Attenuation of Hypertension and Heart Hypertrophy by Recombinant Adeno-Associated Virus Delivering Angiotensinogen Antisense (rAAV-AGT-AS)
Introduction: We have previously shown that injection of antisense oligonucleotides (AS-ODNs) or plasmid producing antisense directed against angiotensinogen (AGT) mRNA, lowers BP in spontaneously hypertensive rats (SHR). The effects with ODNs, however, last ∼7 days. For more prolonged effects we have developed a recombinant adeno-associated viral vector (rAAV) to deliver AS-DNA targeted to AGT. Methods: Since captopril has been shown to inhibit development of hypertension in SHR, we hypothesized that AGT inhibition would prevent hypertension. Five day old SHR were injected with a single dose of 109 ipf (25 μl), intra-cardiacly, with AGT-AS-DNA (n=10) or control sense-AGT-DNA (n=6). Systolic BP was measured weekly from 6 and 26 weeks of age. At 14 weeks and 7 months, tissue DNA was analyzed for presence of the transgene. At 26 weeks, the left ventricle of the heart was dissected, weighed and compared to body weight, to determine the degree of hypertrophy (n=5/group). Every three weeks plasma was sampled for RIA of AGT. Results: The rats injected with rAAV-AGT-AS had lower BP than the control group from 10 weeks of age. The difference was significant at 10,11,12 and 13 weeks of age (p<0.05). The maximum difference was at 12 weeks (172 ± 7 for control versus 150 ± 8 mmHg for the treated group). Plasma AGT levels in the treated group did not correlate to BP, whereas plasma AGT in controls correlated to BP (p<0.05). The treated group had significantly lower BP than controls at 6 months (p<0.05). The presence of AS-transgene was confirmed by nested PCR on the DNA isolated from the heart, kidney and liver. The rAAV-AGT-AS significantly decreased the left ventricular hypertrophy measured at 26 weeks (LV/BW ratio: 3.18 ± 0.02 for control versus 2.92 ± 0.04 for the treated group, p<0.001). No signs of toxicity or body weight loss were observed. Conclusion: The study shows that AAV is a stable, effective vector to deliver AGT antisense and that AGT is an important factor in the development and maintenance of high BP in SHR. We conclude that the rAAV-AGT-AS vector shows promise for long-term attenuation of BP and heart hypertrophy.