Penny Nelson

Adrenergic receptor polymorphisms in patients with stress (tako-tsubo) cardiomyopathy

BACKGROUND Stress (tako-tsubo) cardiomyopathy (SC) is a newly reported condition afflicting older women, characterized by acute left ventricular (LV) systolic dysfunction, triggered by emotionally and physically stressful events, and occurring without significant coronary obstruction. Sympathetic nervous system hyperactivity has been implicated in the pathophysiology of SC. Single nucleotide polymorphisms involving the adrenergic receptors (AR) might result in susceptibility to SC. METHODS Forty-one female SC patients were identified aged 34-89 years (mean 65) and were compared with 43 control females of similar age with respect to AR genotype frequencies for B1 receptor (amino acid positions 389 and 49) and alpha 2c receptor (deletion 322-325). RESULTS For SC patients, initial LV ejection fraction was 32 ± 10% vs. 62 ± 11% in control patients, p < 0.05. Genotype frequencies for SC patients vs. controls were B1 389 Arg/Arg (0.49 vs. 0.51), B1 389 Arg/Gly (0.49 vs. 0.49), B1 389 Gly/Gly (0.02 vs. 0), B1 49 Ser/Ser (0.88 vs. 0.81), B1 49 Ser/Gly (0.12 vs. 0.16), B1 49 Gly/Gly (0 vs. 0.02), alpha 2c Wt/Wt (0.93 vs. 0.86), and alpha 2c Wt/Del 322-325 (0.07 vs. 0.14); p = ns for all comparisons. CONCLUSIONS Genotype polymorphism frequencies for B1 receptor (amino acid positions 389 and 49) and alpha 2c receptor (deletion 322-325) are not significantly different in SC patients compared to female controls. These data suggest that these AR polymorphisms do not mediate the sympathetic nervous system hyperactivity in SC patients.

Beta-adrenergic adaptation in paediatric idiopathic dilated cardiomyopathy

BACKGROUND Although the pathophysiology and treatment of adult heart failure (HF) are well studied, HF in children remains poorly understood. In adults, adrenergic receptor (AR)-mediated adaptation plays a central role in cardiac abnormalities in HF, and these patients respond well to β-blocker (BB) therapy. However, in children with HF, there is a growing body of literature suggesting a lack of efficacy of adult HF therapies. Due to these unanticipated differences in response to therapy and the paucity of data regarding the molecular adaptation of the paediatric heart, we investigated the molecular characteristics of HF in children. METHODS AND RESULTS Explanted hearts from adults and children with idiopathic dilated cardiomyopathy and non-failing controls were used in the study. Our results show that the molecular characteristics of paediatric HF are strikingly different from their adult counterparts. These differences include: (i) down-regulation of β1- and β2-AR in children, whereas β2-AR expression is maintained in adults; (ii) up-regulation of connexin43 in children, whereas down-regulation is observed in adults; (iii) no differences in phosphatase expression, whereas up-regulation is observed in adults; (iv) no decrease in the phosphorylation of phospholamban at the Ser16 or Thr17 sites in children, which are known characteristics of adult HF. CONCLUSION There is a different adaptation of β-AR and adrenergic signalling pathways in children with HF compared with adults. Our results begin to address the disparities in cardiovascular research specific to children and suggest that age-related differences in adaptation could influence the response to therapy. These findings could lead to a paradigm shift in the contemporary management of children with HF.

Combinatorial Pharmacogenetic Interactions of Bucindolol and β1, α2C Adrenergic Receptor Polymorphisms

Background Pharmacogenetics involves complex interactions of gene products affecting pharmacodynamics and pharmacokinetics, but there is little information on the interaction of multiple genetic modifiers of drug response. Bucindolol is a β-blocker/sympatholytic agent whose efficacy is modulated by polymorphisms in the primary target (β1 adrenergic receptor [AR] Arg389 Gly on cardiac myocytes) and a secondary target modifier (α2C AR Ins [wild-type (Wt)] 322–325 deletion [Del] on cardiac adrenergic neurons). The major allele homozygotes and minor allele carriers of each polymorphism are respectively associated with efficacy enhancement and loss, creating the possibility for genotype combination interactions that can be measured by clinical trial methodology. Methodology In a 1,040 patient substudy of a bucindolol vs. placebo heart failure clinical trial, we tested the hypothesis that combinations of β1389 and α2C322–325 polymorphisms are additive for both efficacy enhancement and loss. Additionally, norepinephrine (NE) affinity for β1389 AR variants was measured in human explanted left ventricles. Principal Findings The combination of β1389 Arg+α2C322–325 Wt major allele homozygotes (47% of the trial population) was non-additive for efficacy enhancement across six clinical endpoints, with an average efficacy increase of 1.70-fold vs. 2.32-fold in β1389 Arg homozygotes+α2C322–325 Del minor allele carriers. In contrast, the minor allele carrier combination (13% subset) exhibited additive efficacy loss. These disparate effects are likely due to the higher proportion (42% vs. 8.7%, P = 0.009) of high-affinity NE binding sites in β1389 Arg vs. Gly ARs, which converts α2CDel minor allele-associated NE lowering from a therapeutic liability to a benefit. Conclusions On combination, the two sets of AR polymorphisms 1) influenced bucindolol efficacy seemingly unpredictably but consistent with their pharmacologic interactions, and 2) identified subpopulations with enhanced (β1389 Arg homozygotes), intermediate (β1389 Gly carriers+α2C322–325 Wt homozygotes), and no (β1389 Gly carriers+α2C322–325 Del carriers) efficacy.

Age-Related Differences in Phosphodiesterase Activity and Effects of Chronic Phosphodiesterase Inhibition in Idiopathic Dilated Cardiomyopathy

Background—Despite the application of proven adult heart failure therapies to children with idiopathic dilated cardiomyopathy (IDC), prognosis remains poor. Clinical experience with phosphodiesterase 3 inhibitors (PDE3i) in pediatric patients with IDC, however, demonstrates improved heart failure symptoms without the increased incidence of sudden death seen in adults treated with PDE3i. We sought to determine age-related differences in PDE activity and associated intracellular signaling responsible for the efficacy and relative safety of chronic PDE3i in pediatric heart failure. Methods and Results—cAMP levels, PDE activity, and phospholamban phosphorylation (pPLB) were determined in explanted human left ventricular myocardium (pediatric n=41; adult n=88). Adults and children with IDC (not treated with PDE3i) had lower cAMP and pPLB compared with nonfailing controls. In contrast to their adult counterparts, pediatric IDC patients chronically treated with PDE3i had elevated cAMP (P=0.0403) and pPLB (P=0.0119). In addition, total PDE- and PDE3-specific activities were not altered in pediatric IDC patients on PDE3i, whereas adult IDC patients on PDE3i demonstrated higher total PDE-specific (74.6±13.8 pmol/mg per minute) and PDE3-specific (48.2±15.9 pmol/mg per minute) activities in comparison with those of nonfailing controls (59.5±14.4 and 35.5±12.8 pmol/mg per minute, respectively). Conclusions—Elevated cAMP and higher pPLB may contribute to sustained hemodynamic benefits in pediatric IDC patients treated with PDE3i. In contrast, higher total PDE and PDE3 activities in adult IDC patients treated with PDE3i may perpetuate lower myocardial cAMP and pPLB levels, limiting the potential benefits of PDE3i therapy.

Temporal analysis of mRNA and miRNA expression in transgenic mice overexpressing Arg- and Gly389 polymorphic variants of the β1-adrenergic receptor.

Several studies in humans or transgenic animals have reported that the 389 Arg or Gly polymorphic variation of the β1-adrenergic receptor (AR) is associated with differential responses to beta-blocker therapy and/or myocardial disease progression. Analysis of changes in gene expression is an important means of defining molecular differences associated with structural or functional phenotypic variations. To determine if structural and functional myocardial phenotypic differences between β1389 Arg vs. Gly transgenic overexpressors are associated with qualitative and/or quantitative differences in gene expression, a comprehensive analysis of mRNAs and miRNAs expressed in the hearts of 3 and 6-8 mo old β1-Arg389 and β1-Gly389 overexpressor transgenic mice was performed. Changes in mRNA and miRNA expression were analyzed by arrays and partially confirmed by RT-qPCR. Bioinformatic analysis demonstrated that several genes, including those involved in PKA and CaMK signaling pathways, are regulated in a temporal- or phenotype-specific manner. Furthermore, expression signature analyses indicated that miRNAs have the potential to target expression of a number of genes involved in multiple cardiomyopathy-related pathways, and changes in miRNA expression can precede the onset of disease. Differences in gene expression between β1-Arg389 and β1-Gly389 transgenic mice are largely quantitative rather than qualitative and are associated with the development of cardiomyopathy in a time-dependent manner. Chronic β1-AR overdrive results in increased expression of components of the CaMK pathway, with correspondingly decreased levels of components of the PKA pathway. Based on the temporal and genotype-specific pattern of miRNA expression, miRNAs are likely to be important predictors of disease states, especially when miRNA expression is paired with mRNA expression, and that miRNA/mRNA expression signatures have the potential to be useful in determining the underlying risk associated with cardiac disease progression.

Molecular Changes in Children with Heart Failure Undergoing Left Ventricular Assist Device Therapy

Objective To determine whether left ventricular assist device (LVAD) treatment in children with heart failure would result in the modification of molecular pathways involved in heart failure pathophysiology. Study design Forty‐seven explanted hearts from children were studied (16 nonfailing control, 20 failing, and 11 failing post‐LVAD implantation [F‐LVAD]). Protein expression and phosphorylation states were determined by receptor binding assays and Western blots. mRNA expression was measured with real‐time quantitative polymerase chain reaction. To evaluate for interactions and identify correlations, 2‐way ANOVA and regression analysis were performed. Results Treatment with LVAD resulted in recovery of total &bgr;‐adrenergic receptor expression and &bgr;1‐adrenergic receptor (&bgr;1‐AR) in failing hearts to normal levels (&bgr;‐adrenergic receptor expression: 67.2 ± 11.5 fmol/mg failing vs 99.5 ± 27.7 fmol/mg nonfailing, 104 ± 38.7 fmol/mg F‐LVAD, P ≤ .01; &bgr;1‐AR: 52.2 ± 10.3 fmol/mg failing vs 83.0 ± 23 fmol/mg non‐failing, 76.5 ± 32.1 fmol/mg F‐LVAD P ≤ .03). The high levels of G protein‐coupled receptor kinase‐2 were returned to nonfailing levels after LVAD treatment (5.6 ± 9.0 failing vs 1.0 ± 0.493 nonfailing, 1.0 ± 1.3 F‐LVAD). Interestingly, &bgr;2‐adrenergic receptor expression was significantly greater in F‐LVAD (27.5 ± 12; P < .005) hearts compared with nonfailing (16.4 ± 6.1) and failing (15.1 ± 4.2) hearts. Phospholamban phosphorylation at serine 16 was significantly greater in F‐LVAD (7.7 ± 11.7) hearts compared with nonfailing (1.0 ± 1.2, P = .02) and failing (0.8 ± 1.0, P = .01) hearts. Also, atrial natriuretic factor (0.6 ± 0.8) and brain natriuretic peptide (0.1 ± 0.1) expression in F‐LVAD was significantly lower compared with failing hearts (2.8 ± 3.6, P = .01 and 0.6 ± 0.7, P = .02). Conclusion LVAD treatment in children with heart failure results in reversal of several pathologic myocellular processes, and G protein‐coupled receptor kinase‐2 may regulate &bgr;1‐AR but not &bgr;2‐adrenergic receptor expression in children with heart failure.