Abstract 3669: Transcoronary Gene Transfer of Serca2a Enhances Coronary Blood Flow through an Increase of eNOS Activity in Endothelial Cells
Congestive heart failure (CHF) is associated with impaired endothelium-dependent nitric oxide (NO)-mediated vasodilatation. Clinical trials using AAV1-SERCA2a myocardial gene delivery in CHF patients are currently underway. We hypothesized that transcoronary SERCA2a gene transfer can increase NO production by endothelial cells (EC). To assess the effects of SERCA2a gene transfer on coronary flow and endothelial function in vivo, we used a preclinical porcine volume-overload heart failure model (severe mitral valve regurgitation (MR)). Two months after MR creation, animals underwent intracoronary injection of AAV1-SERCA2a or saline. Four months post MR creation, the average coronary flow peak velocity (APV) was measured in the mid portion of the left anterior descending artery (LAD), and adjusted to the diameter of coronary artery and the weight of the left ventricle. Compared to saline treated animals, SERCA2a has significantly increased the adjusted coronary flow (ml/min/g) (0.467 ± 0.09, n=6 vs. 0.886 ± 0.18, n=7; P<0.01) but no difference was observed compared to sham operated animals (1.003 ± 0.2137, n=4; P=NS). Confocal immunofluorescence of swine coronary artery sections demonstrated that SERCA2a is expressed in endothelial and vascular smooth muscle cells; however, eNOS was expressed only in EC in all animal groups. Western blot analyses showed down-regulation of SERCA2a and eNOS expression in LAD, LCA and RCA from failing saline-injected animals. In coronary arteries from SERCA2a-transduced animals, expression levels of SERCA2a and eNOS were similar to those in sham-operated animals. To further investigate the effect of SERCA2a gene transfer on endothelial function we used cultured human coronary artery EC. RT-PCR analysis, western blot and e-NOS promoter/ luciferase reporter assay have shown that SERCA2a gene transfer increased expression and Ser-1177 phosphorylation of e-NOS. Furthermore, cGMP production was increased in SERCA2a-infected EC, confirming its possible functional effect on eNOS activity. In conclusion, our results demonstrate that transcoronary SERCA2a gene transfer improves endothelial function in failing hearts through regulation of eNOS expression and activity.