Novel roles for G protein-coupled receptor kinases in cardiac injury and repair.

G protein-coupled receptors (GPCRs) are key modulators of cell signaling. Multiple GPCRs are present in the heart where they regulate cardiac homeostasis including processes such as myocyte contraction, heart rate and coronary blood flow. GPCRs are pharmacological targets for several cardiovascular disorders including heart failure (HF) such as beta-adrenergic receptor (ßAR) blockers and angiotensin II receptor (AT1R) antagonists. The activity of GPCRs are finely regulated by GPCR kinases (GRKs), which phosphorylate agonist-occupied receptors and start the process of desensitization. Among the seven members of the GRK family, GRK2 and GRK5 are predominantly expressed in the heart, where they exhibit both canonical and non-canonical functions. Both kinases are known to be increased in cardiac pathologies and contribute to pathogenesis through their roles in different cellular compartments. Lowering or inhibiting their actions mediate cardioprotective effects against pathological cardiac growth and failing heart. Therefore, given their importance in cardiac dysfunction, these kinases are drawing attention as promising targets for the treatment of HF, which needs improved therapies. Over the past three decades, broad knowledge on GRK inhibition in HF has been gained by studies using genetically engineered animal models or through gene therapy with peptide inhibitors or using small molecule inhibitors. In this mini review, we summarize the work focusing on GRK2 and GRK5 but also discuss a couple of the non-abundant cardiac subtypes and their multi-functional roles in the normal and diseased heart and the potential and therapeutic targets.

Sex Differences in Glucose Homeostasis.

Sexual dimorphism has been demonstrated to have an effect on various physiological functions. In this regard, researchers have investigated its impact on glucose homeostasis in both preclinical and clinical studies. Sex differences mainly arise from physiological factors such as sex hormones, body fat and muscle distribution, and sex chromosomes. The sexual dimorphism has also been studied in the context of diabetes. Reflecting the prevalence of the disease among the population, studies focusing on the sex difference in type 1 diabetes (T1D) are not common as the ones in type 2 diabetes (T2D). T1D is reported as the only major specific autoimmune disease that exhibits a male predominance. Clinical studies have demonstrated that impaired fasting glucose is more frequent in men whereas women more commonly exhibit impaired glucose tolerance. Understanding the sex difference in glucose homeostasis becomes more attractive when focusing on the findings that highlight sexual dimorphism on the efficacy or adverse effect profile of antidiabetic medications. Thus, in this chapter, we aimed to discuss the impact of sex on the glucose homeostasis both in health and in diabetes.

Nitric Oxide and S-Nitrosylation in Cardiac Regulation: G Protein-Coupled Receptor Kinase-2 and ß-Arrestins as Targets.

Cardiac diseases including heart failure (HF), are the leading cause of morbidity and mortality globally. Among the prominent characteristics of HF is the loss of ß-adrenoceptor (AR)-mediated inotropic reserve. This is primarily due to the derangements in myocardial regulatory signaling proteins, G protein-coupled receptor (GPCR) kinases (GRKs) and ß-arrestins (ß-Arr) that modulate ß-AR signal termination via receptor desensitization and downregulation. GRK2 and ß-Arr2 activities are elevated in the heart after injury/stress and participate in HF through receptor inactivation. These GPCR regulators are modulated profoundly by nitric oxide (NO) produced by NO synthase (NOS) enzymes through S-nitrosylation due to receptor-coupled NO generation. S-nitrosylation, which is NO-mediated modification of protein cysteine residues to generate an S-nitrosothiol (SNO), mediates many effects of NO independently from its canonical guanylyl cyclase/cGMP/protein kinase G signaling. Herein, we review the knowledge on the NO system in the heart and S-nitrosylation-dependent modifications of myocardial GPCR signaling components GRKs and ß-Arrs.

Role of the ß3-adrenergic receptor subtype in catecholamine-induced myocardial remodeling.

ß3-Adrenoceptors (AR) stimulate cardiac Na+/K+ pump in healthy hearts. ß3-ARs are upregulated by persistent sympathetic hyperactivity; however, their effect on Na+/K+ ATPase activity and ventricular function in this condition is still unknown. Here, we investigate preventive effects of additional ß3-AR activation (BRL) on Na+/K+ ATPase activity and in vivo hemodynamics in a model of noradrenaline-induced hypertrophy. Rats received NA or NA plus simultaneously administered BRL in vivo infusion for 14 days; their cardiac function was investigated by left ventricular pressure-volume analysis. Moreover, fibrosis and apoptosis were also assessed histologically. NA induced an hypertrophic pattern, as detected by morphological, histological, and biochemical markers. Additional BRL exposure reversed the hypertrophic pattern and restored Na+/K+ ATPase activity. NA treatment increased systolic function and depressed diastolic function (slowed relaxation). Additional BRL treatment reversed most NA-induced hemodynamic changes. NA decreased Na+/K+ pump α2 subunit expression selectively, a change also reversed by additional BRL treatment. Increasing ß3-AR stimulation may prevent the consequences of chronic NA exposure on Na+/K+ pump and in vivo hemodynamics. ß3-AR agonism may thus represent a new therapeutic strategy for pharmacological modulation of hypertrophy under conditions of chronically enhanced sympathetic activity.

Onset of decreased heart work is correlated with increased heart rate and shortened QT interval in high-carbohydrate fed overweight rats.

Mechanical activity of the heart is adversely affected in metabolic syndrome (MetS) characterized by increased body mass and marked insulin resistance. Herein, we examined the effects of high carbohydrate intake on cardiac function abnormalities by evaluating in situ heart work, heart rate, and electrocardiograms (ECGs) in rats. MetS was induced in male Wistar rats by adding 32% sucrose to drinking water for 22-24 weeks and was confirmed by insulin resistance, increased body weight, increased blood glucose and serum insulin, and increased systolic and diastolic blood pressures in addition to significant loss of left ventricular integrity and increased connective tissue around myofibrils. Analysis of in situ ECG recordings showed a markedly shortened QT interval and decreased QRS amplitude with increased heart rate. We also observed increased oxidative stress and decreased antioxidant defense characterized by decreases in serum total thiol level and attenuated paraoxonase and arylesterase activities. Our data indicate that increased heart rate and a shortened QT interval concomitant with higher left ventricular developed pressure in response to ß-adrenoreceptor stimulation as a result of less cyclic AMP release could be regarded as a natural compensation mechanism in overweight rats with MetS. In addition to the persistent insulin resistance and obesity associated with MetS, one should consider the decreased heart work, increased heart rate, and shortened QT interval associated with high carbohydrate intake, which may have more deleterious effects on the mammalian heart.

ß3-Adrenoceptor-mediated responses in diabetic rat heart.

ß3-adrenoceptors mediate negative inotropic effect in contrast to classical ß1- and ß2-adrenoceptors. Cardiac ß3-adrenoceptors are upregulated in experimental diabetes. Thus, cardiodepressant effect mediated by ß3-adrenoceptors has been proposed to contribute to the impaired cardiac function in this pathology. In our study, we investigated the influence of streptozotocin-diabetes on cardiac contractility to ß3-adrenoceptors stimulation by using Langendorff-perfused rat hearts. BRL 37344, a selective ß3-adrenoceptor agonist, induced dose-dependent decreases in left ventricular developed pressure (LVDP) in hearts from control rats. BRL 37344 also dose-dependently decreased +dP/dt and -dP/dt values. Effects of BRL 37344 were abolished by SR 59230, but not altered by nadolol pre-treatment. On the other hand, these effects of BRL 37344 were all significantly increased in hearts from diabetic rats. We also observed that diabetes significantly increased the mRNA levels encoding cardiac ß3-adrenoceptors. In addition, Giα2 mRNA expressions were found to be increased in the cardiac tissue of diabetic rats as well. The effect of BRL 37344 on cardiac contractility was normalized upon treatment of diabetic rats with insulin. These data demonstrate an increased effect of ß3-adrenoceptor stimulation on hemodynamic function of the heart in accordance with an increased mRNA levels encoding cardiac ß3-adrenoceptors in 8-week diabetic rats.

A year in pharmacology: new drugs approved by the US Food and Drug Administration in 2023.

With 54 new drugs and seven cellular and gene therapy products, the approvals by the US Food and Drug Administration (FDA) recovered 2023 from the 2022 dent back to the levels of 2020-2021. As in previous years of this annual review, we assign these new drugs to one of three levels of innovation: first drug against a condition ("first-in-indication"), first drug using a novel molecular mechanism ("first-in-class"), and "next-in-class," i.e., a drug using an already exploited molecular mechanism. We identify four (7%) "first-in-indication," 22 (36%) "first-in-class," and 35 (57%) "next-in-class" drugs. By treatment area, rare diseases (54%) and cancer drugs (23%) were once again the most prevalent (and partly overlapping) therapeutic areas. Other continuing trends were the use of accelerated regulatory approval pathways and the reliance on biopharmaceuticals (biologics). 2023 marks the approval of a first therapy based on CRISPR/Cas9 gene editing.

Nebivolol prevents desensitization of ß-adrenoceptor signaling and induction of cardiac hypertrophy in response to isoprenaline beyond ß1-adrenoceptor blockage.

The importance of chronic stimulation of ß-adrenoceptors in the development of cardiac dysfunction is the rationale for the use of ß-blockers in the treatment of heart failure. Nebivolol is a third-generation ß-blocker, which has further properties including stimulation of endothelial nitric oxide synthase and/or ß3-adrenoceptors. The aim of this study was to investigate whether nebivolol has additional effects on ß-adrenoceptor-mediated functional responses along with morphologic and molecular determinants of cardiac hypertrophy compared with those of metoprolol, a selective ß1-adrenoceptor blocker. Rats infused by isoprenaline (100 µg·kg(-1)·day(-1), 14 days) were randomized into three groups according to the treatment with metoprolol (30 mg·kg(-1)·day(-1)), nebivolol (10 mg·kg(-1)·day(-1)), or placebo for 13 days starting on day 1 after implantation of minipump. Both metoprolol and nebivolol caused a similar reduction on heart rate. Nebivolol mediated a significant improvement on cardiac mass, coronary flow, mRNA expression levels of sarcoplasmic reticulum Ca(2+) ATPase (SERCA2a) and atrial natriuretic peptide and phospholamban (PLN)/SERCA2a and phospho-PLN/PLN ratio compared with metoprolol and placebo. Nebivolol prevented the detrimental effects of isoprenaline infusion on isoprenaline (68% of control at 30 µM), BRL37344 (63% of control at 0.1 µM), and forskolin (64% of control at 1 µM) responses compared with metoprolol (isoprenaline, 34% of control; BRL37344, no response; forskolin, 26% of control) and placebo (isoprenaline, 33% of control; BRL37344, 28% of control; forskolin, 12% of control). Both ß-blockers improved the changes in mRNA expressions of ß1- and ß3-adrenoceptors. Our results suggest that nebivolol partially protects the responsiveness of ß-adrenoceptor signaling and the development of cardiac hypertrophy independent of its ß1-adrenoceptor blocking effect.

A year in pharmacology: new drugs approved by the US Food and Drug Administration in 2022.

While new drug approvals by the U.S. Food and Drug Administration (FDA) had remained stable or even increased in the first 2 years of the COVID-19 pandemic, the 37 newly approved drugs in 2022 are considerably less than the 53 and 50 new drugs approved in 2020 and 2021, respectively, and less than the rolling 10-year average of 43. As in previous years of this annual review, we assign these new drugs to one of three levels of innovation: first drug against a condition ("first-in-indication"), first drug using a novel molecular mechanism ("first-in-class"), and "next-in-class," i.e., a drug using an already exploited molecular mechanism. We identify two "first-in-indication" (ganaxolon and teplizumab), 20 (54%) "first-in-class," and 17 (46%) "next-in-class" drugs. By treatment area, rare diseases and cancer drugs were once again the most prevalent (partly overlapping) therapeutic areas. Other continuing trends were the use of accelerated regulatory approval pathways and the reliance on biopharmaceuticals (biologics).

A year in pharmacology: new drugs approved by the US Food and Drug Administration in 2021.

The second year of the COVID-19 pandemic had no adverse effect on the number of new drug approvals by the US Food and Drug Administration (FDA). Quite the contrary, with a total of 50 new drugs, 2021 belongs to the most successful FDA years. We assign these new drugs to one of three levels of innovation: (1) first drug against a condition ("first-in-indication"), (2) first drug using a novel molecular mechanism ("first-in-class"), and (3) "next-in-class", i.e., a drug using an already exploited molecular mechanism. We identify 21 first-in-class, 28 next-in-class, and only one first-in-indication drugs. By treatment area, the largest group is once again cancer drugs, many of which target specific genetic alterations. Every second drug approved in 2021 targets an orphan disease, half of them being cancers. Small molecules continue to dominate new drug approvals, followed by antibodies and non-antibody biopharmaceuticals. In 2021, the FDA continued to approve drugs without strong evidence of clinical effects, best exemplified by the aducanumab controversy.

A year in pharmacology: new drugs approved by the US Food and Drug Administration in 2020.

While the COVID-19 pandemic also affected the work of regulatory authorities, the US Food and Drug Administration approved a total of 53 new drugs in 2020, one of the highest numbers in the past decades. Most newly approved drugs related to oncology (34%) and neurology (15%). We discuss these new drugs by level of innovation they provide, i.e., first to treat a condition, first using a novel mechanisms of action, and "others." Six drugs were first in indication, 15 first using a novel mechanism of action, and 32 other. This includes many drugs for the treatment of orphan indications and some for the treatment of tropical diseases previously neglected for commercial reasons. Small molecules continue to dominate new drug approvals, followed by antibodies. Of note, newly approved drugs also included small-interfering RNAs and antisense oligonucleotides. These data show that the trend for declines in drug discovery and development has clearly been broken.

Cardiac and Vascular α1-Adrenoceptors in Congestive Heart Failure: A Systematic Review.

As heart failure (HF) is a devastating health problem worldwide, a better understanding and the development of more effective therapeutic approaches are required. HF is characterized by sympathetic system activation which stimulates α- and ß-adrenoceptors (ARs). The exposure of the cardiovascular system to the increased locally released and circulating levels of catecholamines leads to a well-described downregulation and desensitization of ß-ARs. However, information on the role of α-AR is limited. We have performed a systematic literature review examining the role of both cardiac and vascular α1-ARs in HF using 5 databases for our search. All three α1-AR subtypes (α1A, α1B and α1D) are expressed in human and animal hearts and blood vessels in a tissue-dependent manner. We summarize the changes observed in HF regarding the density, signaling and responses of α1-ARs. Conflicting findings arise from different studies concerning the influence that HF has on α1-AR expression and function; in contrast to ß-ARs there is no consistent evidence for down-regulation or desensitization of cardiac or vascular α1-ARs. Whether α1-ARs are a therapeutic target in HF remains a matter of debate.

Cardiac ß3 -adrenoceptors-A role in human pathophysiology?

As ß3 -adrenoceptors were first demonstrated to be expressed in adipose tissue they have received much attention for their metabolic effects in obesity and diabetes. After the existence of this subtype had been suggested to be present in the heart, studies focused on its role in cardiac function. While the presence and functional role of ß3 -adrenoceptors in the heart has not uniformly been detected, there is a broad consensus that they become up-regulated in pathological conditions associated with increased sympathetic activity such as heart failure and diabetes. When detected, the ß3 -adrenceptor has been demonstrated to mediate negative inotropic effects in an inhibitory G protein-dependent manner through the NO-cGMP-PKG signalling pathway. Whether these negative inotropic effects provide protection from the adverse effects induced by overstimulation of ß1 /ß2 -adrenoceptors or in themselves are potentially harmful is controversial, but ongoing clinical studies in patients with congestive heart failure are testing the hypothesis that ß3 -adrenceptor agonism has a beneficial effect. LINKED ARTICLES: This article is part of a themed section on Adrenoceptors-New Roles for Old Players. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.14/issuetoc.

Beta-3 adrenoceptors: A potential therapeutic target for heart disease.

As heart failure (HF) is a growing public health problem worldwide, rapid therapeutic development is required to improve HF management. Decreased myocardial contractility in HF is associated with the persistent sympathetic activation of ß1/ß2-adrenoceptors (ß1/ß2-ARs). Although it is initially activated to compensate for a decline in myocardial contractility, it plays a pivotal role in organ damage and functional deterioration over time, resulting in the desensitization of receptors involved. The third ß-AR subtype, ß3-AR, is resistant to desensitization, and as a result, the expression of this subtype is enhanced in human failing myocardium. In addition, this upregulation and the stimulation of this subtype have been demonstrated to mediate cardioprotective effects such as antihypertrophic, antioxidant and antifibrotic effects via various signaling pathways in different cell types. However, the role of this attractive therapeutic intervention in heart diseases must be clarified through clinical trials.

Aging related functional and structural changes in the heart and aorta: MitoTEMPO improves aged-cardiovascular performance.

Aging in humans represents declining in cardio-protective systems, however its mechanisms are not known yet. We aimed to analyse how aging affects key mechanisms responsible for contractile dysfunction via comparing the improperly synchrony between electrical and mechanical activities in male aged-rats (24-month old) comparison to those of adult-rats (6-month old). We determined significantly increased systemic oxidative stress with decreased antioxidant capacity, clear insulin resistance and hypertrophy in aged-rats with normal fasting blood glucose. We also determined significantly high level of reactive oxygen species, ROS production in fluorescent dye chloromethyl-2',7'-dichlorodihydrofluoroscein diacetate (DCFDA) loaded isolated cardiomyocytes from aged-rats, confirming the increased oxidative stress in these hearts. In situ electrocardiograms, ECGs presented significant prolongations in RR- and QT-intervals in the aged-rats. Invasive hemodynamic measurements demonstrated marked increases in the heart rate and mean arterial pressure and decreases in the ejection-fraction and preload-recruitable stroke-work, together with depressed contraction and relaxation activities in aortic rings. In light and electron microscopy examinations in aged-rats, significant increases in muscle fibre radius and amount of collagen fibres were detected in the heart as well as markedly flattened and partial local splitting in elastic lamellas in the aorta, besides irregularly clustered mitochondria and lysosomes around the myofilaments in cardiomyocytes. MitoTEMPO treatment of tissue samples and cardiomyocytes from aged-rats for 1-h induced significant structural improvements. In the second part of our study, we have shown that mitochondria-targeted antioxidant MitoTEMPO antagonized all alterations in the heart samples as well as penylephrine-induced contractile and acetylcholine-induced relaxation responses of aged-rat aortic rings. Overall, the present data strongly support the important role of mitochondrial oxidative stress in the development of aged-related insufficiencies and that antioxidant strategies specifically targeting this organelle could have therapeutic benefit in aging-associated complications.

The role of insulin-thyroid hormone interaction on ß-adrenoceptor-mediated cardiac responses.

ß-adrenoceptor-mediated responses are known to be attenuated in diabetic rat hearts, related to decreased receptor sensitivity and density. These impaired responses were improved with insulin in diabetic rats, but not in thyroidectomized diabetic rats. We aimed to investigate the possible interaction between insulin and thyroid hormones to restore diabetes-induced alterations on ß-adrenoceptor-mediated responses. Male Sprague-Dawley rats were divided into seven groups: control (C), diabetic (D), insulin-treated diabetic (DI), thyroidectomized diabetic (TxD), insulin-treated thyroidectomized diabetic (TxDI), insulin+low dose 3,3',5-triiodo-L-thyronine (T3) treated (TxDIT2.5) or insulin+high dose T3 (TxDIT5) treated thyroidectomized diabetic rats. Diabetes was induced with 38 mg/kg streptozotocin. Cardiac function was assessed through pressure-volume analysis and papillary muscle experiments. QPCR and western blot experiments were performed to evaluate cardiac gene expressions. Hemodynamic parameters were impaired in diabetes, and were mostly corrected in DI and TxDIT5 groups. Isoprenaline- and BRL37344-induced contractile responses were also decreased in diabetes. Isoprenaline responses were improved significantly in DI and TxDIT5 groups, whereas BRL 37344-mediated responses were increased slightly. Reduced ß1-adrenoceptor and SERCA 2A mRNA levels in diabetes were corrected in DI and TxDIT5 groups. Decreased SERCA 2A and increased ß3-adrenoceptor protein levels in diabetes were improved in DI and TxDIT5 groups. No significant changes were found in phospholamban or endothelial nitricoxide synthase protein levels. These results show that the beneficial effects of insulin on ß-adrenoceptor-mediated responses in diabetic rats are dependent upon adequate concentrations of thyroid hormones.