Jonathan A. Hata

Enhancement of cardiac function after adenoviral-mediated in vivo intracoronary β2-adrenergic receptor gene delivery

Exogenous gene delivery to alter the function of the heart is a potential novel therapeutic strategy for treatment of cardiovascular diseases such as heart failure (HF). Before gene therapy approaches to alter cardiac function can be realized, efficient and reproducible in vivo gene techniques must be established to efficiently transfer transgenes globally to the myocardium. We have been testing the hypothesis that genetic manipulation of the myocardial beta-adrenergic receptor (beta-AR) system, which is impaired in HF, can enhance cardiac function. We have delivered adenoviral transgenes, including the human beta2-AR (Adeno-beta2AR), to the myocardium of rabbits using an intracoronary approach. Catheter-mediated Adeno-beta2AR delivery produced diffuse multichamber myocardial expression, peaking 1 week after gene transfer. A total of 5 x 10(11) viral particles of Adeno-beta2AR reproducibly produced 5- to 10-fold beta-AR overexpression in the heart, which, at 7 and 21 days after delivery, resulted in increased in vivo hemodynamic function compared with control rabbits that received an empty adenovirus. Several physiological parameters, including dP/dtmax as a measure of contractility, were significantly enhanced basally and showed increased responsiveness to the beta-agonist isoproterenol. Our results demonstrate that global myocardial in vivo gene delivery is possible and that genetic manipulation of beta-AR density can result in enhanced cardiac performance. Thus, replacement of lost receptors seen in HF may represent novel inotropic therapy.

Targeted β-Adrenergic Receptor Kinase (βARK1) Inhibition by Gene Transfer in Failing Human Hearts

Background—Failing human myocardium is characterized by an attenuated contractile response to &bgr;-adrenergic receptor (&bgr;AR) stimulation due to changes in this signaling cascade, including increased expression and activity of the &bgr;-adrenergic receptor kinase (&bgr;ARK1). This leads to desensitization and downregulation of &bgr;ARs. Previously, expression of a peptide inhibitor of &bgr;ARK1 (&bgr;ARKct) has proven beneficial in several animal models of heart failure (HF). Methods and Results—To test the hypothesis that inhibition of &bgr;ARK1 could improve &bgr;-adrenergic signaling and contractile function in failing human myocytes, the &bgr;ARKct was expressed via adenovirus-mediated (Ad&bgr;ARKct) gene transfer in ventricular myocytes isolated from hearts explanted from 10 patients with end-stage HF undergoing cardiac transplantation. Ad&bgr;ARKct also contained the marker gene, green fluorescent protein, and successful gene transfer was confirmed via fluorescence and immunoblotting. Compared with uninfected failing myocytes (control), the velocities of both contraction and relaxation in the Ad&bgr;ARKct-treated cells were increased in response to the &bgr;-agonist isoproterenol (contraction: 57.5±6.6% versus 37.0±4.2% shortening per second, P <0.05; relaxation: 43.8±5.5% versus 27.5±3.9% lengthening per second, P <0.05). Fractional shortening was similarly enhanced (12.2±1.2% versus 8.0±0.9%, P <0.05). Finally, adenylyl cyclase activity in response to isoproterenol was also increased in Ad&bgr;ARKct-treated myocytes. Conclusions—These results demonstrate that as in animal models of HF, expression of the &bgr;ARKct can improve contractile function and &bgr;-adrenergic responsiveness in failing human myocytes. Thus, &bgr;ARK1 inhibition may represent a therapeutic strategy for human HF.

Impact of Mitral Valve Regurgitation Evaluated by Intraoperative Transesophageal Echocardiography on Long-Term Outcomes After Coronary Artery Bypass Grafting

Background—It is unclear if mild or moderate mitral valve regurgitation (MR) should be repaired at the time of coronary artery bypass grafting (CABG). We sought to determine the long-term effect of uncorrected MR, measured by intraoperative transesophageal echocardiography (TEE), in CABG patients. Methods and Results—Between May 1999 and September 2003, data were gathered for 3264 consecutive patients who underwent isolated CABG and had MR graded by intraoperative TEE. MR was graded on the following 5 levels: none, trace, mild, moderate, and severe. Patients who had severe MR or who underwent mitral valve surgery were eliminated from the analysis. The remaining patients were combined into the following 3 groups: none or trace, mild, and moderate MR. Preoperative and follow-up data were 99% complete. The median length of follow-up was 3.0 years. Multivariable analysis controlling for important preoperative risk factors was performed to determine predictors of death and death/hospitalization for heart failure. Increasing MR was a risk factor for death [hazard ratio (HR), 1.44; P<0.001] and death/heart failure hospitalization (HR, 1.34; P<0.01). When patients with moderate MR were eliminated from the analysis, mild MR was a risk factor for death (HR, 1.34; P=0.011) and death/hospitalization for heart failure (HR, 1.34; P<0.001). Conclusions—Even mild MR, identified by intraoperative TEE, predicts worse outcomes after CABG. Revascularization alone did not eliminate the negative long-term effects of mild MR. CABG patients with uncorrected mild or moderate MR are at increased risk for death and heart-failure hospitalization; consideration for surgical repair or more aggressive medical management and follow-up is warranted.

Restoration of β-Adrenergic Receptor Signaling and Contractile Function in Heart Failure by Disruption of the βARK1/Phosphoinositide 3-Kinase Complex

Background—Desensitization and downregulation of myocardial &bgr;-adrenergic receptors (&bgr;ARs) are initiated by the increase in &bgr;AR kinase 1 (&bgr;ARK1) levels. By interacting with &bgr;ARK1 through the phosphoinositide kinase (PIK) domain, phosphoinositide 3-kinase (PI3K) is targeted to agonist-stimulated &bgr;ARs, where it regulates endocytosis. We tested the hypothesis that inhibition of receptor-targeted PI3K activity would alter receptor trafficking and ameliorate &bgr;AR signaling, ultimately improving contractility of failing cardiomyocytes. Methods and Results—To competitively displace PI3K from &bgr;ARK1, we generated mice with cardiac-specific overexpression of the PIK domain. Seven-day isoproterenol administration in wild-type mice induced desensitization of &bgr;ARs and their redistribution from the plasma membrane to early and late endosomes. In contrast, transgenic PIK overexpression prevented the redistribution of &bgr;ARs away from the plasma membrane and preserved their responsiveness to agonist. We further tested whether PIK overexpression could normalize already established &bgr;AR abnormalities and ameliorate contractile dysfunction in a large animal model of heart failure induced by rapid ventricular pacing in pigs. Failing porcine hearts showed increased &bgr;ARK1-associated PI3K activity and marked desensitization and redistribution of &bgr;ARs to endosomal compartments. Importantly, adenoviral gene transfer of the PIK domain in failing pig myocytes resulted in reduced receptor-localized PI3K activity and restored to nearly normal agonist-stimulated cardiomyocyte contractility. Conclusions—These data indicate that the heart failure state is associated with a maladaptive redistribution of &bgr;ARs away from the plasma membrane that can be counteracted through a strategy that targets the &bgr;ARK1/PI3K complex.

Molecular β-adrenergic signaling abnormalities in failing rabbit hearts after infarction

We studied alterations in the beta-adrenergic receptor (beta-AR) system of rabbit hearts during the development of heart failure (HF) after myocardial infarction (MI) to determine whether the molecular beta-AR abnormalities associated with human HF exist in this animal model. Rabbit HF was established 3 wk after left circumflex coronary artery (LCX) ligation by in vivo physiological measurements, and molecular beta-AR signaling was examined in tissue and cultured ventricular myocytes. We found that there was a significant global reduction in beta-AR density by approximately 50% in both ventricles of MI animals compared with sham-operated control animals and that functional beta-AR coupling was significantly reduced. Importantly, as found in human HF, myocardial protein levels and activity of the beta-AR kinase (beta-ARK1) and Galphai were found to be significantly elevated in MI rabbits, suggesting that these molecules are contributing to myocardial dysfunction. Thus the myocardial beta-AR system of this rabbit model of HF shares important biochemical characteristics with human HF and therefore is an ideal laboratory model to investigate novel therapeutic targets for the treatment of HF.We studied alterations in the β-adrenergic receptor (β-AR) system of rabbit hearts during the development of heart failure (HF) after myocardial infarction (MI) to determine whether the molecular β-AR abnormalities associated with human HF exist in this animal model. Rabbit HF was established 3 wk after left circumflex coronary artery (LCX) ligation by in vivo physiological measurements, and molecular β-AR signaling was examined in tissue and cultured ventricular myocytes. We found that there was a significant global reduction in β-AR density by ∼50% in both ventricles of MI animals compared with sham-operated control animals and that functional β-AR coupling was significantly reduced. Importantly, as found in human HF, myocardial protein levels and activity of the β-AR kinase (β-ARK1) and Gαi were found to be significantly elevated in MI rabbits, suggesting that these molecules are contributing to myocardial dysfunction. Thus the myocardial β-AR system of this rabbit model of HF shares important biochemical characteristics with human HF and therefore is an ideal laboratory model to investigate novel therapeutic targets for the treatment of HF.

Risk-Adjusted Short- and Long-Term Outcomes for On-Pump Versus Off-Pump Coronary Artery Bypass Surgery

Background—Surgeons have adopted off-pump coronary artery bypass grafting (OPCAB) in an effort to reduce the morbidity of surgical revascularization. However, long-term outcome of OPCAB compared with conventional coronary artery bypass grafting (CABG) remains poorly defined. Methods and Results—Using logistic regression analysis and proportional hazards modeling, short-term and long-term outcomes (perioperative mortality and complications, risk-adjusted survival, and survival/freedom from revascularization) were investigated for patients who underwent OPCAB (641 patients) and CABG-cardiopulmonary bypass (5026 patients) from 1998 to 2003 at our institution. For these variables, follow-up was 98% complete. OPCAB patients were less likely to receive transfusion (odds ratio for OPCAB, 0.80; P=0.037), and there were trends toward improvement in other short-term outcomes compared with CABG-cardiopulmonary bypass. Long-term outcomes analysis demonstrated no difference in survival, but OPCAB patients were more likely to require repeat revascularization (OPCAB hazard ratio, 1.29; P=0.020). Conclusions—OPCAB patients were less likely to receive transfusion during their hospitalization for surgery but had higher risk for revascularization in follow-up. These results highlight the need for a large randomized, controlled trial to compare these 2 techniques.

Does a pre-left ventricular assist device screening score predict long-term transplantation success? A 2-center analysis.

BACKGROUND A risk factor summation score was previously validated to successfully predict survival after insertion of a left ventricular assist device (LVAD). We investigated whether this scoring system also predicts clinical outcomes after eventual heart transplantation in LVAD recipients. METHODS A retrospective review was performed on 153 consecutive patients who received an LVAD as a bridge to transplantation at 2 large-volume centers from 1996 to 2003. The scoring system was used to designate low- and high-scoring groups. RESULTS Thirty-day mortality and 5-year survival after transplantation were equivalent between groups (4.46% versus 7.32% and 76% versus 70%, respectively). No difference was seen in length of posttransplantation ventilator dependence (2.83 +/- 0.49 versus 3.3 +/- 0.72 days) or intensive care unit monitoring (6.38 +/- 0.77 versus 6.97 +/- 1.1 days). However, low-scoring patients had a significantly decreased duration of inotrope support (5.57 +/- 0.45 versus 7.74 +/- 1.0 days, P = .035). CONCLUSION A risk factor summation score may predict which LVAD patients will require prolonged inotropic support following heart transplantation. However, survival in high-risk (elevated score) LVAD patients following heart transplantation is comparable to low-risk groups, favoring the continued practice of LVAD implantation as a bridge to transplantation even in high-risk patients.