene transfer for modulation of vascular insufficiency.
Smith
Robert S
RS
Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston 29425-2211, USA.
Lin
Kuei-Fu
KF
Agata
Jun
J
Chao
Lee
L
Chao
Julie
J
D000256:Adenoviridae; D000818:Animals; D002196:Capillaries / Q000502:physiology; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000096:biosynthesis; D004279:DNA, Viral; D004730:Endothelium, Vascular / Q000201:enzymology*; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006454:Hemoglobins / Q000032:analysis; D006614:Hindlimb / Q000098:blood supply; D007511:Ischemia / Q000201:enzymology / Q000503:physiopathology*; D017078:Laser-Doppler Flowmetry; D008297:Male; D051379:Mice; D023421:Models, Animal; D018919:Neovascularization, Physiologic; D019001:Nitric Oxide Synthase / Q000235:genetics* / Q000502:physiology*; D010957:Plasmids; D051381:Rats; D012039:Regional Blood Flow
D004279:DNA, Viral; D006454:Hemoglobins; D019001:Nitric Oxide Synthase; D006152:Cyclic GMP