Expression and function of a recombinant endothelial nitric oxide synthase gene in porcine coronary arteries

David G. Cable, Timothy O'Brien, Iftikhar J. Kullo, Robert S. Schwartz, Hartzell V. Schaff, Vincent J. Pompili

Research output: Contribution to journalArticle

31 Scopus citations

Abstract

Objectives: Direct gene transfer of exogenous nitric oxide synthase, with the subsequent increase in nitric oxide production, could represent a potential therapeutic strategy in the treatment of vascular proliferative disorders. The goal of the present study was to determine if porcine coronary arteries could be transduced with an adenoviral vector encoding endothelial nitric oxide synthase (Ad. CMVeNOS) resulting in functional expression. Methods and Results: Segments of porcine right coronary artery were exposed for 1 h at 37°C to either replication-deficient adenovirus encoding bovine endothelial nitric oxide synthase (Ad. CMVeNOS, 5 x 109 pfu/ml) or control adenovirus encoding Escherichia coli β-galactosidase (Ad. CMVLacZ, 5 x 109 pfu/ml). Immunohistochemistry with a monoclonal antibody specific for nitric oxide synthase (NOS) demonstrated recombinant gene expression in both the endothelial and adventitial layers of Ad.CMVeNOS transduced coronaries with only endogenous NOS confirmed in the endothelium of Ad. CMVLacZ arteries. Coronary arteries transduced with Ad.CMVeNOS yielded 517 ± 110 (mean ± S.E.M.) nM/ng nitrite while vessels transduced with Ad.CMVLacZ yielded 126 ± 84 nM/ng (P < 0.05, n = 6). Isometric tension recording, following prostaglandin F(2α) constriction, documented a reduced tension in Ad.CMVeNOS transduced coronaries, compared to matched Ad.CMVLacZ coronaries (6.10 ± 1.0g g vs. 8.45 ± 1.19 g, respectively, P = 0.05, n = 8). This tension differential was eliminated with prior incubation in N(G)-monomethyl-L- arginine (L-NMMA, 10-4 M). The EC50 for calcium ionophore relaxation of Ad.CMVeNOS coronary arteries was reduced compared to Ad. CMVLacZ (- 7.90 ± 0.03 logM vs. - 7.26 ± 0.11 logM, respectively, P < 0.05, n = 8). Conclusions: These studies demonstrate successful transfer of endothelial nitric oxide synthase into porcine coronary arteries as verified by histochemical localization of recombinant protein with an increase of nitric oxide release as demonstrated by enhanced nitrite production and an alteration in vasomotor function.

Original languageEnglish (US)
Pages (from-to)553-559
Number of pages7
JournalCardiovascular research
Volume35
Issue number3
DOIs
StatePublished - Sep 1 1997

Keywords

  • Adenoviral vector
  • Gene therapy
  • Nitric oxide synthase
  • Porcine coronary artery
  • Vasomotor function

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine
  • Physiology (medical)

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