RESUMO
Recombinant adeno-associated (AAV) viruses have unique properties, which make them ideal vectors for gene transfer targeting the myocardium. Numerous serotypes of AAV have been identified with variable tropisms towards cardiac tissue. In the present study, we investigated the time course of expression of eight different AAV serotypes in rat myocardium and the nature of the immunity against these serotypes. We first assessed whether neutralizing antibodies (NAb) were present for any of the serotype in the rats. We injected 100 microl of each AAV 1-8 serotype (10(12) DNAse resistant particles/ml), encoding LacZ gene, into the apical wall of rat myocardium. At 1, 4, 12 and 24 weeks after gene delivery, the animals were killed and beta-galactosidase (beta-gal) activity was assessed by luminometry. Additionally, LacZ genomic copies and AAV capsids copies were measured through standard polymerase chain reaction analysis and cryo-sections from the area of viral injection were stained for X-gal detection at the same time points. No NAbs were detected against any of AAV serotypes. At all the time points studied, AAV1, 6 and 8 demonstrated the highest efficiency in transducing rat hearts in vivo. Parallel to the results with beta-gal activity, the highest levels LacZ and AAV DNA genomic copies were with AAV1, 6 and 8. The positive X-gal staining depicted by these serotypes confirmed these results. These results indicate that among the various AAV serotypes, AAV1, 6 and 8 have differential tropism for the heart unaffected by pre-existing NAb in the rat. Although AAV 1 and 6 vectors induced rapid and robust expression and reach a plateau at 4 weeks, AAV 8 continued increasing until the end of the study. AAV 2, 5 and 7 vectors were slower to induce expression of the reporter gene, but did reach levels of expression comparable to AAV1 and AAV6 vectors after 3 months.
