Introduction:
Single ventricle congenital heart disease (SV) is the leading cause of cardiovascular death and indication for heart transplantation in infancy. There are no proven therapies for SV heart failure (HF). Human and animal models of HF demonstrate that myocardial phosphodiesterase-5 (PDE5) is increased with cardiac stress and treatment with a PDE5 inhibitor (PDE5i) results in enhanced cardiac function and prevents remodeling. Sildenafil, a PDE5i, is increasingly utilized for the treatment of patients suffering from failing SV. The objective of this study was to determine myocardial PDE5 expression and activity in children transplanted for failing SV.
Methods:
At the time of cardiac transplantation, explanted pediatric hearts were immediately cooled in ice cold oxygenated Tyrodes in the operating room. The tissue is rapidly dissected, flash frozen and stored at -80
0
C until further use. RNA, protein and cytosolic fractions were isolated from explanted right ventricle (RV) tissue from SV and non-failing (NF) donors. RTqPCR for PDE5, Western blot (normalized to calnexin loading control) for PDE5 and PDE5 activity assays were performed. For PDE 5 activity, cGMP hydrolysis was measured using [
3
H]cGMP as the substrate. Sildenafil was added to measure PDE5 specific activity, and activity was calculated using nonlinear regression.
Results:
All patients used in the SV analysis had a failing morphologic RV and were selected from a cohort of 17 SV (median age 0.5, range 0.05-10 yrs) and 8 NF controls (median age 7, range 1.3-13 yrs). PDE5 gene expression was higher in SV myocardium compared to NF (1.9±0.7, n=17 SV vs 1.1±0.5 ct fold change, n=8 NF, p=0.02). There was a trend towards higher PDE5 protein expression in SV myocardium compared to NF (1.5±0.7, n=4 SV vs 1.0±0.4 protein expression normalized to NF, n=3 NF; p=ns). There was increased PDE5 activity in SV compared to NF (22.3±1.2, n=3 SV vs 11.9±4.2 pmol/mg/min, n=2 NF; p=0.02).
Conclusions:
There is increasing evidence that PDE5i has beneficial direct myocardial effects. There is increased PDE5 gene expression and activity in failing SV myocardium compared to NF control suggesting that PDE5 may represent a promising therapeutic target in this challenging population.
Right ventricular cyclic nucleotide signaling is decreased in hyperoxia-induced pulmonary hypertension in neonatal mice
