The octapeptide angiotensin II (Ang II) is a circulating hormone as well as a locally formed agonist synthesized by the angiotensin-converting enzyme (ACE) of endothelial cells. It forms a powerful mechanism to control the amount and pressure of body fluids. All main effects are directed to save body salt and water and ensure blood pressure under basic conditions and in emergencies. All blood vessels respond to stimulation by Ang II; the immediate response is smooth muscle contraction, increasing vascular resistance, and elevating blood pressure. Such effects are conveyed by type 1 angiotensin receptors (AT1Rs) located in the plasma membrane of both endothelial and vascular smooth muscle cells. AT1Rs are heterotrimeric G protein-coupled receptors (GPCRs), but their signal pathways are much more complicated than other GPCRs. In addition to Gq/11, the G12/13, JAK/STAT, Jnk, MAPK, and ERK 1/2, and arrestin-dependent and -independent pathways are activated because of the promiscuous attachment of different signal proteins to the intracellular G protein binding site and to the intracellular C terminal loop. Substantial changes in protein expression follow, including the intracellular inflammation signal protein NF-κB, endothelial contact proteins, cytokines, matrix metalloproteinases (MMPs), and type I protocollagen, eliciting the inflammatory transformation of endothelial and vascular smooth muscle cells and fibrosis. Ang II is an important contributor to vascular pathologies in hypertensive, atherosclerotic, and aneurysmal vascular wall remodeling. Such direct vascular effects are reviewed. In addition to reducing blood pressure, AT1R antagonists and ACE inhibitors have a beneficial effect on the vascular wall by inhibiting pathological wall remodeling.

The discovery and characterization of the signal cascades of the β-adrenergic receptors have made it possible to effectively target the receptors for drug development. β-Adrenergic receptors are a class A rhodopsin type of G protein-coupled receptors (GPCRs) that are stimulated mainly by catecholamines and therefore mediate diverse effects of the parasympathetic nervous system in eliciting "fight or flight" type responses. They are detectable in several human tissues where they control a plethora of physiological processes and therefore contribute to the pathogenesis of several disease conditions. Given the relevance of the β-adrenergic receptor as a molecular target for many pathological conditions, this comprehensive review aims at providing an in-depth exploration of the recent advancements in β3-adrenergic receptor research. More importantly, we delve into the prospects of the β3-adrenergic receptor as a therapeutic target across a variety of clinical domains.

G protein-coupled receptors (GPCRs) regulate every aspect of kidney function by mediating the effects of various endogenous and exogenous substances. A key concept in GPCR function is biased signalling, whereby certain ligands may selectively activate specific pathways within the receptor's signalling repertoire. For example, different agonists may induce biased signalling by stabilizing distinct active receptor conformations - a concept that is supported by advances in structural biology. However, the processes underlying functional selectivity in receptor signalling are extremely complex, involving differences in subcellular compartmentalization and signalling dynamics. Importantly, the molecular mechanisms of spatiotemporal bias, particularly its connection to ligand binding kinetics, have been detailed for GPCRs critical to kidney function, such as the AT1 angiotensin receptor (AT1R), V2 vasopressin receptor (V2R) and the parathyroid hormone 1 receptor (PTH1R). This expanding insight into the multifaceted nature of biased signalling paves the way for innovative strategies for targeting GPCR functions; the development of novel biased agonists may represent advanced pharmacotherapeutic approaches to the treatment of kidney diseases and related systemic conditions, such as hypertension, diabetes and heart failure.

Cellular ACE2 (cACE2), a vital component of the renin-angiotensin system (RAS), possesses catalytic activity to maintain AngII and Ang 1-7 balance, which is necessary to prevent harmful effects of AngII/AT2R and promote protective pathways of Ang (1-7)/MasR and Ang (1-7)/AT2R. Hemostasis of the brain-RAS is essential for maintaining normal central nervous system (CNS) function. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a viral disease that causes multi-organ dysfunction. SARS-CoV-2 mainly uses cACE2 to enter the cells and cause its downregulation. This, in turn, prevents the conversion of Ang II to Ang (1-7) and disrupts the normal balance of brain-RAS. Brain-RAS disturbances give rise to one of the pathological pathways in which SARS-CoV-2 suppresses neuroprotective pathways and induces inflammatory cytokines and reactive oxygen species. Finally, these impairments lead to neuroinflammation, neuronal injury, and neurological complications. In conclusion, the influence of RAS on various processes within the brain has significant implications for the neurological manifestations associated with COVID-19. These effects include sensory disturbances, such as olfactory and gustatory dysfunctions, as well as cerebrovascular and brain stem-related disorders, all of which are intertwined with disruptions in the RAS homeostasis of the brain.

Salt sensitivity of blood pressure (SSBP) is an intermediate phenotype of hypertension and is a predictor of long-term cardiovascular events and death. However, the genetic structures of SSBP are uncertain, and it is difficult to precisely diagnose SSBP in population. So, we aimed to identify genes related to susceptibility to the SSBP, construct a risk evaluation model, and explore the potential functions of these genes.

A genome-wide association study of the systemic epidemiology of salt sensitivity (EpiSS) cohort was performed to obtain summary statistics for SSBP. Then, we conducted a transcriptome-wide association study (TWAS) of 12 tissues using FUSION software to predict the genes associated with SSBP and verified the genes with an mRNA microarray. The potential roles of the genes were explored. Risk evaluation models of SSBP were constructed based on the serial P value thresholds of polygenetic risk scores (PRSs), polygenic transcriptome risk scores (PTRSs) and their combinations of the identified genes and genetic variants from the TWAS. The TWAS revealed that 2605 genes were significantly associated with SSBP. Among these genes, 69 were differentially expressed according to the microarray analysis. The functional analysis showed that the genes identified in the TWAS were enriched in metabolic process pathways. The PRSs were correlated with PTRSs in the heart atrial appendage, adrenal gland, EBV-transformed lymphocytes, pituitary, artery coronary, artery tibial and whole blood. Multiple logistic regression models revealed that a PRS of P < 0.05 had the best predictive ability compared with other PRSs and PTRSs. The combinations of PRSs and PTRSs did not significantly increase the prediction accuracy of SSBP in the training and validation datasets.

Several known and novel susceptibility genes for SSBP were identified via multitissue TWAS analysis. The risk evaluation model constructed with the PRS of susceptibility genes showed better diagnostic performance than the transcript levels, which could be applied to screen for SSBP high-risk individuals.

β-arrestins are a family of intracellular signaling proteins that play a key role in regulating the activity of G protein-coupled receptors. The angiotensin-II type 1 receptor is an important G protein-coupled receptor involved in the regulation of cardiovascular function and has been implicated in the progression of cardiovascular diseases. In addition to canonical G protein signaling, G protein-coupled receptors including the angiotensin-II type 1 receptor can signal via β-arrestin. Dysregulation of β-arrestin signaling has been linked to several cardiovascular diseases including hypertension, atherosclerosis, and heart failure. Understanding the role of β-arrestins in these conditions is critical to provide new therapeutic targets for the treatment of cardiovascular disease. In this review, we will discuss the beneficial and maladaptive physiological outcomes of angiotensin-II type 1 receptor-dependent β-arrestin activation in different cardiovascular diseases.

Angiotensin II (AngII), as an octapeptide hormone normally ionized at physiological pH, cannot cross cell membranes and thus, relies on, two (mainly) G protein-coupled receptor (GPCR) types, AT1R and AT2R, to exert its intracellular effects in various organ systems including the cardiovascular one. Although a lot remains to be elucidated about the signaling of the AT2R, AT1R signaling is known to be remarkably versatile, mobilizing a variety of G protein-dependent and independent signal transduction pathways inside cells to produce a biological outcome. Cardiac AT1R signaling leads to hypertrophy, adverse remodeling, fibrosis, while vascular AT1R signaling raises blood pressure via vasoconstriction, but also elicits hypertrophic, vascular growth/proliferation, and pathological remodeling sets of events. In addition, adrenal AT1R is the major physiological stimulus (alongside hyperkalemia) for secretion of aldosterone, a mineralocorticoid hormone that contributes to hypertension, electrolyte abnormalities, and to pathological remodeling of the failing heart. Regulator of G protein Signaling (RGS) proteins, discovered about 25 years ago as GTPase-activating proteins (GAPs) for the Gα subunits of heterotrimeric G proteins, play a central role in silencing G protein signaling from a plethora of GPCRs, including the AngII receptors. Given the importance of AngII and its receptors, but also of several RGS proteins, in cardiovascular homeostasis, the physiological and pathological significance of RGS protein-mediated modulation of cardiovascular AngII signaling comes as no surprise. In the present review, we provide an overview of the current literature on the involvement of RGS proteins in cardiovascular AngII signaling, by discussing their roles in cardiac (cardiomyocyte and cardiofibroblast), vascular (smooth muscle and endothelial cell), and adrenal (medulla and cortex) AngII signaling, separately. Along the way, we also highlight the therapeutic potential of enhancement of, or, in some cases, inhibition of each RGS protein involved in AngII signaling in each one of these cell types.

In traditional Chinese medicine (TCM), the liver is the "general organ" that is responsible for governing/maintaining the free flow of qi over the entire body and storing blood. According to the classic five elements theory, zang-xiang theory, yin-yang theory, meridians and collaterals theory, and the five-viscera correlation theory, the liver has essential relationships with many extrahepatic organs or tissues, such as the mother-child relationships between the liver and the heart, and the yin-yang and exterior-interior relationships between the liver and the gallbladder. The influences of the liver to the extrahepatic organs or tissues have been well-established when treating the extrahepatic diseases from the perspective of modulating the liver by using the ancient classic prescriptions of TCM and the acupuncture and moxibustion. In modern medicine, as the largest solid organ in the human body, the liver has the typical functions of filtration and storage of blood; metabolism of carbohydrates, fats, proteins, hormones, and foreign chemicals; formation of bile; storage of vitamins and iron; and formation of coagulation factors. The liver also has essential endocrine function, and acts as an immunological organ due to containing the resident immune cells. In the perspective of modern human anatomy, physiology, and pathophysiology, the liver has the organ interactions with the extrahepatic organs or tissues, for example, the gut, pancreas, adipose, skeletal muscle, heart, lung, kidney, brain, spleen, eyes, skin, bone, and sexual organs, through the circulation (including hemodynamics, redox signals, hepatokines, metabolites, and the translocation of microbiota or its products, such as endotoxins), the neural signals, or other forms of pathogenic factors, under normal or diseases status. The organ interactions centered on the liver not only influence the homeostasis of these indicated organs or tissues, but also contribute to the pathogenesis of cardiometabolic diseases (including obesity, type 2 diabetes mellitus, metabolic [dysfunction]-associated fatty liver diseases, and cardio-cerebrovascular diseases), pulmonary diseases, hyperuricemia and gout, chronic kidney disease, and male and female sexual dysfunction. Therefore, based on TCM and modern medicine, the liver has the bidirectional interaction with the extrahepatic organ or tissue, and this established bidirectional interaction system may further interact with another one or more extrahepatic organs/tissues, thus depicting a complex "pan-hepatic network" model. The pan-hepatic network acts as one of the essential mechanisms of homeostasis and the pathogenesis of diseases.

The adrenal cortex is responsible for production of adrenal steroid hormones and is anatomically divided into three distinct zones: zona glomerulosa secreting mineralocorticoids (mainly aldosterone), zona fasciculata secreting glucocorticoids (cortisol), and zona reticularis producing androgens. Importantly, due to their high lipophilicity, no adrenal steroid hormone (including aldosterone) is stored in vesicles but rather gets synthesized and secreted instantly upon cell stimulation with specific stimuli. Aldosterone is the most potent mineralocorticoid hormone produced from the adrenal cortex in response to either angiotensin II (AngII) or elevated K+ levels in the blood (hyperkalemia). AngII, being a peptide, cannot cross cell membranes and thus, uses two distinct G protein-coupled receptor (GPCR) types, AngII type 1 receptor (AT1R) and AT2R to exert its effects inside cells. In zona glomerulosa cells, AT1R activation by AngII results in aldosterone synthesis and secretion via two main pathways: (a) Gq/11 proteins that activate phospholipase C ultimately raising intracellular free calcium concentration; and (b) βarrestin1 and -2 (also known as Arrestin-2 and -3, respectively) that elicit sustained extracellular signal-regulated kinase (ERK) activation. Both pathways induce upregulation and acute activation of StAR (steroidogenic acute regulatory) protein, the enzyme that catalyzes the rate-limiting step in aldosterone biosynthesis. This chapter describes these two salient pathways underlying AT1R-induced aldosterone production in zona glomerulosa cells. We also highlight some pharmacologically important notions pertaining to the efficacy of the currently available AT1R antagonists, also known as angiotensin receptor blockers (ARBs) or sartans at suppressing both pathways, i.e., their inverse agonism efficacy at G proteins and βarrestins.

Angiotensin II receptors are members of G protein-coupled receptor superfamily that manifest biased signals toward G protein- and β-arrestin-dependent pathways. However, the role of angiotensin II receptor-biased ligands and the mechanisms underlying myofibroblast differentiation in human cardiac fibroblasts have not been fully elucidated. Our results demonstrated that antagonism of angiotensin II type 1 receptor (AT₁ receptor) and blockade of G_αq protein suppressed angiotensin II (Ang II)-induced fibroblast proliferation, overexpression of collagen I and α-smooth muscle actin (α-SMA), and stress fibre formation, indicating the AT₁ receptor/G_αq axis is necessary for fibrogenic effects of Ang II. Stimulation of AT₁ receptors by their G_αq-biased ligand (TRV120055), but not β-arrestin-biased ligand (TRV120027), substantially exerted fibrogenic effects at a level similar to that of Ang II, suggesting that AT₁ receptor induced cardiac fibrosis in a G_αq-dependent and β-arrestin-independent manner. Valsartan prevents TRV120055-mediated fibroblast activation. TRV120055 mediated the upregulation of transforming growth factor-beta1 (TGF-β1) through the AT₁ receptor/G_αq cascade. In addition, G_αq protein and TGF-β1 were necessary for ERK1/2 activation induced by Ang II and TRV120055. Collectively, TGF-β1 and ERK1/2 are downstream effectors of the G_αq-biased ligand of AT₁ receptor for the induction of cardiac fibrosis.

GPCRs (G protein-coupled receptors) are implicated in blood pressure (BP) and fluid intake regulation. There is a developing concept that these effects are mediated by both canonical G protein signaling and noncanonical β-arrestin mediated signaling, but the contributions of each remain largely unexplored. Here, we hypothesized that β-arrestin contributes to fluid homeostasis and blood pressure (BP) regulation in deoxycorticosterone acetate (DOCA) salt hypertension, a prototypical model of salt-sensitive hypertension.

Global β-arrestin1 (Arrb1) and β-arrestin2 (Arrb2) knockout mice were employed to evaluate drinking behavior, and BP was evaluated in Arrb2-knockout mice. Age- and sex-matched C57BL/6 mice served as controls. We measured intake of water and different sodium chloride solutions and BP employing a 2-bottle choice paradigm with and without DOCA.

Without DOCA (baseline), Arrb2-knockout mice exhibited a significant elevation in saline intake with no change in water intake. With DOCA treatment, Arrb2-knockout mice exhibited a significant increase in both saline and water intake. Although Arrb2-knockout mice exhibited hypernatremia at baseline conditions, we did not find significant changes in total body sodium stores or sodium palatability. In a separate cohort, BP was measured via telemetry in Arrb2-knockout and C57BL/6 mice with and without DOCA. Arrb2-knockout did not exhibit significant differences in BP before DOCA treatment when provided water alone, or when provided a choice of water and saline. However, Arrb2-knockout exhibited an increased pressor response to DOCA-salt.

These findings suggest that in salt-sensitive hypertension, ARRB2, but not ARRB1 (β-arrestin 1), might counterbalance the canonical signaling of GPCRs.

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.

Systolic heart failure (HF) is a chronic clinical syndrome characterized by the reduction in cardiac function and still remains the disease with the highest mortality worldwide. Despite considerable advances in pharmacological treatment, HF represents a severe clinical and social burden. Chronic human HF is characterized by several important neurohormonal perturbations, emanating from both the autonomic nervous system and the adrenal glands. Circulating catecholamines (norepinephrine and epinephrine) and aldosterone elevations are among the salient alterations that confer significant hormonal burden on the already compromised function of the failing heart. This is why sympatholytic treatments (such as β-blockers) and renin-angiotensin system inhibitors or mineralocorticoid receptor antagonists, which block the effects of angiotensin II (AngII) and aldosterone on the failing heart, are part of the mainstay HF pharmacotherapy presently. The adrenal gland plays an important role in the modulation of cardiac neurohormonal stress because it is the source of almost all aldosterone, of all epinephrine, and of a significant amount of norepinephrine reaching the failing myocardium from the blood circulation. Synthesis and release of these hormones in the adrenals is tightly regulated by adrenal G protein-coupled receptors (GPCRs), such as adrenergic receptors and AngII receptors. In this review, we discuss important aspects of adrenal GPCR signaling and regulation, as they pertain to modulation of cardiac function in the context of chronic HF, by focusing on the 2 best studied adrenal GPCR types in that context, adrenergic receptors and AngII receptors (AT 1 Rs). Particular emphasis is given to findings from the past decade and a half that highlight the emerging roles of the GPCR-kinases and the β-arrestins in the adrenals, 2 protein families that regulate the signaling and functioning of GPCRs in all tissues, including the myocardium and the adrenal gland.

Aldosterone (Aldo) breakthrough is a well-known phenomenon that occurs in patients with long-term renin-angiotensin aldosterone system (RAAS) blockade using inhibitors of renin or angiotensin converting enzyme or angiotensin II type 1 receptor blockers. The blockade of the mineralocorticoid receptor (MR), an Aldo binding receptor, is effective in managing patients with resistant hypertension, defined as uncontrollable blood pressure despite the concurrent use of three antihypertensive drugs. In other words, MR inhibitors are not used as first-line antihypertensive drugs in most guidelines for hypertension management. Aldo breakthrough puts hypertensive patients at higher risk of cardiovascular disease and worsens future outcomes. This review discusses Aldo secretion and the mechanism of Aldo breakthrough, dependent or independent of the RAAS, with consideration of the pharmacological aspects of this phenomenon, as well as hypothetical views.

α2-Adrenergic receptors (ARs) mediate many cellular actions of epinephrine and norepinephrine, including inhibition of their secretion (sympathetic inhibition) from adrenal chromaffin cells. Like many other G protein-coupled receptors (GPCRs), they undergo agonist-dependent phosphorylation and desensitization by GPCR kinases (GRKs), a phenomenon recently shown to play a major role in the sympathetic overdrive that accompanies and aggravates chronic heart failure. A three-glutamic acid deletion polymorphism in the human α2B-AR subtype gene (Glu301-303) causes impaired agonist-promoted receptor phosphorylation and desensitization, resulting in enhanced signaling to inhibition of cholinergic-induced catecholamine secretion in adrenal chromaffin cells. One of the various pharmacological assays that can be used to quantify and quantitatively compare the degrees of agonist-dependent desensitization, i.e., G protein decoupling, of these two polymorphic α2B-AR variants (or of any two GPCRs for that matter) is the guanosine-5'-O-3-thiotriphosphate (GTPγS) assay that can directly quantify heterotrimeric G protein activation.

Cis-bifenthrin (cis-BF) is a common-used pyrethroid insecticide frequently detected in environmental and biological matrices. Mounting evidence highlights the endocrine disrupting effects of cis-BF due to anti-estrogenic or anti-androgenic activity. However, little is known about the exposure effects of cis-BF on adrenal cortex function. In this study, effects of cis-BF on biosynthesis of adrenal steroids, as well as the potential mechanisms were investigated in human adrenocortical carcinoma (H295R) cells. Cis-BF decreased basal production levels of cortisol and aldosterone, as well as cAMP-induced production of cortisol. Both he basal and cAMP-stimulated transcriptional levels of several steroidogenic genes were significantly down-regulated by cis-BF. As an important rate-limiting enzyme in steroidogenesis, the protein level of StAR was prohibited by cis-BF on both basal and cAMP-induced conditions. Intracellular level of cAMP was significantly reduced by cis-BF. Overall, these data suggest that cis-BF may inhibit the biosynthesis of cortisol and aldosterone via disrupting cAMP signaling cascade.

The physiological and pathophysiological roles of the angiotensin II type 1 (AT1) receptor, a G protein-coupled receptor ubiquitously expressed throughout the cardiovascular system, have been the focus of intense investigations for decades. The success of angiotensin converting enzyme inhibitors (ACEIs) and of angiotensin receptor blockers (ARBs), which are AT1R-selective antagonists/inverse agonists, in the treatment of heart disease is a testament to the importance of this receptor for cardiovascular homeostasis. Given the pleiotropic signaling of the cardiovascular AT1R and, in an effort to develop yet better drugs for heart disease, the concept of biased signaling has been exploited to design and develop biased AT1R ligands that selectively activate β-arrestin transduction pathways over Gq protein-dependent pathways. However, by focusing solely on Gq or β-arrestins, studies on AT1R "biased" signaling & agonism tend to largely ignore other non-Gq-, non β-arrestin-dependent signaling modalities the very versatile AT1R employs in cardiovascular tissues, including two very important types of signal transducers/regulators: other G protein types (e.g., Gi/o, G12/13) & the Regulator of G protein Signaling (RGS) proteins. In this review, we provide a brief overview of the current state of cardiovascular AT1R biased signaling field with a special focus on the non-Gq-, non β-arrestin-dependent signaling avenues of this receptor in the cardiovascular system, which usually get left out of the conversation of "biased" AT1R signal transduction.

Compelling evidence has revealed that biased activation of G protein-coupled receptor (GPCR) signaling, including angiotensin II (AngII) receptor type 1 (AT1) signaling, plays pivotal roles in vascular homeostasis and injury, but whether a clinically relevant endogenous biased antagonism of AT1 signaling exists under physiological and pathophysiological conditions has not been clearly elucidated. Here, we show that an extracellular matrix protein, cartilage oligomeric matrix protein (COMP), acts as an endogenous allosteric biased modulator of the AT1 receptor and its deficiency is clinically associated with abdominal aortic aneurysm (AAA) development. COMP directly interacts with the extracellular N-terminus of the AT1 via its EGF domain and inhibits AT1-β-arrestin-2 signaling, but not Gq or Gi signaling, in a selective manner through allosteric regulation of AT1 intracellular conformational states. COMP deficiency results in activation of AT1a-β-arrestin-2 signaling and subsequent exclusive AAA formation in response to AngII infusion. AAAs in COMP^-/- or ApoE^-/- mice are rescued by AT1a or β-arrestin-2 deficiency, or the application of a peptidomimetic mimicking the AT1-binding motif of COMP. Explorations of the endogenous biased antagonism of AT1 receptor or other GPCRs may reveal novel therapeutic strategies for cardiovascular diseases.

The mineralocorticoid aldosterone regulates electrolyte and blood volume homeostasis, but it also adversely modulates the structure and function of the chronically failing heart, through its elevated production in chronic human post-myocardial infarction (MI) heart failure (HF). By activating the mineralocorticoid receptor (MR), a ligand-regulated transcription factor, aldosterone promotes inflammation and fibrosis of the heart, while increasing oxidative stress, ultimately induding mitochondrial dysfunction in the failing myocardium. To reduce morbidity and mortality in advanced stage HF, MR antagonist drugs, such as spironolactone and eplerenone, are used. In addition to the MR, aldosterone can bind and stimulate other receptors, such as the plasma membrane-residing G protein-coupled estrogen receptor (GPER), further complicating it signaling properties in the myocardium. Given the salient role that adrenergic receptor (ARs)-particularly βARs-play in cardiac physiology and pathology, unsurprisingly, that part of the impact of aldosterone on the failing heart is mediated by its effects on the signaling and function of these receptors. Aldosterone can significantly precipitate the well-documented derangement of cardiac AR signaling and impairment of AR function, critically underlying chronic human HF. One of the main consequences of HF in mammalian models at the cellular level is the presence of mitochondrial dysfunction. As such, preventing mitochondrial dysfunction could be a valid pharmacological target in this condition. This review summarizes the current experimental evidence for this aldosterone/AR crosstalk in both the healthy and failing heart, and the impact of mitochondrial dysfunction in HF. Recent findings from signaling studies focusing on MR and AR crosstalk via non-conventional signaling of molecules that normally terminate the signaling of ARs in the heart, i.e., the G protein-coupled receptor-kinases (GRKs), are also highlighted.

Heart failure (HF) is a complicated clinical syndrome that is considered an increasingly frequent reason for hospitalization, characterized by a complex therapeutic regimen, reduced quality of life, and high morbidity. Long-standing hypertension ultimately paves the way for HF. Recently, there have been improvements in the treatment of hypertension and overall management not limited to only conventional medications, but several novel pathways and their pharmacological alteration are also conducive to the treatment of hypertension. Beta-arrestin (β-arrestin), a protein responsible for beta-adrenergic receptors' (β-AR) functioning and trafficking, has recently been discovered as a potential regulator in hypertension. β-arrestin isoforms, namely β-arrestin1 and β-arrestin2, mainly regulate cardiac function. However, there have been some controversies regarding the function of the two β-arrestins in hypertension regarding HF. In the present review, we try to figure out the paradox between the roles of two isoforms of β-arrestin in the treatment of HF.

Angiotensin II (AngII) uses two distinct G protein-coupled receptor (GPCR) types, AT1R and AT2R, to exert a plethora of physiologic effects in the body and to significantly affect cardiovascular homeostasis. Although not much is known about the signaling of the AT2R, AT1R signaling is known to be quite pleiotropic, mobilizing a variety of signal transducers inside cells to produce a biological outcome. When the outcome in question is aldosterone production from the adrenal cortex, the main transducers activated specifically by the adrenocortical AT1R to signal toward that cellular effect are the Gq/11 protein alpha subunits and the β-arrestins (also known as Arrestin-2 and -3). The existence of various downstream pathways the AT1R signal can travel down on has led to the ever-expanding filed of GPCR pharmacology termed "biased" signaling, which refers to a ligand preferentially activating one signaling pathway over others downstream of the same receptor in the same cell. However, "biased" signaling or "biased" agonism is therapeutically desirable only when the downstream pathways lead to different or opposite cellular outcomes, so the pathway promoting the beneficial effect can be selectively activated over the pathway that leads to detrimental consequences. In the case of the adrenal AT1R, both Gq/11 proteins and β-arrestins mediate signaling to the same end-result: aldosterone synthesis and secretion. Therefore, both pathways need to remain inactive in the adrenal cortex to fully suppress the production of aldosterone, which is one of the culprit hormones elevated in chronic heart failure, hypertension, and various other cardiovascular diseases. Variations in the effectiveness of the AT1R antagonists, which constitute the angiotensin receptor blocker (ARB) class of drugs (also known as sartans), at the relative blockade of these two pathways downstream of the adrenal AT1R opens the door to the flip term "biased" inverse agonism at the AT1R. ARBs that are unbiased and equipotent inverse agonists for both G proteins and β-arrestins at this receptor, like candesartan and valsartan, are the most preferred agents with the best efficacy at reducing circulating aldosterone, thereby ameliorating heart failure. In the present review, the biased signaling of the adrenal AT1R, particularly in relation to aldosterone production, is examined and the term "biased" inverse agonism at the AT1R is introduced and explained, as a means of pharmacological categorization of the various agents within the ARB drug class.

The treatment of pediatric heart failure is a long-standing unmet medical need. Angiotensin II supports mammalian perinatal circulation by activating cardiac L-type Ca²⁺ channels through angiotensin type 1 receptor (AT₁R) and β-arrestin. TRV027, a β-arrestin-biased AT₁R agonist, that has been reported to be safe but not effective for adult patients with heart failure, activates the AT₁R/β-arrestin pathway. We found that TRV027 evokes a long-acting positive inotropic effect specifically on immature cardiac myocytes through the AT₁R/β-arrestin/L-type Ca²⁺ channel pathway with minimum effect on heart rate, oxygen consumption, reactive oxygen species production, and aldosterone secretion. Thus, TRV027 could be utilized as a valuable drug specific for pediatric heart failure.

Tobacco-related products, containing the highly addictive nicotine together with numerous other harmful toxicants and carcinogens, have been clearly associated with coronary artery disease, heart failure, stroke, and other heart diseases. Among the mechanisms by which nicotine contributes to heart disease is elevation of the renin-angiotensin-aldosterone system (RAAS) activity. Nicotine, and its major metabolite in humans cotinine, have been reported to induce RAAS activation, resulting in aldosterone elevation in smokers. Aldosterone has various direct and indirect adverse cardiac effects. It is produced by the adrenal cortex in response to angiotensin II (AngII) activating AngII type 1 receptors. RAAS activity increases in chronic smokers, causing raised aldosterone levels (nicotine exposure causes the same in rats). AngII receptors exert their cellular effects via either G proteins or the two βarrestins (βarrestin1 and-2).

Since adrenal ßarrestin1 is essential for adrenal aldosterone production and nicotine/cotinine elevate circulating aldosterone levels in humans, we hypothesized that nicotine activates adrenal ßarrestin1, which contributes to RAAS activation and heart disease development.

We studied human adrenocortical zona glomerulosa H295R cells and found that nicotine and cotinine upregulate βarrestin1 mRNA and protein levels, thereby enhancing AngII-dependent aldosterone synthesis and secretion.

In contrast, siRNA-mediated βarrestin1 knockdown reversed the effects of nicotine on AngII-induced aldosterone production in H295R cells. Importantly, nicotine promotes hyperaldosteronism via adrenal βarrestin1, thereby precipitating cardiac dysfunction, also in vivo, since nicotine-exposed experimental rats with adrenal-specific βarrestin1 knockdown display lower circulating aldosterone levels and better cardiac function than nicotine-exposed control animals with normal adrenal βarrestin1 expression.

Adrenal βarrestin1 upregulation is one of the mechanisms by which tobacco compounds, like nicotine, promote cardio-toxic hyperaldosteronism in vitro and in vivo. Thus, adrenal βarrestin1 represents a novel therapeutic target for tobacco-related heart disease prevention or mitigation.

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.

Aldosterone is produced by adrenocortical zona glomerulosa (AZG) cells in response to angiotensin II (AngII) acting through its type I receptors (AT1Rs). AT1R is a G protein-coupled receptor (GPCR) that induces aldosterone via both G proteins and the adapter protein βarrestin1, which binds the receptor following its phosphorylation by GPCR-kinases (GRKs) to initiate G protein-independent signaling. β-adrenergic receptors (ARs) also induce aldosterone production in AZG cells. Herein, we investigated whether GRK2 or GRK5, the two major adrenal GRKs, is involved in the catecholaminergic regulation of AngII-dependent aldosterone production. In human AZG (H295R) cells in vitro, the βAR agonist isoproterenol significantly augmented both AngII-dependent aldosterone secretion and synthesis, as measured by the steroidogenic acute regulatory (StAR) protein and CYP11B2 (aldosterone synthase) mRNA inductions. Importantly, GRK2, but not GRK5, was indispensable for the βAR-mediated enhancement of aldosterone in response to AngII. Specifically, GRK2 inhibition with Cmpd101 abolished isoproterenol's effects on AngII-induced aldosterone synthesis/secretion, whereas the GRK5 knockout via CRISPR/Cas9 had no effect. It is worth noting that these findings were confirmed in vivo, since rats overexpressing GRK2, but not GRK5, in their adrenals had elevated circulating aldosterone levels compared to the control animals. However, treatment with the β-blocker propranolol prevented hyperaldosteronism in the adrenal GRK2-overexpressing rats. In conclusion, GRK2 mediates a βAR-AT1R signaling crosstalk in the adrenal cortex leading to elevated aldosterone production. This suggests that adrenal GRK2 may be a molecular link connecting the sympathetic nervous and renin-angiotensin systems at the level of the adrenal cortex and that its inhibition might be therapeutically advantageous in hyperaldosteronism-related conditions.

To study the ability of a novel bovine serum albumin-angiotensin II (BSA-Ang II) conjugate to effect responses of the AT₁ angiotensin II receptor subtype mediated by the G-protein-coupled and the beta-arrestin pathways.

Angiotensin II (Ang II) was conjugated with bovine serum albumin and compared with Ang II for competition binding to AT₁ receptors, to stimulate aldosterone release from adrenocortical cells, to promote beta-arrestin binding to AT₁ receptors, to promote calcium mobilization, and stimulate drinking of water and saline by rats.

The BSA-Ang II conjugate was less potent competing for AT₁R binding, but was equally efficacious at stimulating aldosterone release from H295R adrenocortical cells. Both BSA-Ang II and Ang II stimulated calcium mobilization and beta-arrestin binding to AT₁ receptors. BSA-Ang II and Ang II stimulated water appetite equivalently but BSA-Ang II stimulated saline appetite more than Ang II. Both BSA-Ang II and Ang II were considerably more potent at causing calcium mobilization than β-arrestin binding.

Addition of a high molecular weight molecule to Ang II reduced its AT₁ receptor binding affinity, but did not significantly alter stimulation of aldosterone release or water consumption. The BSA-Ang II conjugate caused a greater saline appetite than Ang II suggesting that it may be a more efficacious agonist of this beta-arrestin-mediated response than Ang II. The higher potency calcium signaling response suggests that the G-protein-coupled responses predominate at physiological concentrations of Ang II, while the beta-arrestin response requires pathophysiological or pharmacological concentrations of Ang II to occur.

Tobacco kills 6 million people annually and its global health costs are continuously rising. The main addictive component of every tobacco product is nicotine. Among the mechanisms by which nicotine, and its major metabolite, cotinine, contribute to heart disease is the renin-angiotensin-aldosterone system (RAAS) activation. This increases aldosterone production from the adrenals and circulating aldosterone levels. Aldosterone is a mineralocorticoid hormone with various direct harmful effects on the myocardium, including increased reactive oxygen species (ROS) generation, which contributes significantly to cardiac mitochondrial dysfunction and cardiac aging. Aldosterone is produced in the adrenocortical zona glomerulosa (AZG) cells in response to angiotensin II (AngII), activating its type 1 receptor (AT1R). The AT1R is a G protein-coupled receptor (GPCR) that leads to aldosterone biosynthesis and secretion, via signaling from both Gq/11 proteins and the GPCR adapter protein βarrestin1, in AZG cells. Adrenal βarrestin1 is essential for AngII-dependent adrenal aldosterone production, which aggravates heart disease. Since adrenal βarrestin1 is essential for raising circulating aldosterone in the body and tobacco compounds are also known to elevate aldosterone levels in smokers, accelerating heart disease progression, our central hypothesis is that nicotine and cotinine increase aldosterone levels to induce cardiac injury by stimulating adrenal βarrestin1. In the present review, we provide an overview of the current literature of the physiology and pharmacology of adrenal aldosterone production regulation, of the effects of tobacco on this process and, finally, of the effects of tobacco and aldosterone on cardiac structure and function, with a particular focus on cardiac mitochondrial function. We conclude our literature account with a brief experimental outline, as well as with some therapeutic perspectives of our pharmacological hypothesis, that is that adrenal βarrestin1 is a novel molecular target for preventing tobacco-induced hyperaldosteronism, thereby also ameliorating tobacco-related heart disease development.

Agonist-induced internalization of G protein-coupled receptors (GPCRs) is a significant step in receptor kinetics and is known to be involved in receptor down-regulation. However, the dopamine D3 receptor (D3R) has been an exception wherein agonist induces D3Rs to undergo desensitization followed by pharmacological sequestration - which is defined as the sequestration of cell surface receptors into a more hydrophobic fraction within the plasma membrane without undergoing the process of receptor internalization. Pharmacological sequestration renders the receptor in an inactive state on the membrane. In our previous study we demonstrated that a novel class of D3R agonists exemplified by SK608 have biased signaling properties via the G-protein dependent pathway and do not induce D3R desensitization. In this study, using radioligand binding assay, immunoblot or immunocytochemistry methods, we observed that SK608 induced internalization of human D3R stably expressed in CHO, HEK and SH-SY5Y cells which are derived from neuroblastoma cells, suggesting that it is not a cell-type specific event. Further, we have evaluated the potential mechanism of D3R internalization induced by these biased signaling agonists. SK608-induced D3R internalization was time- and concentration-dependent. In comparison, dopamine induced D3R upregulation and pharmacological sequestration in the same assays. GRK2 and clathrin/dynamin I/II are the key molecular players in the SK608-induced D3R internalization process, while β-arrestin 1/2 and GRK-interacting protein 1(GIT1) are not involved. These results suggest that SK608-promoted D3R internalization is similar to the type II internalization observed among peptide binding GPCRs.

The two ubiquitous, outside the retina, G protein-coupled receptor (GPCR) adapter proteins, β-arrestin-1 and -2 (also known as arrestin-2 and -3, respectively), have three major functions in cells: GPCR desensitization, i.e., receptor decoupling from G-proteins; GPCR internalization via clathrin-coated pits; and signal transduction independently of or in parallel to G-proteins. Both β-arrestins are expressed in the heart and regulate a large number of cardiac GPCRs. The latter constitute the single most commonly targeted receptor class by Food and Drug Administration-approved cardiovascular drugs, with about one-third of all currently used in the clinic medications affecting GPCR function. Since β-arrestin-1 and -2 play important roles in signaling and function of several GPCRs, in particular of adrenergic receptors and angiotensin II type 1 receptors, in cardiac myocytes, they have been a major focus of cardiac biology research in recent years. Perhaps the most significant realization coming out of their studies is that these two GPCR adapter proteins, initially thought of as functionally interchangeable, actually exert diametrically opposite effects in the mammalian myocardium. Specifically, the most abundant of the two β-arrestin-1 exerts overall detrimental effects on the heart, such as negative inotropy and promotion of adverse remodeling post-myocardial infarction (MI). In contrast, β-arrestin-2 is overall beneficial for the myocardium, as it has anti-apoptotic and anti-inflammatory effects that result in attenuation of post-MI adverse remodeling, while promoting cardiac contractile function. Thus, design of novel cardiac GPCR ligands that preferentially activate β-arrestin-2 over β-arrestin-1 has the potential of generating novel cardiovascular therapeutics for heart failure and other heart diseases.

Chronic obstructive pulmonary disease (COPD) is a frequent comorbid disease in patients undergoing coronary artery bypass grafting (CABG) surgery, with an incidence ranging from 4% to 20.5%. Conventionally, COPD was recognized as a surgical contraindication to CABG. Because of the recent improvements in surgical techniques, anesthesia, and postoperative management, CABG has been performed more commonly in patients with COPD. However, studies have shown the various effects of COPD on postoperative morbidity and mortality after CABG, and this remains to be well defined.

To compare the postoperative outcomes after CABG between patients with and those without COPD.

A systematic search was conducted in the Cochrane Library, PubMed, EmBase, and Ovid databases (until May 10, 2018). Studies comparing perioperative results and mortality outcomes after CABG between patients with and those without COPD were evaluated independently by 2 reviewers to identify the potentially eligible studies. Review Manager and STATA software were used for statistical analyses.

No significant difference in the mortality rates were found between patients with and those without COPD. COPD was associated with a higher respiratory failure rate (odds ratio [OR] = 4.01; 95% CI: 1.19-13.51, P = .03; P <.001 for heterogeneity), higher pneumonia rate (OR = 2.92; 95% CI: 2.37-3.60, P <.00001; P = .73 for heterogeneity), higher stroke rate (OR = 2.91; 95% CI: 1.37-6.18, P = .005; P = .60 for heterogeneity), higher renal failure rate (OR = 1.60; 95% CI: 1.30-1.97, P <.00001; P = .19 for heterogeneity), and higher wound infection rate (OR = 2.16; 95% CI: 1.21-3.88, P = .01; P = .53 for heterogeneity) after CABG.

Patients with COPD were at higher risks for developing postoperative morbidities, particularly pneumonia, respiratory failure, stroke, renal failure, and wound infection. Although COPD was not associated with a higher risk of mortality, caution should be taken when a patient with COPD is indicated for CABG, considering the higher odds of postoperative complications involving the respiratory system and others.

βarrestin (βarr)-1 and -2 are ubiquitously (outside the retina) expressed G-protein-coupled receptor adapter proteins. They uncouple G-protein-coupled receptors from G proteins, internalize the receptor, and subsequently initiate their own wave of signaling independently of G proteins. Angiotensin (Ang) II type 1 receptor (AT1 R) is a well-established example of a receptor signaling through βarrs. Despite the pivotal role of brain AT1 Rs in the regulation of blood pressure, the involvement of βarr-dependent signaling, mediated by AT1 Rs is not well studied. Particularly, in brain astrocytes very little is known about the effects of βarrs in AT1 R signaling. Herein, we utilized a combination of pharmacological and gene manipulation approaches to investigate the role of βarrs in AT1 R-mediated signaling in isolated brainstem astrocytes from spontaneously hypertensive rats (SHRs) and Wistar rats. We observed that βarr1 is the predominant arrestin isoform at the protein level in these cells. Its expression was down-regulated in SHR astrocytes compared to Wistar rat astrocytes. Ang II, contrary to observations in SHR astrocytes where it had no effect, up-regulates βarr1 protein in Wistar rat astrocytes. We observed differential involvement of βarr1 in MAPK activation in brainstem astrocytes of SHR versus Wistar rats. The βarr-biased agonist peptide [Sar1 , Il4 , Il8] Ang-II (SII), induced AT1 R-mediated ERK and p38 activation in Wistar rat astrocytes. SII had no effect on ERK and p38 activation in SHRs brainstem astrocytes. Our results indicate, reduced involvement of βarr1 in dampening Ang II-induced MAPKs activation and diminished βarr1-mediated ERK and p38 activation in SHR brainstem astrocytes. These findings might be mechanistically related to the development of the brain renin-angiotensin-aldosterone system hyperactivity, which leads to pathogenesis of the hypertensive state of the SHR model.

AT1 angiotensin receptor plays important physiological and pathophysiological roles in the cardiovascular system. Renin-angiotensin system represents a target system for drugs acting at different levels. The main effects of ATR1 stimulation involve activation of Gq proteins and subsequent IP3, DAG, and calcium signaling. It has become evident in recent years that besides the well-known G protein pathways, AT1R also activates a parallel signaling pathway through β-arrestins. β-arrestins were originally described as proteins that desensitize G protein-coupled receptors, but they can also mediate receptor internalization and G protein-independent signaling. AT1R is one of the most studied receptors, which was used to unravel the newly recognized β-arrestin-mediated pathways. β-arrestin-mediated signaling has become one of the most studied topics in recent years in molecular pharmacology and the modulation of these pathways of the AT1R might offer new therapeutic opportunities in the near future. In this paper, we review the recent advances in the field of β-arrestin signaling of the AT1R, emphasizing its role in cardiovascular regulation and heart failure.

Since the launch of the first orally available angiotensin II (AngII) type 1 receptor (AT1R) blocker (ARB) losartan (Cozaar) in the late 1990s, the class of ARBs (or 'sartans', short for Angiotensin-RecepTor-ANtagonistS) quickly expanded to include candesartan, eprosartan, irbesartan, valsartan, telmisartan, and olmesartan. All ARBs have high affinity for the AT1 receptor, expressed in various tissues, including smooth muscle cells, heart, kidney, and brain. Since activation of AT1R, the target of these drugs, leads, among other effects, to vascular smooth muscle cell growth, proliferation and contraction, activation of fibroblasts, cardiac hypertrophy, aldosterone secretion from the adrenal cortex, thirst-fluid intake (hypervolemia), etc., the ARBs are nowadays one of the most useful cardiovascular drug classes used in clinical practice. However, significant differences in their pharmacological and clinical properties exist that may favor use of particular agents over others within the class, and, in fact, two of these drugs, candesartan and valsartan, continuously appear to distinguish themselves from the rest of the 'pack' in recent clinical trials. The reason(s) for the potential superiority of these two agents within the ARB class are currently unclear but under intense investigation. The present short review gives an overview of the clinical properties of the ARBs currently approved by the United States Food and Drug Administration, with a particular focus on candesartan and valsartan and the areas where these two drugs seem to have a therapeutic edge. In the second part of our review, we outline recent data from our laboratory (mainly) on the molecular effects of the ARB drugs on aldosterone production and on circulating aldosterone levels, which may underlie (at least in part) the apparent clinical superiority of candesartan (and valsartan) over most other ARBs currently in clinical use.

The activation of the mitogen-activated protein (MAP) kinases extracellular signal-regulated kinase (ERK)1/2 was traditionally used as a readout of signaling of G protein-coupled receptors (GPCRs) via arrestins, as opposed to conventional GPCR signaling via G proteins. Several recent studies using HEK293 cells where all G proteins were genetically ablated or inactivated, or both non-visual arrestins were knocked out, demonstrated that ERK1/2 phosphorylation requires G protein activity, but does not necessarily require the presence of non-visual arrestins. This appears to contradict the prevailing paradigm. Here we discuss these results along with the recent data on gene edited cells and arrestin-mediated signaling. We suggest that there is no real controversy. G proteins might be involved in the activation of the upstream-most MAP3Ks, although in vivo most MAP3K activation is independent of heterotrimeric G proteins, being initiated by receptor tyrosine kinases and/or integrins. As far as MAP kinases are concerned, the best-established role of arrestins is scaffolding of the three-tiered cascades (MAP3K-MAP2K-MAPK). Thus, it seems likely that arrestins, GPCR-bound and free, facilitate the propagation of signals in these cascades, whereas signal initiation via MAP3K activation may be independent of arrestins. Different MAP3Ks are activated by various inputs, some of which are mediated by G proteins, particularly in cell culture, where we artificially prevent signaling by receptor tyrosine kinases and integrins, thereby favoring GPCR-induced signaling. Thus, there is no reason to change the paradigm: Arrestins and G proteins play distinct non-overlapping roles in cell signaling.

There is heated debate about the benefits of using mineralocorticoid receptor antagonists (MRAs) in addition to standard therapy in patients admitted for myocardial infarction (MI) with or without left ventricular dysfunction (LVD).

Randomized controlled trials (RCTs) were scanned by a formal search of electronic databases (PubMed, EMBASE, Cochrane Library, Ovid, and clinical trials) from their inception to April 2018. A meta-analysis was conducted using Review Manager 5.3 to identify studies reporting the efficacy of MRAs use in post-MI patients with or without LVD.

Thirteen RCTs involving 11,365 individuals were eligible for this study. MRAs treatment reduced all-cause mortality by 16%, cardiovascular death by 16%, and death from heart failure (HF) by 22% in post-MI patients. MRAs use reduced all-cause mortality by 13% and cardiovascular death by 15% in post-MI patients with LVD, but there was no significant difference in all-cause mortality and cardiovascular death in post-MI patients without LVD (relative ratios [RR] 0.83, 95% confidence interval [CI] 0.26-2.69, P = .76, I = 0%; RR 1.01, 95% CI 0.33-3.09, P = .99, I = 0%). In 6 RCTs involving post-MI patients, MRAs treatment had a significant effect on improving left ventricular ejection fraction (LVEF) (mean difference 3.33, 95% CI 0.91-5.75, P = .007, I = 94%). Patients treated with MRAs did not show a decrease in recurrent MI or repeat revascularization compared with patients treated without MRAs (RR 0.95, 95% CI [0.80-1.12], P = .54, I = 0%; RR 1.09, 95% CI [0.79-1.50], P = .61, I = 0%). However, MRAs treatment significantly increased the incidence of hyperkalemia compared with patients treated without MRAs (RR 2.05, 95% CI [1.60, 2.61], P < .00001, I = 49%).

MRAs treatment reduced all-cause mortality, cardiovascular death, and death from HF in post-MI patients. MRAs treatment also demonstrated a significant improvement in LVEF. MRAs reduced cardiovascular death and all-cause mortality in patients with LVD. Eplerenone significantly reduced all-cause mortality and cardiovascular death in post-MI patients. However, MRAs failed to show any cardiovascular benefit in post-MI patients without LVD.

The mineralocorticoid hormone aldosterone regulates sodium and potassium homeostasis but also adversely modulates the maladaptive process of cardiac adverse remodeling post-myocardial infarction. Through activation of its mineralocorticoid receptor (MR), a classic steroid hormone receptor/transcription factor, aldosterone promotes inflammation and fibrosis of the heart, the vasculature, and the kidneys. This is why MR antagonists reduce morbidity and mortality of heart disease patients and are part of the mainstay pharmacotherapy of advanced human heart failure. A plethora of animal studies using cell type⁻specific targeting of the MR gene have established the importance of MR signaling and function in cardiac myocytes, vascular endothelial and smooth muscle cells, renal cells, and macrophages. In terms of its signaling properties, the MR is distinct from nuclear receptors in that it has, in reality, two physiological hormonal agonists: not only aldosterone but also cortisol. In fact, in several tissues, including in the myocardium, cortisol is the primary hormone activating the MR. There is a considerable amount of evidence indicating that the effects of the MR in each tissue expressing it depend on tissue- and ligand-specific engagement of molecular co-regulators that either activate or suppress its transcriptional activity. Identification of these co-regulators for every ligand that interacts with the MR in the heart (and in other tissues) is of utmost importance therapeutically, since it can not only help elucidate fully the pathophysiological ramifications of the cardiac MR's actions, but also help design and develop novel better MR antagonist drugs for heart disease therapy. Among the various proteins the MR interacts with are molecules involved in cardiac G protein-coupled receptor (GPCR) signaling. This results in a significant amount of crosstalk between GPCRs and the MR, which can affect the latter's activity dramatically in the heart and in other cardiovascular tissues. This review summarizes the current experimental evidence for this GPCR-MR crosstalk in the heart and discusses its pathophysiological implications for cardiac adverse remodeling as well as for heart disease therapy. Novel findings revealing non-conventional roles of GPCR signaling molecules, specifically of GPCR-kinase (GRK)-5, in cardiac MR regulation are also highlighted.

Angiotensin II (AngII) type 1 receptor (AT₁R) is a G protein-coupled receptor known for its role in numerous physiological processes and its implication in many vascular diseases. Its functions are mediated through G protein dependent and independent signaling pathways. AT₁R has several endogenous peptidic agonists, all derived from angiotensinogen, as well as several synthetic ligands known to elicit biased signaling responses. Here, surface plasmon resonance (SPR) was used as a cell-based and label-free technique to quantify, in real time, the response of HEK293 cells stably expressing the human AT₁R. The goal was to take advantage of the integrative nature of this assay to identify specific signaling pathways in the features of the response profiles generated by numerous endogenous and synthetic ligands of AT₁R. First, we assessed the contributions of Gq, G12/13, Gi, Gβγ, ERK1/2 and β-arrestins pathways in the cellular responses measured by SPR where Gq, G12/Rho/ROCK together with β-arrestins and ERK1/2 were found to play significant roles. More specifically, we established a major role for G12 in the early events of the AT₁R-dependent response, which was followed by a robust ERK1/2 component associated to the later phase of the signal. Interestingly, endogenous AT₁R ligands (AngII, AngIII and AngIV) exhibited distinct responses signatures with a significant increase of the ERK1/2-like components for both AngIII and AngIV, which points toward possibly distinct physiological roles for the later. We also tested AT₁R biased ligands, all of which affected both the early and later events. Our results support SPR-based integrative cellular assays as a powerful approach to delineate the contribution of specific signaling pathways for a given cell response and reveal response differences associated with ligands with distinct pharmacological properties.

β-arrestin1 (or arrestin2) and β-arrestin2 (or arrestin3) are ubiquitously expressed cytosolic adaptor proteins that were originally discovered for their inhibitory role in G protein-coupled receptor (GPCR) signaling through heterotrimeric G proteins. However, further biochemical characterization revealed that β-arrestins do not just "block" the activated GPCRs, but trigger endocytosis and kinase activation leading to specific signaling pathways that can be localized on endosomes. The signaling pathways initiated by β-arrestins were also found to be independent of G protein activation by GPCRs. The discovery of ligands that blocked G protein activation but promoted β-arrestin binding, or vice-versa, suggested the exciting possibility of selectively activating intracellular signaling pathways. In addition, it is becoming increasingly evident that β-arrestin-dependent signaling is extremely diverse and provokes distinct cellular responses through different GPCRs even when the same effector kinase is involved. In this review, we summarize various signaling pathways mediated by β-arrestins and highlight the physiologic effects of β-arrestin-dependent signaling.

AT₁ angiotensin receptor (AT₁R), a prototypical G protein-coupled receptor (GPCR), is the main receptor, which mediates the effects of the renin-angiotensin system (RAS). AT₁R plays a crucial role in the regulation of blood pressure and salt-water homeostasis, and in the development of pathological conditions, such as hypertension, heart failure, cardiovascular remodeling, renal fibrosis, inflammation, and metabolic disorders. Stimulation of AT₁R leads to pleiotropic signal transduction pathways generating arrays of complex cellular responses. Growing amount of evidence shows that AT₁R is a versatile GPCR, which has multiple unique faces with distinct conformations and signaling properties providing new opportunities for functionally selective pharmacological targeting of the receptor. Biased ligands of AT₁R have been developed to selectively activate the β-arrestin pathway, which may have therapeutic benefits compared to the conventional angiotensin converting enzyme inhibitors and angiotensin receptor blockers. In this review, we provide a summary about the most recent findings and novel aspects of the AT₁R function, signaling, regulation, dimerization or oligomerization and its cross-talk with other receptors, including epidermal growth factor (EGF) receptor, adrenergic receptors and CB₁ cannabinoid receptor. Better understanding of the mechanisms and structural aspects of AT₁R activation and cross-talk can lead to the development of novel type of drugs for the treatment of cardiovascular and other diseases.