IN-VIVO GENE-TRANSFER INTO RAT ARTERIAL-WALLS WITH NOVEL ADENOASSOCIATED VIRUS VECTORS

Purpose: We studied the ability of recombinant adeno-associated virus (rAAV) vectors to achieve gene transfer in vivo to intact rat carotid arteries. Methods: Isolated segments of uninjured rat carotid arteries were incubated with (1) rAAV vectors that expressed a beta-galactosid ase gene, (2) a related vector with no promoter, or (3) a normal salin e solution. Gene transfer was evaluated with in situ polymerase chain reaction (PCR). Transgene expression was assessed at intervals that ra nged from 24 hours to 2 months by measurement of beta-galactosidase ac tivity and protein mass in tissue extracts with fluorometric and enzym e-linked immunosorbent assays, respectively. Dose dependence of expres sion was determined for virus concentrations that ranged from 5 x 10(4 ) to 5 x 10(5) infectious units (iu)/ml. Results: Light microscopic an alysis of in situ PCR-stained histologic sections of transduced vessel walls showed approximately 90% of intimal and medial cell nuclei cont ained the beta-galactosidase gene, compared with none in control arter ies. In vivo beta-galactosidase expression was (1) highest 24 hours af ter gene transfer, (2) elevated for 1 month, and (3) dose responsive. Conclusions: rAAV vectors can mediate focal gene transfer into the int act rat carotid artery with detectable levels of transgene expression for 1 month and are potentially useful agents for in vivo gene transfe r into intact arteries.