Differential Downregulation of ß1-Adrenergic Receptor Signaling in the Heart.

BACKGROUND: Chronic sympathetic stimulation drives desensitization and downregulation of ß1 adrenergic receptor (ß1AR) in heart failure. We aim to explore the differential downregulation subcellular pools of ß1AR signaling in the heart. METHODS AND RESULTS: We applied chronic infusion of isoproterenol to induced cardiomyopathy in male C57BL/6J mice. We applied confocal and proximity ligation assay to examine ß1AR association with L-type calcium channel, ryanodine receptor 2, and SERCA2a ((Sarco)endoplasmic reticulum calcium ATPase 2a) and Förster resonance energy transfer-based biosensors to probe subcellular ß1AR-PKA (protein kinase A) signaling in ventricular myocytes. Chronic infusion of isoproterenol led to reduced ß1AR protein levels, receptor association with L-type calcium channel and ryanodine receptor 2 measured by proximity ligation (puncta/cell, 29.65 saline versus 14.17 isoproterenol, P<0.05), and receptor-induced PKA signaling at the plasma membrane (Förster resonance energy transfer, 28.9% saline versus 1.9% isoproterenol, P<0.05) and ryanodine receptor 2 complex (Förster resonance energy transfer, 30.2% saline versus 10.6% isoproterenol, P<0.05). However, the ß1AR association with SERCA2a was enhanced (puncta/cell, 51.4 saline versus 87.5 isoproterenol, P<0.05), and the receptor signal was minimally affected. The isoproterenol-infused hearts displayed decreased PDE4D (phosphodiesterase 4D) and PDE3A and increased PDE2A, PDE4A, and PDE4B protein levels. We observed a reduced role of PDE4 and enhanced roles of PDE2 and PDE3 on the ß1AR-PKA activity at the ryanodine receptor 2 complexes and myocyte shortening. Despite the enhanced ß1AR association with SERCA2a, the endogenous norepinephrine-induced signaling was reduced at the SERCA2a complexes. Inhibiting monoamine oxidase A rescued the norepinephrine-induced PKA signaling at the SERCA2a and myocyte shortening. CONCLUSIONS: This study reveals distinct mechanisms for the downregulation of subcellular ß1AR signaling in the heart under chronic adrenergic stimulation.

ß1 Adrenergic Receptor Autoantibodies and IgG Subclasses: Current Status and Unsolved Issues.

A wide range of anti-myocardial autoantibodies have been reported since the 1970s. Among them, autoantibodies against the ß1-adrenergic receptor (ß1AR-AAb) have been the most thoroughly investigated, especially in dilated cardiomyopathy (DCM). Β1AR-Aabs have agonist effects inducing desensitization of ß1AR, cardiomyocyte apoptosis, and sustained calcium influx which lead to cardiac dysfunction and arrhythmias. Β1AR-Aab has been reported to be detected in approximately 40% of patients with DCM, and the presence of the antibody has been associated with worse clinical outcomes. The removal of anti-myocardial autoantibodies including ß1AR-AAb by immunoadsorption is beneficial for the improvement of cardiac function for DCM patients. However, several studies have suggested that its efficacy depended on the removal of AAbs belonging to the IgG3 subclass, not total IgG. IgG subclasses differ in the structure of the Fc region, suggesting that the mechanism of action of ß1AR-AAb differs depending on the IgG subclasses. Our previous clinical research demonstrated that the patients with ß1AR-AAb better responded to ß-blocker therapy, but the following studies found that its response also differed among IgG subclasses. Further studies are needed to elucidate the possible pathogenic role of IgG subclasses of ß1AR-AAbs in DCM, and the broad spectrum of cardiovascular diseases including HF with preserved ejection fraction.

Increased ß1-adrenergic receptor antibody confers a vulnerable substrate for atrial fibrillation via mediating Ca2+ mishandling and atrial fibrosis in active immunization rabbit models.

BACKGROUND: Autoimmune disorder is the emerging mechanism of atrial fibrillation (AF). The ß1-adrenergic receptor antibody (ß1-AAb) is associated with AF progress. Our study aims to investigate whether ß1-AAbs involves in atrial vulnerable substrate by mediating Ca2+ mishandling and atrial fibrosis in autoimmune associated AF. METHODS: Active immunization models were established via subcutaneous injection of the second extracellular loop (ECL2) peptide for ß1 adrenergic receptor (ß1AR). Invasive electrophysiologic study and ex vivo optical mapping were used to evaluate the changed electrophysiology parameters and calcium handling properties. Phospho-proteomics combined with molecular biology assay were performed to identify the potential mechanisms of remodeled atrial substrate elicited by ß1-AAbs. Exogenous ß1-AAbs were used to induce the cellular phenotypes of HL-1 cells and atrial fibroblasts to AF propensity. RESULTS: ß1-AAbs aggravated the atrial electrical instability and atrial fibrosis. Bisoprolol alleviated the alterations of action potential duration (APD), Ca2+ transient duration (CaD), and conduction heterogeneity challenged by ß1-AAbs. ß1-AAbs prolonged calcium transient refractoriness and promoted arrhythmogenic atrial alternans and spatially discordant alternans, which were partly counteracted through blocking ß1AR. Its underlying mechanisms are related to ß1AR-drived CaMKII/RyR2 activation of atrial cardiomyocytes and the myofibroblasts phenotype formation of fibroblasts. CONCLUSION: Suppressing ß1-AAbs effectively protects the atrial vulnerable substrate by ameliorating intracellular Ca2+ mishandling and atrial fibrosis, preventing the process of the autoimmune associated AF.

Monoamine oxidase A and organic cation transporter 3 coordinate intracellular ß1AR signaling to calibrate cardiac contractile function.

We have recently identified a pool of intracellular ß1 adrenergic receptors (ß1ARs) at the sarcoplasmic reticulum (SR) crucial for cardiac function. Here, we aim to characterize the integrative control of intracellular catecholamine for subcellular ß1AR signaling and cardiac function. Using anchored Förster resonance energy transfer (FRET) biosensors and transgenic mice, we determined the regulation of compartmentalized ß1AR-PKA signaling at the SR and plasma membrane (PM) microdomains by organic cation transporter 3 (OCT3) and monoamine oxidase A (MAO-A), two critical modulators of catecholamine uptake and homeostasis. Additionally, we examined local PKA substrate phosphorylation and excitation-contraction coupling in cardiomyocyte. Cardiac-specific deletion of MAO-A (MAO-A-CKO) elevates catecholamines and cAMP levels in the myocardium, baseline cardiac function, and adrenergic responses. Both MAO-A deletion and inhibitor (MAOi) selectively enhance the local ß1AR-PKA activity at the SR but not PM, and augment phosphorylation of phospholamban, Ca2+ cycling, and myocyte contractile response. Overexpression of MAO-A suppresses the SR-ß1AR-PKA activity and PKA phosphorylation. However, deletion or inhibition of OCT3 by corticosterone prevents the effects induced by MAOi and MAO-A deletion in cardiomyocytes. Deletion or inhibition of OCT3 also negates the effects of MAOi and MAO-A deficiency in cardiac function and adrenergic responses in vivo. Our data show that MAO-A and OCT3 act in concert to fine-tune the intracellular SR-ß1AR-PKA signaling and cardiac fight-or-flight response. We reveal a drug contraindication between anti-inflammatory corticosterone and anti-depressant MAOi in modulating adrenergic regulation in the heart, providing novel perspectives of these drugs with cardiac implications.

Prognostic Value of ß1 Adrenergic Receptor Autoantibody and Soluble Suppression of Tumorigenicity-2 in Patients With Acutely Decompensated Heart Failure.

BACKGROUND: Both ß1 adrenergic receptor autoantibody (ß1-AA) and soluble suppression of tumorigenicity-2 (sST2) take a role in the pathological remodeling of heart failure. However, limited studies investigated the correlation between the expression of ß1-AA and sST2 in patients with acutely decompensated heart failure (ADHF). OBJECTIVE: To explore the correlation between ß1-AA and sST2, and evaluate their prognostic value in patients with ADHF. METHODS: Patients who were admitted for ADHF were included. The N-terminal pro-brain natriuretic peptide (NT-proBNP), sST2, and ß1-AA in blood samples were tested at hospital admission and then followed up for assessing the outcomes. Pearson correlation analysis was used to explore the correlation between ß1-AA and sST2. The effects of ß1-AA, sST2, or the combination of them on the all-cause mortality of patients with ADHF were assessed by Multivariate Cox regression analysis. RESULTS: There were 96 patients with ADHF and 96 control populations enrolled. The ß1-AA was significantly higher in ADHF than in the control group (0.321 ± 0.06 vs. 0.229 ± 0.04, P = 0.000). Pearson correlation analysis showed that ß1-AA was positively correlated with sST2 (r = 0.593), NT-proBNP (r = 0.557), Procalcitonin (r = 0.176), and left ventricular end-diastolic diameter (r = 0.315), but negatively correlated with triglycerides (r = -0.323), and left ventricular ejection fraction (r = -0.430) (all P < 0.05) in ADHF. Patients with ADHF, complicated with both high ß1-AA and sST2, showed the highest all-cause mortality during an average of 25.5 months of follow-up. Multivariate Cox regression showed the combination of both high ß1-AA and sST2 independently correlated with the all-cause mortality after adjustment for other risk factors (hazard ratio 3.348, 95% CI 1.440 to 7.784, P = 0.005). After adding with ß1-AA and sST2, the area under the curves for the prognostic all-cause mortality could increase from 0.642 to 0.748 (P = 0.011). CONCLUSION: The ß1-AA is positively correlated with sST2 in patients with ADHF. Elevated plasma ß1-AA and sST2 level in patients with ADHF are associated with poorer prognoses.

Expression of ß1 adrenergic receptor in vascular anomalies in children.

OBJECTIVE: Controversial, heterogeneous, and inconsistent responses to beta-blockers have been reported in some cases of infantile proliferative hemangiomas. On the basis of these clinical observations, we aimed to examine the ß1 adrenergic receptor (ß1-AR) protein expression distribution among different types of pediatric vascular anomalies. METHODS: Immunohistochemistry (IHC) was performed for ß1-AR on 43 surgical specimens. RESULTS: We found positive ß1-AR IHC staining in all intramuscular hemangiomas, capillary-lymphatic, lymphatic, venous, and combined malformations, and Masson's tumor cases, as well as in 7 of 10 cases of proliferative infantile hemangiomas. CONCLUSIONS: Our research demonstrates, for the first time, the degree of heterogeneous expression of ß1-AR among pediatric vascular malformations. Our results support the need for ß1-AR assessment in pediatric vascular anomalies to select cases with a robust response to ß1-selective blockers. ß1-AR assessment may have a strong impact on therapeutic refinement for pediatric vascular anomalies by selecting cases with a stronger response to beta-blockers.

CLOCK-BMAL1 regulates circadian oscillation of ventricular arrhythmias in failing hearts through ß1 adrenergic receptor.

The incidence of ventricular arrhythmias (VAs) in chronic heart failure (CHF) exhibits a notable circadian rhythm, for which the underlying mechanism has not yet been well defined. Thus, we aimed to investigate the role of cardiac core circadian genes on circadian VAs in CHF. First, a guinea pig CHF model was created by transaortic constriction. Circadian oscillation of core clock genes was evaluated by RT-PCR and was found to be unaltered in CHF (P > 0.05). Using programmed electrical stimulation in Langendorff-perfused failing hearts, we discovered that the CHF group exhibited increased VAs with greater incidence at CT3 compared to CT15 upon isoproterenol (ISO) stimulation. Circadian VAs was blunted by a ß1-AR-selective blocker rather than a ß2-AR-selective blocker. Circadian oscillation of ß1-AR was retained in CHF (P > 0.05) and a 4-h phase delay between ß1-AR and CLOCK-BMAL1 was recorded. Therefore, when CLOCK-BMAL1 was overexpressed using adenovirus infection, an induced overexpression of ß1-AR also ensued, which resulted in prolonged action potential duration (APD) and enhanced arrhythmic response to ISO stimulation in cardiomyocytes (P < 0.05). Finally, chromatin immunoprecipitation and luciferase assays confirmed that CLOCK-BMAL1 binds to the enhancer of ß1-AR gene and upregulates ß1-AR expression. Therefore, in this study, we discovered that CLOCK-BMAL1 regulates the expression of ß1-AR on a transcriptional level and subsequently modulates circadian VAs in CHF.

Relationship between catecholamine level and gene polymorphism of ß1 adrenergic receptor G1165C in children with EV71 infection in hand foot and mouth disease.

OBJECTIVE: To investigate the relationship between the levels of plasma adrenaline and norepinephrine and gene polymorphism of ß1 adrenergic receptor G1165C in children with enterovirus 71 (EV71) infection in hand foot and mouth disease (HFMD). METHODS: The polymerase chain reaction (PCR) was used to detect the expression of gene polymorphism of ß1 adrenergic receptor G1165C in vitro. The levels of plasma adrenaline and norepinephrine were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: The plasma norepinephrine level of severe group was significantly higher than the mild group in children with EV71 infection in HFMD (P < 0.05); however, the levels of plasma adrenaline in two groups had no statistical differences (P > 0.05); There was no significant difference in the distribution of ß1 adrenergic receptor G1165C genotype and allele between EV71 infection group and healthy control group (P > 0.05). Further analysis of EV71 infection group by dividing it into mild and severe groups showed that there was no significant difference in the distribution of genotype and allele between these two groups as well (P > 0.05). There was no significant difference in the levels of epinephrine and norepinephrine in different genotypes of EV71 infection group (P > 0.05), and in the levels of plasma epinephrine and norepinephrine in the mild and severe groups (P > 0.05). CONCLUSIONS: As the disease gets worse, the plasma norepinephrine level has a rising trend in children with EV71 infection in HFMD, which is an important indicator to evaluate the progress of the disease. However, the gene polymorphism of ß1 adrenergic receptor G1165C have no significant correlation, not only with the susceptibility and severity of EV71 infection in hand, foot and mouth disease, but also with the levels of catecholamine.