EFFECTS OF IN-VIVO ADVENTITIAL EXPRESSION OF RECOMBINANT ENDOTHELIAL NITRIC-OXIDE SYNTHASE GENE IN CEREBRAL-ARTERIES

Nitric oxide (NO), synthesized from L-arginine by NO synthases (NOS), plays an essential role in the regulation of cerebrovascular tone, Ade noviral vectors have been widely used to transfer recombinant genes to different vascular beds, To determine whether the recombinant endothe lial NOS (eNOS) gene can be delivered in vivo to the adventitia of cer ebral arteries and functionally expressed, a replication-incompetent a denoviral vector encoding eNOS gene (AdCMVNOS) or beta-galactosidase r eporter gene (AdCMVLacZ) was injected into canine cerebrospinal fluid (CSF) via the cisterna magna (final viral titer in CSF, 10(9) pfu/ml), Adventitial transgene expression was demonstrated 24 h later by beta- galactosidase histochemistry and quantification, eNOS immunohistochemi stry, and Western blot analysis of recombinant eNOS. Electron microsco py immunogold labeling indicated that recombinant eNOS protein was exp ressed in adventitial fibroblasts, In AdCMVNOS-transduced arteries, ba sal cGMP production and bradykinin-induced relaxations were significan tly augmented when compared with AdCMVLacZ-transduced vessels (P < 0.0 5), The increased receptor-mediated relaxations and cGMP production we re inhibited by eNOS inhibitors, In addition, the increase in cGMP pro duction was reversed in the absence of calcium, suggesting that the in creased NO production did not result from inducible NOS expression, Th e present study demonstrates the successful in vivo transfer and funct ional expression of recombinant eNOS gene in large cerebral arteries, It also suggests that perivascular eNOS gene delivery via the CSF is a feasible approach that does not require interruption of cerebral bloo d flow.