IN VIVO eNOS gene transfer in cerebral arteries via cerebrospinal fluid

To determine if recombinant endothelial nitric oxide synthase (eNOS) gene can be expressed in cerebral arteries following in vivo gene transfer, a replication-incompetent adenoviral vector encoding β-galactosidase reporter gene (AdCMVLacZ), or eNOS gene (AdCMVNOS), was injected into canine cerebrospinal fluid (CSF) via cisterna magna (final viral titer in CSF 10⁹ pfu/ml). Dog heads were maintained in a nose-up position for 30 minutes following the injections. After 24 and 72 hours, X-gal histochemistry and eNOS immunohistochemistry revealed adventitial transgene expression in all major cerebral arteries. Electron microscopy immunogold labeling further indicated that recombinant eNOS proteins were mainly expressed in caveolae of the adventitial fibroblasts. While the contractions to DTP (10⁷-10³ mol/mL), relaxations to bradykinin (10¹⁰-10⁶ mol/mL), and cGMP production were not affected in arteries transduced with AdCMVLacZ reporter gene when compared to non-transduced controls, the bradykinin-induced relaxations were significantly enhanced in AdCMVNOS-transduced vessels with significant increase in cGMP (n=5, p<0.05, RM-ANOVA). The increase in cGMP was abolished in the absence of calcium and reversed by L-NMMA. The present study represents the first successful in vivo eNOS gene transfer in cerebral arteries, and suggests that adventitial gene transfer in CSF via cisterna magna is a feasible approach.