Correction: Maning et al. Antagonistic Roles of GRK2 and GRK5 in Cardiac Aldosterone Signaling Reveal GRK5-Mediated Cardioprotection via Mineralocorticoid Receptor Inhibition. Int. J. Mol. Sci. 2020, 21, 2868.

The authors wish to make the following correction to this paper [...].

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 ß2-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. ß2AR-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 ß2AR activation) might be of therapeutic value for heart disease treatment via boosting the efficacy of MR antagonists against Aldo-mediated cardiac injury.

GRK2-Mediated Crosstalk Between ß-Adrenergic and Angiotensin II Receptors Enhances Adrenocortical Aldosterone Production In Vitro and In Vivo.

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 ßarrestin1, which binds the receptor following its phosphorylation by GPCR-kinases (GRKs) to initiate G protein-independent signaling. ß-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 ßAR 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 ßAR-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 ß-blocker propranolol prevented hyperaldosteronism in the adrenal GRK2-overexpressing rats. In conclusion, GRK2 mediates a ßAR-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.

A case report: Pharmacology and resistance patterns of three generations of ALK inhibitors in metastatic inflammatory myofibroblastic sarcoma.

BACKGROUND: Little exists currently in research about the mechanisms of resistance of ALK inhibitors in inflammatory myofibroblastic sarcoma. It is known, however, that ALK gene rearrangements are common in inflammatory myofibroblastic tumors, similar to non-small cell lung cancer. In roughly 50% of inflammatory myofibroblastic tumors, gene rearrangement has been found to occur on chromosome 2 at band 2p23. In non-small cell lung cancer, it has been shown that about a third of patients who progress on the first generation ALK inhibitor, crizotinib develops mutations in the ALK kinase domain. The remaining two-thirds of patients tend to develop amplification of ALK or activation of alternative signaling pathways. Chromoplexy has also been described as a mechanism of resistance, where multiple closed chain rearrangements cause loss-of-function of tumor suppressor genes and gain-in-function of oncogenic fusions. Partner genes that have been identified in IMTs are tropomyosin 3 (TPM3), tropomyosin 4 (TPM4), clathrin heavy chain (CLTC), Ran-binding protein 2 (RANBP2), cysteinyl-tRNA synthetase (CARS), 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase (ATIC), and SEC31L1. All are active promoters for the fusion gene, in response to NPM binding. Several inflammatory myofibroblastic tumor case reports indicated that fusion of ALK and RANBP2 led to a more aggressive clinical course. Although the majority of inflammatory myofibroblastic tumor case reports have utilized first and second generation ALK inhibitors, all generations of ALK inhibitors have demonstrated some ability to impair disease progression and extend life expectancy. However, at some point in the course of therapy with each generation of ALK inhibitor, resistance ultimately developed. In order to better understand the pharmacology and resistance patterns behind three generations of ALK inhibitors, we sought to examine a patient with metastatic anaplastic lymphoma kinase-1-rearranged inflammatory myofibroblastic sarcoma to the brain. We also explored the similarities and differences of this clinical case to other inflammatory myofibroblastic sarcoma case reports involving the use of ALK inhibitors. CASE REPORT: A rare case of pulmonary IMS with ALK-1 gene rearrangement and multiple brain metastases responded to three generations of ALK inhibitors. However, similar to other case reports, due to the development of resistance and recurrence, the patient eventually succumbed to the disease. CONCLUSIONS: ALK inhibitors are beneficial in the temporary prevention of progression of disease in patients with inflammatory myofibroblastic tumors. In this case, due to the inability to reveal the fusion partner in this patient via DNA sequencing, it is unknown exactly if that partner was RANBP2 or another ALK partner gene. Brain biopsy tissue was also unobtainable during sequence of ALK due to risk versus benefit, which would have provided insight as which type of ALK resistance mutations the patient was developing. It is likely that this patient had some form of chromoplexy occurring.

Deletion of Osteopontin Enhances ß2-Adrenergic Receptor-Dependent Anti-Fibrotic Signaling in Cardiomyocytes.

Cardiac ß2-adrenergic receptors (ARs) are known to inhibit collagen production and fibrosis in cardiac fibroblasts and myocytes. The ß2AR is a Gs protein-coupled receptor (GPCR) and, upon its activation, stimulates the generation of cyclic 3',5'-adenosine monophosphate (cAMP). cAMP has two effectors: protein kinase A (PKA) and the exchange protein directly activated by cAMP (Epac). Epac1 has been shown to inhibit cardiac fibroblast activation and fibrosis. Osteopontin (OPN) is a ubiquitous pro-inflammatory cytokine, which also mediates fibrosis in several tissues, including the heart. OPN underlies several cardiovascular pathologies, including atherosclerosis and cardiac adverse remodeling. We found that the cardiotoxic hormone aldosterone transcriptionally upregulates OPN in H9c2 rat cardiac myoblasts-an effect prevented by endogenous ß2AR activation. Additionally, CRISPR-mediated OPN deletion enhanced cAMP generation in response to both ß1AR and ß2AR activation in H9c2 cardiomyocytes, leading to the upregulation of Epac1 protein levels. These effects rendered ß2AR stimulation capable of completely abrogating transforming growth factor (TGF)-ß-dependent fibrosis in OPN-lacking H9c2 cardiomyocytes. Finally, OPN interacted constitutively with Gαs subunits in H9c2 cardiac cells. Thus, we uncovered a direct inhibitory role of OPN in cardiac ß2AR anti-fibrotic signaling via cAMP/Epac1. OPN blockade could be of value in the treatment and/or prevention of cardiac fibrosis.

Novel Insights into the Crosstalk between Mineralocorticoid Receptor and G Protein-Coupled Receptors in Heart Adverse Remodeling and Disease.

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.

Mannitol Anaphylaxis in the Setting of Septic Emboli-Induced Intracranial Hemorrhage.

Neurological complications are a significant problem in bacterial endocarditis. Cerebral embolism is the most frequent concern. Acute embolic disease may trigger focal seizures or mycotic aneurysms. Miliary infection is also common, and lumbar puncture can guide in determining the infective organism. Purulent cerebrospinal fluid (CSF) consists often of Staphylococcus aureus, a virulent organism, whereas non-virulent organisms (i.e., viridans streptococci) have normal CSF formulae. Microscopic abscesses suggest the potential for aneurysm from bacterial endocarditis amplifying the risk of intracranial hemorrhage. Mannitol and hypertonic (3%) saline are intravenous medications used as a rescue treatment for brain hemorrhage. A patient diagnosed with mycoplasma pneumonia and septic shock secondary to tricuspid endocarditis with extensive pulmonary emboli and metastatic infection to his spine was initiated on antibiotics. He developed a massive intracranial bleed from the rupture of mycotic septic emboli and was given mannitol to decrease intracranial pressure, which caused anaphylaxis.

Calcium Alkali Syndrome Treated With Hemodialysis.

Malignancy, primary hyperparathyroidism, and vitamin D intoxication are the most common causes of hypercalcemia. Symptoms of hypercalcemia are nonspecific and require a plasma calcium level to diagnose. Undiagnosed hypercalcemia can cause renal failure long-term. Here, we describe a unique case of hypercalcemia resulting in acute kidney injury (AKI) secondary to overconsumption of calcium carbonate (Tums).

Bleeding After a Single Dose of Ketorolac in a Postoperative Patient.

A 74-year-old male patient with extensive cardiac history and hepatitis on apixaban and clopidogrel presented with adhesive small bowel obstruction. The patient underwent an exploratory laparotomy and adhesiolysis for small bowel obstruction. On postoperative day 6, after a single dose of ketorolac, the patient had an intra-abdominal bleed requiring exploratory laparotomy and washout.

Methotrexate for Cornual Ectopic Pregnancy.

A 27-year-old pregnant female presented to the emergency department with pelvic pain and vaginal bleeding. At the time of diagnosis, she had an ectopic pregnancy in the right cornua 4 cm in size with a human chorionic gonadotropin (hCG) value of 14,438 international units per mL. The ectopic pregnancy was initially managed by intramuscular methotrexate. However, after 10 days, despite an hCG value drop to 409 international units per mL, an indication that methotrexate was working, the patient had a subsequent rupture with hemoperitoneum necessitating exploratory laparotomy and abdominal hysterectomy.

Cardiac Adverse Events With Remdesivir in COVID-19 Infection.

Since December 2019, coronavirus has gradually progressed to a pandemic with no efficacious treatment. Remdesivir is an antiviral medication and inhibitor of viral RNA dependent RNA polymerase with inhibitory action against SARS-CoV virus. Two patients diagnosed with coronavirus infection with worsening respiratory status were initiated with multimodality therapy with antibiotics, steroids and remdesivir. After initiation of remdesivir, the patients' developed bradycardia, with one of the two also showing signs of worsening QT interval. This reverted upon stopping remdesvir therapy. The prevalence of bradycardia with prolonged QT interval is not well-known yet with this medication.

Rectus Sheath Hematoma Causing Bladder Outlet Obstruction.

A 93-year-old woman on Coumadin with history of atrial fibrillation and chronic obstructive pulmonary disease (COPD) presented with urinary retention for one day. Computed tomography (CT) of abdomen and pelvis demonstrated grade 3 rectus sheath hematoma (RSH), with the hematoma dissecting between the transversalis fascia and muscle into the prevesical space. The large-sized hematoma caused compression at the bladder outflow tract causing urinary retention. In view of age and the patient being a poor surgical candidate, the patient was managed by percutaneous drain of the hematoma to reduce size to relieve urinary symptoms. The hematoma shrunk in size over the period of next few weeks and thereby avoided surgical intervention.

Biased Agonism/Antagonism of Cardiovascular GPCRs for Heart Failure Therapy.

G protein-coupled receptors (GPCRs) are among the most important drug targets currently used in clinic, including drugs for cardiovascular indications. We now know that, in addition to activating heterotrimeric G protein-dependent signaling pathways, GPCRs can also activate G protein-independent signaling, mainly via the ßarrestins. The major role of ßarrestin1 and -2, also known as arrestin2 or -3, respectively, is to desensitize GPCRs, i.e., uncoupled them from G proteins, and to subsequently internalize the receptor. As the ßarrestin-bound GPCR recycles inside the cell, it serves as a signalosome transducing signals in the cytoplasm. Since both G proteins and ßarrestins can transduce signals from the same receptor independently of each other, any given GPCR agonist might selectively activate either pathway, which would make it a biased agonist for that receptor. Although this selectivity is always relative (never absolute), in cases where the G protein- and ßarrestin-dependent signals emanating from the same GPCR result in different cellular effects, pharmacological exploitation of GPCR-biased agonism might have therapeutic potential. In this chapter, we summarize the GPCR signaling pathways and their biased agonism/antagonism examples discovered so far that can be exploited for heart failure treatment. We also highlight important issues that need to be clarified along the journey of these ligands from bench to the clinic.