Antagonistic Roles of GRK2 and GRK5 in Cardiac Aldosterone Signaling Reveal GRK5-Mediated Cardioprotection via Mineralocorticoid Receptor Inhibition

Aldosterone (Aldo), when overproduced, is a cardiotoxic hormone underlying heart failure and hypertension. Aldo exerts damaging effects via the mineralocorticoid receptor (MR) but also activates the antiapoptotic G protein-coupled estrogen receptor (GPER) in the heart. G protein-coupled receptor (GPCR)-kinase (GRK)-2 and -5 are the most abundant cardiac GRKs and phosphorylate GPCRs as well as non-GPCR substrates. Herein, we investigated whether they phosphorylate and regulate cardiac MR and GPER. To this end, we used the cardiomyocyte cell line H9c2 and adult rat ventricular myocytes (ARVMs), in which we manipulated GRK5 protein levels via clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 and GRK2 activity via pharmacological inhibition. We report that GRK5 phosphorylates and inhibits the cardiac MR whereas GRK2 phosphorylates and desensitizes GPER. In H9c2 cardiomyocytes, GRK5 interacts with and phosphorylates the MR upon β2-adrenergic receptor (AR) activation. In contrast, GRK2 opposes agonist-activated GPER signaling. Importantly, GRK5-dependent MR phosphorylation of the MR inhibits transcriptional activity, since aldosterone-induced gene transcription is markedly suppressed in GRK5-overexpressing cardiomyocytes. Conversely, GRK5 gene deletion augments cardiac MR transcriptional activity. β2AR-stimulated GRK5 phosphorylates and inhibits the MR also in ARVMs. Additionally, GRK5 is necessary for the protective effects of the MR antagonist drug eplerenone against Aldo-induced apoptosis and oxidative stress in ARVMs. In conclusion, GRK5 blocks the cardiotoxic MR-dependent effects of Aldo in the heart, whereas GRK2 may hinder beneficial effects of Aldo through GPER. Thus, cardiac GRK5 stimulation (e.g., via β2AR activation) might be of therapeutic value for heart disease treatment via boosting the efficacy of MR antagonists against Aldo-mediated cardiac injury

Food and Drug Administration Malfunction Recalls of Left Ventricular Assist Devices

Left ventricular assist devices (LVADs) are being increasingly implanted given the increasing prevalence of patients with advanced heart failure stages. However, they are not exempt from device malfunctions. A PubMed search for the key words (left ventricular assist device malfunction) (ventricular assist system malfunction) was performed. We identified 28 publications in the US Food and Drug Administration (FDA) website database that addressed LVAD malfunction. Twenty-nine FDA recalls were identified regarding LVAD malfunctions: 17 regarding HeartWare ventricular assist device, six for HeartMate II, three for HeartMate 3, and three for total artificial heart. Mechanisms involved in LVAD malfunction include battery malfunction, loose driveline connector, malfunction of the system controller, loose power supply connector ports, malfunction of the driveline splice kit, problems with the percutaneous lead connection, disconnection of the bend relief and outflow graft and outflow graft occlusion among others. Multiple mechanisms could be linked to LVAD malfunction. However, multiple device modifications have been developed over the past decade to avoid recurrent malfunctions. Constant improvements and research in biotechnology are needed to prevent these complications. It remains to be seen if newer generation devices will lead to improved patient outcomes over the long term

Carvedilol Selectively Stimulates βArrestin2-Dependent SERCA2a Activity in Cardiomyocytes to Augment Contractility

Heart failure (HF) carries the highest mortality in the western world and β-blockers [β-adrenergic receptor (AR) antagonists] are part of the cornerstone pharmacotherapy for post-myocardial infarction (MI) chronic HF. Cardiac β1AR-activated βarrestin2, a G protein-coupled receptor (GPCR) adapter protein, promotes Sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA)2a SUMO (small ubiquitin-like modifier)-ylation and activity, thereby directly increasing cardiac contractility. Given that certain β-blockers, such as carvedilol and metoprolol, can activate βarrestins and/or SERCA2a in the heart, we investigated the effects of these two agents on cardiac βarrestin2-dependent SERCA2a SUMOylation and activity. We found that carvedilol, but not metoprolol, acutely induces βarrestin2 interaction with SERCA2a in H9c2 cardiomyocytes and in neonatal rat ventricular myocytes (NRVMs), resulting in enhanced SERCA2a SUMOylation. However, this translates into enhanced SERCA2a activity only in the presence of the β2AR-selective inverse agonist ICI 118,551 (ICI), indicating an opposing effect of carvedilol-occupied β2AR subtype on carvedilol-occupied β1AR-stimulated, βarrestin2-dependent SERCA2a activation. In addition, the amplitude of fractional shortening of NRVMs, transfected to overexpress βarrestin2, is acutely enhanced by carvedilol, again in the presence of ICI only. In contrast, metoprolol was without effect on NRVMs’ shortening amplitude irrespective of ICI co-treatment. Importantly, the pro-contractile effect of carvedilol was also observed in human induced pluripotent stem cell (hIPSC)-derived cardiac myocytes (CMs) overexpressing βarrestin2, and, in fact, it was present even without concomitant ICI treatment of human CMs. Metoprolol with or without concomitant ICI did not affect contractility of human CMs, either. In conclusion, carvedilol, but not metoprolol, stimulates βarrestin2-mediated SERCA2a SUMOylation and activity through the β1AR in cardiac myocytes, translating into direct positive inotropy. However, this unique βarrestin2-dependent pro-contractile effect of carvedilol may be opposed or masked by carvedilol-bound β2AR subtype signaling

Biased Agonism/Antagonism at the AngII-AT1 Receptor: Implications for Adrenal Aldosterone Production and Cardiovascular Therapy

Many of the effects of angiotensin II (AngII), including adrenocortical aldosterone release, are mediated by the AngII type 1 receptor (AT 1 R), a receptor with essential roles in cardiovascular homeostasis. AT 1 R belongs to the G protein-coupled receptor (GPCR) superfamily, mainly coupling to the G q/11 type of G proteins. However, it also signals through βarrestins, oftentimes in parallel to eliciting G protein-dependent signaling. This has spurred infinite possibilities for cardiovascular pharmacology, since various beneficial effects are purportedly exerted by AT 1 R via βarrestins, unlike AT 1 R-induced G protein-mediated pathways that usually result in damaging cardiovascular effects, including hypertension and aldosterone elevation. Over the past decade however, a number of studies from our group and others have suggested that AT 1 R-induced βarrestin signaling can also be damaging for the heart, similarly to the G protein-dependent one, with regard to aldosterone regulation. Additionally, AT 1 R-induced βarrestin signaling in astrocytes from certain areas of the brain may also play a significant role in central regulation of blood pressure and hypertension pathogenesis. These findings have provided the impetus for testing available angiotensin receptor blockers (ARBs) in their efficacy towards blocking both routes (i.e. both G protein- and βarrestin-dependent) of AT 1 R signaling in vitro and in vivo and also have promoted structure-activity relationship (SAR) studies for the AngII molecule in terms of βarrestin signaling to certain cellular effects, e.g. adrenal aldosterone production. In the present review, we will recount all of these recent studies on adrenal and astrocyte AT 1 R-dependent βarrestin signaling while underlining their implications for cardiovascular pathophysiology and therapy

Differential Roles of GRK2 and GRK5 in Cardiac Aldosterone Signaling

Background: Heart failure (HF) is the leading cause of death in the western world and new and innovative treatments are urgently needed. Aldosterone (Aldo) is one of several cardio-toxic hormones, whose levels are increased in post-myocardial infarction (MI) HF, and contributes significantly to the morbidity and mortality of the disease. Aldo binds to its cognate receptor, the mineralocorticoid receptor (MR), a nuclear receptor expressed in various tissues including the heart. However, Aldo also exerts MR-independent (its so-called “non-genomic”) actions, which are very rapid, take place at the plasma membrane and do not involve gene transcription. Several of these non-genomic actions of Aldo are mediated by the G protein-coupled receptor 30 (GPR30) or G proteincoupled estrogen receptor (GPER), a plasma membrane-residing G protein-coupled receptor (GPCR). Aldo causes a plethora of detrimental effects in the myocardium, such as abnormal activation of L-type Ca2+ currents and activation of the ryanodine receptor in ventricular myocytes, while elevated levels of Aldo are associated with essential hypertension and can trigger cardiac hypertrophy, fibrosis, and adverse remodeling. Whether these pathophysiological effects of Aldo in the heart result from its non-genomic or genomic (MR-dependent) signaling pathways (or both) is still a matter of debate. Recent studies suggest that cardiac GPER activation by Aldo may actually be beneficial in the myocardium, as it activates the anti-apoptotic extracellular signal-regulated kinases (ERKs). Most GPCRs undergo functional desensitization due to phosphorylation by GPCR-kinases (GRKs), followed by binding of βarrestins which uncouple the receptor from its cognate G proteins. GRKs are a family of seven serine/threonine protein kinases, whose most abundant members in the heart are GRK2 and GRK5. GRK2 upregulation in the failing myocardium is one of the molecular hallmarks of HF and cardiac GRK2 inhibition has already been established as a therapeutic strategy for this disease. In contrast, the role of cardiac GRK5 in the pathophysiology of HF is not as clearly defined. Both GRK2 and -5 are also known to phosphorylate a variety of non-GPCR substrates. Recently, both GRK2 and -5 were shown to get activated by the MR in the heart to induce apoptosis and hypertrophy, respectively. On the other hand, the MR was recently shown to get phosphorylated by unidentified kinase(s) at Ser-843, inside its ligand binding domain, an event that diminishes the MR`s capacity to activate transcription, i.e. essentially inhibits its function. Hypothesis: In the heart, GRK5 phosphorylates the MR at Ser-843 and inhibits its function, whereas GRK2 phosphorylates and desensitizes GPER. Methods: We utilized the rat cardiomyocyte cell line H9c2, which endogenously expresses MR, GPER, GRK2 and GRK5. We performed co-immunoprecipitation experiments to detect GRK-GPER and GRK-MR interactions. We measured protein phosphorylation via western blotting with anti-phosphoserine antibodies. MR transcriptional activity was measured via a luciferase reporter assay. Results: GRK5, but not GRK2, interacts with, and phosphorylates the MR in H9c2 cardiomyocytes, constitutively and in response to beta-adrenoceptor stimulation. In contrast, GRK2, but not GRK5, phosphorylates and desensitizes GPER upon beta-adrenoceptor stimulation in H9c2 cells. The GRK5-phosphorylated MR appears incapable of activating gene transcription, since MR luciferase reporter activity is markedly depressed upon GRK5 overexpression. In contrast, it is markedly enhanced upon GRK5 siRNA-mediated knockdown in H9c2 cells. Conclusions: The pathological MR-dependent Aldo signaling in the heart may be blocked by GRK5, while its beneficial signaling through GPER may be inhibited by GRK2. Thus, a strategy combining GRK5 potentiation with simultaneous GRK2 inhibition may beneficially modulate cardiac Aldo signaling for HF therapy

Bicaval vs biatrial anastomosis techniques in orthotopic heart transplantation: An updated analysis of the UNOS database

Despite the significant increase in the number of orthotopic heart transplants (OHT) performed yearly using the bicaval anastomosis technique, the impact on long-term outcomes remains a topic of debate. We analyzed the United Network for Organ Sharing (UNOS) database in search of the latest insight. We performed a retrospective analysis of the UNOS database from 2006 to 2016 to identify first-time OHT recipients. Patients were primarily stratified according to anastomosis technique: bicaval vs biatrial. Baseline characteristics and clinical status were recorded. The primary endpoint was all-cause mortality. Secondary outcomes included need for permanent pacemaker (PPM), and length of hospital stay (LOS). The Kaplan-Meier method was used to compare survival between the two groups. The Cox proportional hazards regression model was used to conduct multivariable analysis. Statistical significance established at P < .0001. A total of 26 990 patients were identified. Of those who met the inclusion criteria (21 597), 16 573 (77%) underwent bicaval anastomosis. There were no major differences in baseline characteristics between the two groups. The bicaval anastomosis technique was not associated with increased survival during the study period (hazard ratio: 0.97; P = .3557), but the bicaval group required postoperative PPM less often (2.51% vs 5.79%, P < .0001) and was associated with shorter LOS on multivariable analysis. The use of either bicaval or biatrial anastomosis during OHT offers comparable survival advantage. Nonetheless, bicaval anastomosis is associated with less need for postoperative PPM and slightly shorter LOS

Cardiac βarrestin2 Improves Contractility and Adverse Remodeling in Heart Failure, But Is Underexpressed in Humans

In summary, our present study aims to bring the attention of clinicians and pharmacologists to the remarkable functional divergence of the 2 βarrestins in the heart, which, coupled with the virtual absence of the “good” cardiac βarrestin2 protein in humans, highlights potential causes of 2 recent clinical failures of novel, otherwise promising therapies for human HF. Importantly, it points to a “missing link” (boosting endogenous cardiac βarrestin2 levels) for these therapeutics that is necessary to attain efficacy for human HF treatment

Evaluating the impact of chronic obstructive pulmonary disease on in‐hospital outcomes following left ventricular assist device implantation

Background Chronic obstructive pulmonary disease (COPD) is a cause of ventricular dysfunction. However, in the setting of patients with heart failure undergoing left ventricular assist device (LVAD) implantation, there is a paucity of data on the association between COPD and in‐hospital outcomes. Methods and Results Retrospective cohort study based on the NIS including patients ≥18 years who underwent LVAD implantation from 2011 to 2017. Multivariate regression was used to evaluate the impact of COPD on in‐hospital outcomes. A total of 25,503 patients underwent LVAD implantation, of which 13.8% also had COPD. COPD group was older (median 62 vs. 58 years), and more males (82% vs. 76.4%, p < .001 for both). COPD group had more hypertension, diabetes, atrial tachyarrhythmias, dyslipidemia, prior stroke, coronary artery diseases, pulmonary hypertension, and chronic kidney disease (p < .001 for all). No differences in strokes, infections, mechanical circulatory support, and LVAD thrombosis. There was a higher incident of inpatient acute kidney injury, major bleeding, cardiac complications, thromboembolism, and cardiac arrest in patients without COPD (p < .05 for all). Compared with no‐COPD group, COPD group had a lower mortality (6.2% vs. 12.4%; odds ratio, 0.59; confidence interval, 0.512–0.685; p < .05). Conclusion Patients with COPD undergoing LVAD implantation have more comorbidities, without an associated increase mortality