Bastian Spallek

RBM20 , a gene for hereditary cardiomyopathy, regulates titin splicing

Alternative splicing has a major role in cardiac adaptive responses, as exemplified by the isoform switch of the sarcomeric protein titin, which adjusts ventricular filling. By positional cloning using a previously characterized rat strain with altered titin mRNA splicing, we identified a loss-of-function mutation in the gene encoding RNA binding motif protein 20 (Rbm20) as the underlying cause of pathological titin isoform expression. The phenotype of Rbm20-deficient rats resembled the pathology seen in individuals with dilated cardiomyopathy caused by RBM20 mutations. Deep sequencing of the human and rat cardiac transcriptome revealed an RBM20-dependent regulation of alternative splicing. In addition to titin (TTN), we identified a set of 30 genes with conserved splicing regulation between humans and rats. This network is enriched for genes that have previously been linked to cardiomyopathy, ion homeostasis and sarcomere biology. Our studies emphasize the key role of post-transcriptional regulation in cardiac function and provide mechanistic insights into the pathogenesis of human heart failure.

Interferon-γ Signaling Inhibition Ameliorates Angiotensin II–Induced Cardiac Damage

Angiotensin (Ang) II induces vascular injury in part by activating innate and adaptive immunity; however, the mechanisms are unclear. We investigated the role of interferon (IFN)-&ggr; and interleukin (IL)-23 signaling. We infused Ang II into IFN-&ggr; receptor (IFN-&ggr;R) knockout mice and wild-type controls, as well as into mice treated with neutralizing antibodies against IL-23 receptor and IL-17A. Ang II–treated IFN-&ggr;R knockout mice exhibited reduced cardiac hypertrophy, reduced cardiac macrophage and T-cell infiltration, less fibrosis, and less arrhythmogenic electric remodeling independent of blood pressure changes. In contrast, IL-23 receptor antibody treatment did not reduce cardiac hypertrophy, fibrosis, or electric remodeling despite mildly reduced inflammation. IL-17A antibody treatment behaved similarly. In the kidney, IFN-&ggr;R deficiency reduced inflammation and tubulointerstitial damage and improved glomerular filtration rate. Nonetheless, albuminuria was increased compared with Ang II–treated wild-type controls. The glomeruli of Ang II–treated IFN-&ggr;R knockout mice exhibited fewer podocytes, less nephrin and synaptopodin staining, and impaired podocyte autophagy. Thus, IFN-&ggr; blockade, but not IL-23 receptor antibody treatment, protects from Ang II–induced cardiac damage and electric remodeling. In the kidney, IFN-&ggr; signaling acts in a cell type–specific manner. Glomerular filtration rate is preserved in the absence of the IFN-&ggr;R, whereas podocytes may require the IFN-&ggr;R in the presence of Ang II for normal integrity and function.

Coxsackie and Adenovirus Receptor Is a Modifier of Cardiac Conduction and Arrhythmia Vulnerability in the Setting of Myocardial Ischemia

OBJECTIVES The aim of this study was to investigate the modulatory effect of the coxsackie and adenovirus receptor (CAR) on ventricular conduction and arrhythmia vulnerability in the setting of myocardial ischemia. BACKGROUND A heritable component in the risk of ventricular fibrillation during myocardial infarction has been well established. A recent genome-wide association study of ventricular fibrillation during acute myocardial infarction led to the identification of a locus on chromosome 21q21 (rs2824292) in the vicinity of the CXADR gene. CXADR encodes the CAR, a cell adhesion molecule predominantly located at the intercalated disks of the cardiomyocyte. METHODS The correlation between CAR transcript levels and rs2824292 genotype was investigated in human left ventricular samples. Electrophysiological studies and molecular analyses were performed using CAR haploinsufficient (CAR⁺/⁻) mice. RESULTS In human left ventricular samples, the risk allele at the chr21q21 genome-wide association study locus was associated with lower CXADR messenger ribonucleic acid levels, suggesting that decreased cardiac levels of CAR predispose to ischemia-induced ventricular fibrillation. Hearts from CAR⁺/⁻ mice displayed slowing of ventricular conduction in addition to an earlier onset of ventricular arrhythmias during the early phase of acute myocardial ischemia after ligation of the left anterior descending artery. Expression and distribution of connexin 43 were unaffected, but CAR⁺/⁻ hearts displayed increased arrhythmia susceptibility on pharmacological electrical uncoupling. Patch-clamp analysis of isolated CAR⁺/⁻ myocytes showed reduced sodium current magnitude specifically at the intercalated disk. Moreover, CAR coprecipitated with NaV1.5 in vitro, suggesting that CAR affects sodium channel function through a physical interaction with NaV1.5. CONCLUSIONS CAR is a novel modifier of ventricular conduction and arrhythmia vulnerability in the setting of myocardial ischemia. Genetic determinants of arrhythmia susceptibility (such as CAR) may constitute future targets for risk stratification of potentially lethal ventricular arrhythmias in patients with coronary artery disease.

Bcl10 Mediates Angiotensin II–Induced Cardiac Damage and Electrical Remodeling

Angiotensin (Ang) II is a potent mediator of both hypertension and cardiac damage; however, the mechanisms by which this occur remain unclear. B-cell lymphoma/leukemia 10 (Bcl10) is a member of the CBM signalosome, which links Ang II and nuclear factor-&kgr;B signaling. We hypothesized that Bcl10 is pivotal in the pathogenesis of Ang II–induced cardiac damage. Ang II infusion in mice lacking Bcl10 resulted in reduced cardiac fibrosis, less cellular infiltration, and improved arrhythmogenic electric remodeling, despite a similar degree of hypertension or cardiac hypertrophy. Adoptive transfer of bone marrow (BM), whereby Bcl10 knockout or wildtype BM was transferred to their opposite genotype recipients, revealed the dual importance of Bcl10 within both cardiac and immune cells. Loss of Bcl10 in cardiac cells resulted in reduced expression of genes important for the adhesion and recruitment of immune cells. In vitro experiments demonstrated that adhesion of monocytes to Ang II–treated endothelial cells also required Bcl10. Additionally, Bcl10 deficiency in macrophages reduced their intrinsic migratory ability. To address the role of BM-derived fibroblasts in the formation of cardiac fibrosis, we explored whether Bcl10 is also important for the infiltration of BM-derived (myo)fibroblasts into the heart. The transfer of green fluorescent protein positive wildtype BM into Bcl10 knockout recipient mice revealed a reduced number of noncardiac (myo)fibroblasts compared with those wildtype recipients. Our results demonstrate the significant role of Bcl10 in multiple cell types important for the generation of Ang II–induced cardiac damage and electric remodeling and may provide a new avenue for therapeutic intervention.

Impaired myocardial development resulting in neonatal cardiac hypoplasia alters postnatal growth and stress response in the heart

AIMS Foetal growth has been proposed to influence cardiovascular health in adulthood, a process referred to as foetal programming. Indeed, intrauterine growth restriction in animal models alters heart size and cardiomyocyte number in the perinatal period, yet the consequences for the adult or challenged heart are largely unknown. The aim of this study was to elucidate postnatal myocardial growth pattern, left ventricular function, and stress response in the adult heart after neonatal cardiac hypoplasia in mice. METHODS AND RESULTS Utilizing a new mouse model of impaired cardiac development leading to fully functional but hypoplastic hearts at birth, we show that myocardial mass is normalized until early adulthood by accelerated physiological cardiomyocyte hypertrophy. Compensatory hypertrophy, however, cannot be maintained upon ageing, resulting in reduced organ size without maladaptive myocardial remodelling. Angiotensin II stress revealed aberrant cardiomyocyte growth kinetics in adult hearts after neonatal hypoplasia compared with normally developed controls, characterized by reversible overshooting hypertrophy. This exaggerated growth mainly depends on STAT3, whose inhibition during angiotensin II treatment reduces left ventricular mass in both groups but causes contractile dysfunction in developmentally impaired hearts only. Whereas JAK/STAT3 inhibition reduces cardiomyocyte cross-sectional area in the latter, it prevents fibrosis in control hearts, indicating fundamentally different mechanisms of action. CONCLUSION Impaired prenatal development leading to neonatal cardiac hypoplasia alters postnatal cardiac growth and stress response in vivo, thereby linking foetal programming to organ size control in the heart.

Amyloid-β peptides activate α1-adrenergic cardiovascular receptors

Alzheimer disease features amyloid-&bgr; (A&bgr;) peptide deposition in brain and blood vessels and is associated with hypertension. A&bgr; peptide can cause vasoconstriction and endothelial dysfunction. We observed that A&bgr; peptides exert a chronotropic effect in neonatal cardiomyocytes, similar to &agr;1-adrenergic receptor autoantibodies that we described earlier. Recently, it was shown that &agr;1-adrenergic receptor could impair blood–brain flow. We hypothesized that A&bgr; peptides might elicit a signal transduction pathway in vascular cells, induced by &agr;1-adrenergic receptor activation. A&bgr; (25–35) and A&bgr; (10–35) induced a positive chronotropic effect in the cardiac contraction assay (28.75±1.15 and 29.40±0.98 bpm), which was attenuated by &agr;1-adrenergic receptor blockers (urapidil, 1.53±1.17 bpm; prazosin, 0.30±0.96 bpm). Both A&bgr; peptides induced an intracellular calcium release in vascular smooth muscle cells. Chronotropic activity and calcium response elicited by A&bgr; (25–35) were blocked with peptides corresponding to the first extracellular loop of the &agr;1-adrenergic receptor. We observed an induction of extracellular-regulated kinase 1/2 phosphorylation by A&bgr; (25–35) in Chinese hamster ovary cells overexpressing &agr;1-adrenergic receptor, vascular smooth muscle cells, and cardiomyocytes. We generated an activation-state–sensitive &agr;1-adrenergic receptor antibody and visualized activation of the &agr;1-adrenergic receptor by A&bgr; peptide. A&bgr; (25–35) induced vasoconstriction of mouse aortic rings and in coronary arteries in Langendorff-perfused rat hearts that resulted in decreased coronary flow. Both effects could be reversed by &agr;1-adrenergic receptor blockade. Our data are relevant to the association between Alzheimer disease and hypertension. They may explain impairment of vascular responses by A&bgr; and could have therapeutic implications.Alzheimer disease features amyloid-β (Aβ) peptide deposition in brain and blood vessels and is associated with hypertension. Aβ peptide can cause vasoconstriction and endothelial dysfunction. We observed that Aβ peptides exert a chronotropic effect in neonatal cardiomyocytes, similar to α 1 -adrenergic receptor autoantibodies that we described earlier. Recently, it was shown that α 1 -adrenergic receptor could impair blood–brain flow. We hypothesized that Aβ peptides might elicit a signal transduction pathway in vascular cells, induced by α 1 -adrenergic receptor activation. Aβ (25–35) and Aβ (10–35) induced a positive chronotropic effect in the cardiac contraction assay (28.75±1.15 and 29.40±0.98 bpm), which was attenuated by α 1 -adrenergic receptor blockers (urapidil, 1.53±1.17 bpm; prazosin, 0.30±0.96 bpm). Both Aβ peptides induced an intracellular calcium release in vascular smooth muscle cells. Chronotropic activity and calcium response elicited by Aβ (25–35) were blocked with peptides corresponding to the first extracellular loop of the α 1 -adrenergic receptor. We observed an induction of extracellular-regulated kinase 1/2 phosphorylation by Aβ (25–35) in Chinese hamster ovary cells overexpressing α 1 -adrenergic receptor, vascular smooth muscle cells, and cardiomyocytes. We generated an activation-state–sensitive α 1 -adrenergic receptor antibody and visualized activation of the α 1 -adrenergic receptor by Aβ peptide. Aβ (25–35) induced vasoconstriction of mouse aortic rings and in coronary arteries in Langendorff-perfused rat hearts that resulted in decreased coronary flow. Both effects could be reversed by α 1 -adrenergic receptor blockade. Our data are relevant to the association between Alzheimer disease and hypertension. They may explain impairment of vascular responses by Aβ and could have therapeutic implications.

Amyloid-β Peptides Activate α 1 -Adrenergic Cardiovascular Receptors

Alzheimer disease features amyloid-&bgr; (A&bgr;) peptide deposition in brain and blood vessels and is associated with hypertension. A&bgr; peptide can cause vasoconstriction and endothelial dysfunction. We observed that A&bgr; peptides exert a chronotropic effect in neonatal cardiomyocytes, similar to &agr;1-adrenergic receptor autoantibodies that we described earlier. Recently, it was shown that &agr;1-adrenergic receptor could impair blood–brain flow. We hypothesized that A&bgr; peptides might elicit a signal transduction pathway in vascular cells, induced by &agr;1-adrenergic receptor activation. A&bgr; (25–35) and A&bgr; (10–35) induced a positive chronotropic effect in the cardiac contraction assay (28.75±1.15 and 29.40±0.98 bpm), which was attenuated by &agr;1-adrenergic receptor blockers (urapidil, 1.53±1.17 bpm; prazosin, 0.30±0.96 bpm). Both A&bgr; peptides induced an intracellular calcium release in vascular smooth muscle cells. Chronotropic activity and calcium response elicited by A&bgr; (25–35) were blocked with peptides corresponding to the first extracellular loop of the &agr;1-adrenergic receptor. We observed an induction of extracellular-regulated kinase 1/2 phosphorylation by A&bgr; (25–35) in Chinese hamster ovary cells overexpressing &agr;1-adrenergic receptor, vascular smooth muscle cells, and cardiomyocytes. We generated an activation-state–sensitive &agr;1-adrenergic receptor antibody and visualized activation of the &agr;1-adrenergic receptor by A&bgr; peptide. A&bgr; (25–35) induced vasoconstriction of mouse aortic rings and in coronary arteries in Langendorff-perfused rat hearts that resulted in decreased coronary flow. Both effects could be reversed by &agr;1-adrenergic receptor blockade. Our data are relevant to the association between Alzheimer disease and hypertension. They may explain impairment of vascular responses by A&bgr; and could have therapeutic implications.Alzheimer disease features amyloid-β (Aβ) peptide deposition in brain and blood vessels and is associated with hypertension. Aβ peptide can cause vasoconstriction and endothelial dysfunction. We observed that Aβ peptides exert a chronotropic effect in neonatal cardiomyocytes, similar to α 1 -adrenergic receptor autoantibodies that we described earlier. Recently, it was shown that α 1 -adrenergic receptor could impair blood–brain flow. We hypothesized that Aβ peptides might elicit a signal transduction pathway in vascular cells, induced by α 1 -adrenergic receptor activation. Aβ (25–35) and Aβ (10–35) induced a positive chronotropic effect in the cardiac contraction assay (28.75±1.15 and 29.40±0.98 bpm), which was attenuated by α 1 -adrenergic receptor blockers (urapidil, 1.53±1.17 bpm; prazosin, 0.30±0.96 bpm). Both Aβ peptides induced an intracellular calcium release in vascular smooth muscle cells. Chronotropic activity and calcium response elicited by Aβ (25–35) were blocked with peptides corresponding to the first extracellular loop of the α 1 -adrenergic receptor. We observed an induction of extracellular-regulated kinase 1/2 phosphorylation by Aβ (25–35) in Chinese hamster ovary cells overexpressing α 1 -adrenergic receptor, vascular smooth muscle cells, and cardiomyocytes. We generated an activation-state–sensitive α 1 -adrenergic receptor antibody and visualized activation of the α 1 -adrenergic receptor by Aβ peptide. Aβ (25–35) induced vasoconstriction of mouse aortic rings and in coronary arteries in Langendorff-perfused rat hearts that resulted in decreased coronary flow. Both effects could be reversed by α 1 -adrenergic receptor blockade. Our data are relevant to the association between Alzheimer disease and hypertension. They may explain impairment of vascular responses by Aβ and could have therapeutic implications.

Amyloid-β Peptides Activate α1-Adrenergic Cardiovascular ReceptorsNovelty and Significance

Alzheimer disease features amyloid-&bgr; (A&bgr;) peptide deposition in brain and blood vessels and is associated with hypertension. A&bgr; peptide can cause vasoconstriction and endothelial dysfunction. We observed that A&bgr; peptides exert a chronotropic effect in neonatal cardiomyocytes, similar to &agr;1-adrenergic receptor autoantibodies that we described earlier. Recently, it was shown that &agr;1-adrenergic receptor could impair blood–brain flow. We hypothesized that A&bgr; peptides might elicit a signal transduction pathway in vascular cells, induced by &agr;1-adrenergic receptor activation. A&bgr; (25–35) and A&bgr; (10–35) induced a positive chronotropic effect in the cardiac contraction assay (28.75±1.15 and 29.40±0.98 bpm), which was attenuated by &agr;1-adrenergic receptor blockers (urapidil, 1.53±1.17 bpm; prazosin, 0.30±0.96 bpm). Both A&bgr; peptides induced an intracellular calcium release in vascular smooth muscle cells. Chronotropic activity and calcium response elicited by A&bgr; (25–35) were blocked with peptides corresponding to the first extracellular loop of the &agr;1-adrenergic receptor. We observed an induction of extracellular-regulated kinase 1/2 phosphorylation by A&bgr; (25–35) in Chinese hamster ovary cells overexpressing &agr;1-adrenergic receptor, vascular smooth muscle cells, and cardiomyocytes. We generated an activation-state–sensitive &agr;1-adrenergic receptor antibody and visualized activation of the &agr;1-adrenergic receptor by A&bgr; peptide. A&bgr; (25–35) induced vasoconstriction of mouse aortic rings and in coronary arteries in Langendorff-perfused rat hearts that resulted in decreased coronary flow. Both effects could be reversed by &agr;1-adrenergic receptor blockade. Our data are relevant to the association between Alzheimer disease and hypertension. They may explain impairment of vascular responses by A&bgr; and could have therapeutic implications.Alzheimer disease features amyloid-β (Aβ) peptide deposition in brain and blood vessels and is associated with hypertension. Aβ peptide can cause vasoconstriction and endothelial dysfunction. We observed that Aβ peptides exert a chronotropic effect in neonatal cardiomyocytes, similar to α 1 -adrenergic receptor autoantibodies that we described earlier. Recently, it was shown that α 1 -adrenergic receptor could impair blood–brain flow. We hypothesized that Aβ peptides might elicit a signal transduction pathway in vascular cells, induced by α 1 -adrenergic receptor activation. Aβ (25–35) and Aβ (10–35) induced a positive chronotropic effect in the cardiac contraction assay (28.75±1.15 and 29.40±0.98 bpm), which was attenuated by α 1 -adrenergic receptor blockers (urapidil, 1.53±1.17 bpm; prazosin, 0.30±0.96 bpm). Both Aβ peptides induced an intracellular calcium release in vascular smooth muscle cells. Chronotropic activity and calcium response elicited by Aβ (25–35) were blocked with peptides corresponding to the first extracellular loop of the α 1 -adrenergic receptor. We observed an induction of extracellular-regulated kinase 1/2 phosphorylation by Aβ (25–35) in Chinese hamster ovary cells overexpressing α 1 -adrenergic receptor, vascular smooth muscle cells, and cardiomyocytes. We generated an activation-state–sensitive α 1 -adrenergic receptor antibody and visualized activation of the α 1 -adrenergic receptor by Aβ peptide. Aβ (25–35) induced vasoconstriction of mouse aortic rings and in coronary arteries in Langendorff-perfused rat hearts that resulted in decreased coronary flow. Both effects could be reversed by α 1 -adrenergic receptor blockade. Our data are relevant to the association between Alzheimer disease and hypertension. They may explain impairment of vascular responses by Aβ and could have therapeutic implications.