G protein-coupled receptor kinase 2 (GRK2) with its multidomain structure performs various crucial cellular functions under both normal and pathological conditions. Overexpression of GRK2 is linked to cardiovascular diseases, and its inhibition or deletion has been shown to be protective. The functions of GRK2 extend beyond G protein-coupled receptor (GPCR) signaling, influencing non-GPCR substrates as well. Increased GRK2 in heart failure (HF) initially may be protective but ultimately leads to maladaptive effects such as GPCR desensitization, insulin resistance, and apoptosis. The multifunctional nature of GRK2, including its action in hypertrophic gene expression, insulin signaling, and cardiac fibrosis, highlights its complex role in HF pathogenesis. Additionally, GRK2 is involved in mitochondrial biogenesis and lipid metabolism. GRK2 also regulates epinephrine secretion from the adrenal gland and its increase in circulating lymphocytes can be used to monitor HF status. Overall, GRK2 is a multifaceted protein with significant implications for HF and the regulation of GRK2 is crucial for understanding and treating cardiovascular diseases.

AbstractSmoking, particularly chronic smoking (CS), is a threat to global health, contributing to increased mortality and morbidity associated with cardiovascular disease (CVD). CS induces oxidative stress and endothelial dysfunction, which has a profound impact on cardiac structure and function. While the protective effects of estrogen, particularly 17β-estradiol (E2), on cardiovascular health are well-documented in premenopausal women, the interaction between estrogen and CS remains poorly understood. The aim of this study is to investigate the impact of chronic cigarette smoking on cardiac health in relation to ethinylestradiol (EE) oral contraceptive (OC) usage in premenopausal females. Female mice were exposed to chronic cigarette smoke and co-administered EE. Cardiac structural and functional parameters were assessed alongside inflammatory markers, oxidative stress indicators, and histological changes. Results revealed that the combination of EE and CS led to adverse cardiac remodeling characterized by increased left ventricular end-diastolic volume and elevated left ventricular mass. In addition, an inflammatory state was evident, marked by increased expression of IL-4, IL-1β, IL-13, IL-10, and PARP-1, as well as increased interstitial collagen deposition. These findings suggest a progression towards adverse cardiac remodeling resembling dilated cardiomyopathy. Furthermore, our observations highlight the complexity of the inflammatory response triggered by smoking, potentially exacerbated by estrogen supplementation. The main finding of this study is that the combination of CS and EE enhanced adverse cardiac remodeling, which was shown structurally, histologically, and biochemically.

Acute kidney injury (AKI) is a common complication of cardiovascular diseases (CVDs) in both males and females, increasing mortality rate substantially. Premenopausal females appear to be more protected, suggesting a potential protective role of female sex hormones. Here, we tested the hypothesis that ovariectomy (OVX) eliminates the beneficial effect of female sex on renal protection following acute myocardial infarction (MI). Seven days post-MI, both sexes exhibited worsened kidney function and a substantial decrease in total kidney NAD levels. UnlikeMI femalemice, MI males showed exacerbated morphological alterations with increased proinflammatory, proapoptotic, and profibrotic biomarkers. The expression of NAD(+) biosynthetic enzymes NAMPT and NMRK-1 was increased in MI females only, while males showed a substantial increase in NAD(+) consuming enzyme PARP-1. OVX did not eliminate the female-sex protection of glomerular morphology but was associated with swelling of proximal convoluted tubules with MI as in males. With OVX, MI females had enhanced proinflammatory cytokine release, and a further decrease in creatinine clearance and urine output was observed. Our findings suggest that MI induced AKI in both sexes with pre-menopausal female mice being more protected. Ovariectomy worsens aspects of AKI in females after MI, which may portend increased risk for development of chronic kidney disease.

ABSTRACT:Inflammation is a major player in many cardiovascular diseases including hypertension, atherosclerosis, myocardial infarction, and heart failure. In many individuals, these conditions coexist and mutually exacerbate each other's progression. The pathophysiology of these diseases entails the active involvement of both innate and adaptive immune cells. Immune cells that possess the α7 subunit of the nicotinic acetylcholine receptor on their surface have the potential to be targeted through both pharmacological and electrical stimulation of the cholinergic system. The cholinergic system regulates the inflammatory response to various stressors in different organ systems by systematically suppressing spleen-derived monocytes and chemokines and locally improving immune cell function. Research on the cardiovascular system has demonstrated the potential for atheroma plaque stabilization and regression as favorable outcomes. Smaller infarct size and reduced fibrosis have been associated with improved cardiac function and a decrease in adverse cardiac remodeling. Furthermore, enhanced electrical stability of the myocardium can lead to a reduction in the incidence of ventricular tachyarrhythmia. In addition, improving mitochondrial dysfunction and decreasing oxidative stress can result in less myocardial tissue damage caused by reperfusion injury. Restoring baroreflex activity and reduction in renal damage can promote blood pressure regulation and help counteract hypertension. Thus, the present review highlights the potential of nicotinic acetylcholine receptor activation as a natural approach to alleviate the adverse consequences of inflammation in the cardiovascular system.

Ranolazine was approved by the US Food and Drug Administration as an antianginal drug in 2006, and has been used since in certain groups of patients with stable angina. The therapeutic action of ranolazine was initially attributed to inhibitory effects on fatty acids metabolism. As investigations went on, however, it developed that the main beneficial effects of ranolazine arise from its action on the late sodium current in the heart. Since late sodium currents were discovered to be involved in various heart pathologies such as ischemia, arrhythmias, systolic and diastolic dysfunctions, and all these conditions are associated with heart failure, ranolazine has in some way been tested either directly or indirectly on heart failure in numerous experimental and clinical studies. As the heart continuously remodels following any sort of severe injury, the inhibition by ranolazine of the underlying mechanisms of cardiac remodeling including ion disturbances, oxidative stress, inflammation, apoptosis, fibrosis, metabolic dysregulation, and neurohormonal impairment are discussed, along with unresolved issues. A projection of pathologies targeted by ranolazine from cellular level to clinical is provided in this review.

Background Whether cigarette smoking affects the heart post-myocardial infarction (MI) in a sex-dependent way remains controversial. Using a mouse model, we investigated cardiac remodeling under the influence of acute cigarette smoke (CS) exposure following ischemic injury in both sexes. Methods Ten cigarettes were smoked twice daily for 2 weeks followed by MI and then 1 additional week post permanent LAD ligation. Cardiac function, histology, and infarct size were assessed, and inflammatory markers quantified by RT–PCR. Statistical comparisons were performed using an unpaired t test or ANOVA followed by Tukey post hoc test. Results We observed that cigarette smoking exacerbated both left and right ventricular remodeling only in males at an early stage of post-MI. Females did not display a significant structural and/or functional alteration within 7 days of cardiac remodeling post-MI upon CS exposure. Worsened right ventricular remodeling in males was independent of pulmonary congestion. CS-exposed males exhibited enhanced increases in left ventricular end systolic and diastolic volumes, as well as reductions in ejection fraction and fractional area changes of left ventricular base. At day 7, infarct size was increased by cigarette smoking in males only, which was accompanied by enhanced collagen deposition in both the infarcted and peri-infarcted areas. Both IL-6 and TNF-α mRNA expression significantly increased in CS-exposed MI male group only at day 7 post-MI suggestive of prolonged inflammation. Conclusions These findings indicate that CS exposure worsens the progression of cardiac remodeling post-MI in male sex in a significant manner compared to female sex at least at early stages.

Heart failure with preserved ejection fraction (HFpEF) remains a medical anomaly that baffles researchers and physicians alike. The overall phenotypical changes of diastolic function and left ventricular hypertrophy observed in HFpEF are definable; however, the metabolic and molecular alterations that ultimately produce these changes are not well established. Comorbidities such as obesity, hypertension, and diabetes, as well as general aging, play crucial roles in its development and progression. Various animal models have recently been developed to better understand the pathophysiological and metabolic developments in HFpEF and to illuminate novel avenues for pharmacotherapy. These models include multi-hit rodents and feline aortic constriction animals. Recently, genomic, proteomic, and metabolomic approaches have been used to define altered signaling pathways in the heart associated with HFpEF, including those involved in inflammation, cGMP-related, Ca2+ handling, mitochondrial respiration, and the unfolded protein response in endoplasmic reticulum stress. This article aims to present an overview of what has been learnt by these studies, focusing mainly on the findings in common while highlighting unresolved issues. The knowledge gained from these research models will not simply be of benefit for treating HFpEF but will undoubtedly provide new insights into the mechanisms by which the heart deals with external stresses and how the processes involved can fail.

A 64 -year-old female was admitted to our inpatient clinic with a 4-month-history of dyspnea and chest pain both at rest and on exertion. One year ago, she had coronary artery by-pass surgery due to triple vessel disease. Electrocardiogram displayed ST segment elevation in leads V2, D1 and aVL (A). A series of cardiac enzyme tests showed no signs of acute myocardial damage. Chest x-ray revealed mild cardiomegaly with a mass image adjacent to the left ventricle (B, arrow). Echocardiogram demonstrated mildly reduced left ventricular function and an aneurysm cavity with a wide neck in the anterobasal region of the left ventricle, partly filled with thrombus (C and D, arrows). Coronary angiography revealed triple vessel disease, total occluded mid LAD, patent LIMA-LAD and safenCx. On a cardiac magnetic resonance imaging (MRI); a left sided two-chamber cine image clearly showed a thin-walled saccular anterobasal aneurysm partly filled with thrombus (E, arrow), and an inversion-recovery-prepared breath-hold gradient echo cine MRI fifteen minutes after material injection showed delayed enhancement of the wall of the aneurysm and thrombus (F, arrow). There were no coronary vessels surrounding the aneurysm.

Introduction: Tobacco smoke is a major risk factor for coronary artery diseases (CAD) and chronic tobacco smoking (CS) increase the risk of CAD by 2 to 4 folds. CS effect on cardiac homeostasis has never been evaluated in premenopausal females taking oral contraceptive (OC). This study investigates CS effect on cardiac remodeling in female mice in the presence or absence of Ethinyl Estradiol (EE). Methods: Blood pressure and echocardiography were recorded for female C57BL/6J mice on EE, EE-CS, vehicle, and vehicle-CS at baseline and after 8-weeks of exposure. Cardiac inflammatory cytokines and oxidative stress markers were evaluated by real time PCR and western blots. Interstitial collagen deposition was assessed by Masson Trichrome staining. Results: Eight weeks of EE-CS treated mice showed no effect on BP but significant adverse structural and functional cardiac effects, represented by increased systolic (0.981 ± 0.035 N=11 (p<0.0332)) and diastolic (1.450 ± 0.037 N=11 (p<0.0002)) areas when compared to the vehicle and EE groups, along with increased systolic (1.981 ± 0.109 N=11 (p<0.033)) and diastolic ( 3.911 ± 0.159 N=11 (p<0.033)) volumes when compared to all groups. Functional changes were accompanied with a decreased fractional shortening (9.194 ± 1.006 N=11 (p<0.033)) when compared to vehicle and EE groups with no changes in ejection fraction parameter. Moreover EE-CS treatment significantly increased NOX-4 expression (2.297 ± 0.643 N=5 (p<0.033)) when compared to EE treatment alone, along with an increased pro-inflammatory profile including IL-1β (0.750 ±0.171N=5 (p<0.033)) and IL-4 (4.110 ± 0.623 N= 5 (p<0.002)) when compared to all groups, and IL-6 (3.045±0.910 N=5 (p<0.002)) and IL-13 (2.686 ± 0.648 N=5 (p<0.033)) when compared to vehicle-CS group. Morphologically, EE-CS exhibited a significant increase in interstitial fibrosis (1.186± 0.020 N=5, 3.2285 ± 0.683 N=5, (p<0.033)) when compared to the vehicle-CS group. Conclusion: This study provides clear evidence that CS-exposed premenopausal female mice on OC regimen exhibited significant adverse cardiac events that could be classified under cardiac dysfunction with preserved ejection fraction. Additional experiments are warranted to further elaborate on these findings.

Introduction: Acute kidney injury (AKI) is a common complication of acute myocardial infarction (AMI) in both males and females, increasing mortality rate substantially. Premenopausal females appear to be more protected, suggesting a potential protective role of estrogen. Hypothesis: We hypothesized that premenopausal female mice are more protected against AMI-induced AKI and that is potentially due to female sex hormones. Materials and Methods: AMI was induced by ligating the left anterior coronary artery in male and premenopausal female mice. Kidney histological, molecular, and functional parameters were assessed after 7 days. Surgical menopause was induced in AMI-female mice to evaluate estrogen-mediation of the observed renal protection. Results: 7days post-AMI enhanced fibrosis and glomerular retraction were observed in AMI-males, only. Both sexes exhibited a comparable marked increase in glomerular reactive oxygen species (ROS) and DNA fragmentation that was associated with a marked decrease in total NAD kidney levels. Increased release of IL-1β, a pro-inflammatory cytokine, and caspase-3, a proapoptotic marker, and α-smooth muscle actin, a profibrotic biomarker, was seen in AMI-male mice, only. NAD + biosynthetic enzymes, including nicotinamide phosphoribosyl transferase and nicotinamide riboside kinase-1, increased only in AMI-females, whereas male counterparts showed a substantial decrease in mitochondrial bioenergetic enzymes such as Sirtuins 1 and 3, along with an increase in Poly [ADP-ribose] polymerase 1, an NAD + consuming enzyme. Exacerbated kidney damage in AMI-females post-ovariectomy was seen with aggravated morphological alterations, increased ROS generation, and enhanced pro-inflammatory cytokine release, and altered NAD + biosynthetic and mitochondrial bioenergetic enzymes. These changes translated into exacerbated kidney function as evidenced by a decrease in creatinine clearance in AMI-male and female mice, while urine output declined in AMI-male mice only. A further decrease in creatinine clearance and urine output was observed in AMI-female mice following ovariectomy. Conclusion: Our findings suggest that AMI induced AKI in both sexes with pre-menopausal female mice being more protected.

Myocardial infarction (MI) is the leading cause of mortality worldwide. Interleukin (IL)-33 (IL-33) is a cytokine present in most cardiac cells and is secreted on necrosis where it acts as a functional ligand for the ST2 receptor. Although IL-33/ST2 axis is protective against various forms of cardiovascular diseases, some studies suggest potential detrimental roles for IL-33 signaling. The aim of the present study was to examine the effect of IL-33 administration on cardiac function post-MI in mice. MI was induced by coronary artery ligation. Mice were treated with IL-33 (1 μg/day) or vehicle for 4 and 7 days. Functional and molecular changes of the left ventricle (LV) were assessed. Single cell suspensions were obtained from bone marrow, heart, spleen, and peripheral blood to assess the immune cells using flow cytometry at 1, 3, and 7 days post-MI in IL-33 or vehicle-treated animals. The results of the present study suggest that IL-33 is effective in activating a type 2 cytokine milieu in the damaged heart, consistent with reduced early inflammatory and pro-fibrotic response. However, IL-33 administration was associated with worsened cardiac function and adverse cardiac remodeling in the MI mouse model. IL-33 administration increased infarct size, LV hypertrophy, cardiomyocyte death, and overall mortality rate due to cardiac rupture. Moreover, IL-33-treated MI mice displayed a significant myocardial eosinophil infiltration at 7 days post-MI when compared with vehicle-treated MI mice. The present study reveals that although IL-33 administration is associated with a reparative phenotype following MI, it worsens cardiac remodeling and promotes heart failure.

Multiple clinical studies documented renal damage in chronic cigarette smokers (CS) irrespective of their age and gender. Premenopausal female smokers are known to exert a certain cardiovascular and renal protection with undefined mechanisms. Given the multiple demographic variables within clinical studies, this experimental study was designed to be the first to assess whether gender‐biased CS‐induced kidney damage truly exists between premenopausal female and age‐matched C57Bl6J male mice when compared to their relative control groups. Following 6 weeks of CS exposure, cardiac function, inflammatory marker production, fibrosis formation, total and glomerular ROS levels, and glomerulotubular homeostasis were assessed in both genders. Although both CS‐exposed male and female mice exhibited comparable ROS fold change relative to their respective control groups, CS‐exposed male mice showed a more pronounced fibrotic deposition, inflammation, and glomerulotubular damage profile. However, the protection observed in CS‐exposed female group was not absolute. CS‐exposed female mice exhibited a significant increase in fibrosis, ROS production, and glomerulotubular alteration but with a pronounced anti‐inflammatory profile when compared to their relative control groups. Although both CS‐exposed genders presented with altered glomerulotubular homeostasis, the alteration phenotype between genders was different. CS‐exposed males showed a significant decrease in Bowman's space along with reduced tubular diameter consistent with an endocrinization pattern of chronic tubular atrophy, suggestive of an advanced stage of glomerulotubular damage. CS‐exposed female group, on the other hand, displayed glomerular hypertrophy with a mild tubular dilatation profile suggestive of an early stage of glomerulotubular damage that generally precedes collapse. In conclusion, both genders are prone to CS‐induced kidney damage with pronounced female protection due to a milder damage slope.

The impact of cigarette smoking (CS) on kidney homeostasis in the presence of myocardial infarction (MI) in both males and females remains poorly elucidated. C57BL6/J mice were exposed to 2 weeks of CS prior to MI induction followed by 1 week of CS exposure in order to investigate the impact of CS on kidney damage in the presence of MI. Cardiac hemodynamic analysis revealed a significant decrease in ejection fraction (EF) in CS-exposed MI male mice when compared with the relative female subjects, whereas cardiac output (CO) comparably decreased in CS-exposed MI mice of both sexes. Kidney structural alterations, including glomerular retraction, proximal convoluted tubule (PCT) cross-sectional area, and total renal fibrosis were more pronounced in CS-exposed MI male mice when compared with the relative female group. Although renal reactive oxygen species (ROS) generation and glomerular DNA fragmentation significantly increased to the same extent in CS-exposed MI mice of both sexes, alpha-smooth muscle actin (α-SMA) and connective tissue growth factor (CTGF) significantly increased in CS-exposed MI male mice, only. Metabolically, nicotinamide phosphoribosyltransferase (NAMPT) and nicotinamide riboside-1 (NMRK-1) substantially increased in CS-exposed MI female mice only, whereas sirtuin (SIRT)-1 and SIRT-3 substantially decreased in CS-exposed MI male mice compared with their relative female group. Additionally, renal NAD levels significantly decreased only in CS-exposed MI male mice. In conclusion, MI female mice exhibited pronounced renal protection following CS when compared with the relative male groups.

Cardiac autonomic neuropathy (CAN) is an early cardiovascular manifestation of type 2 diabetes (T2D) that constitutes an independent risk factor for cardiovascular mortality and morbidity. Nevertheless, its underlying pathophysiology remains poorly understood. We recently showed that localized perivascular adipose tissue (PVAT) inflammation underlies the incidence of parasympathetic CAN in prediabetes. Here, we extend our investigation to provide a mechanistic framework for the evolution of autonomic impairment as the metabolic insult worsens. Early metabolic dysfunction was induced in rats fed a mild hypercaloric diet. Two low-dose streptozotocin injections were used to evoke a state of late decompensated T2D. Cardiac autonomic function was assessed by invasive measurement of baroreflex sensitivity using the vasoactive method. Progression into T2D was associated with aggravation of CAN to include both sympathetic and parasympathetic arms. Unlike prediabetic rats, T2D rats showed markers of brainstem neuronal injury and inflammation as well as increased serum levels of IL-1β. Experiments on PC12 cells differentiated into sympathetic-like neurons demonstrated that brainstem injury observed in T2D rats resulted from exposure to possible proinflammatory mediators in rat serum rather than a direct effect of the altered metabolic profile. CAN and the associated cardiovascular damage in T2D only responded to combined treatment with insulin to manage hyperglycemia in addition to a nonhypoglycemic dose of metformin or pioglitazone providing an anti-inflammatory effect, coincident with the effect of these combinations on serum IL-1β. Our present results indicate that CAN worsening upon progression to T2D involves brainstem inflammatory changes likely triggered by systemic inflammation.

Despite increased social awareness, marketing restraints, tobacco taxation, and available smoking cessation rehab programs, active and passive smoking remain a worldwide challenging epidemic and a key risk factor for cardiovascular diseases development. Although cardiovascular (CV) protection is more pronounced in women than in men due to estrogenic effects, tobacco cigarette smoking exposure seems to alter this protection by modulating estrogen actions via undefined mechanisms. Premenopausal cigarette smoking women are at higher risk of adverse CV effects than non-smokers. In this study, we investigated the impact of cigarette smoking on early CV injury after myocardial infarction (MI) in non-menopausal female mice. Aortic arch calcification, fibrosis, reactive oxygen species, and gene expression of inflammatory and calcification genes were exaggerated in mice exposed to cigarette smoke (CS). These findings suggest that aortic injury following MI, characterized by vascular smooth muscle cells transdifferentiation, calcification, inflammation, and collagen deposition but not cardiac dysfunction is exacerbated with CS exposure. The novel findings of this study highlight the importance of aortic injury on short and long-term prognosis in CS-exposed MI females. Linking those findings to estrogen alteration is probable and entails investigation.

HomeHypertensionVol. 72, No. 4IL-33 (Interleukin 33)/sST2 Axis in Hypertension and Heart Failure Free AccessReview ArticlePDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissions ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toFree AccessReview ArticlePDF/EPUBIL-33 (Interleukin 33)/sST2 Axis in Hypertension and Heart Failure Rana Ghali, Raffaele Altara, William E. Louch, Alessandro Cataliotti, Ziad Mallat, Abdullah Kaplan, Fouad A. Zouein and George W. Booz Rana GhaliRana Ghali From the Department of Pharmacology and Toxicology, American University of Beirut Medicine Center, Lebanon (R.G., A.K., F.A.Z.) *R. Ghali and R. Altara contributed equally to this work as first authors. Search for more papers by this author , Raffaele AltaraRaffaele Altara Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, Norway (R.A., W.E.L., A.C.) KG Jebsen Center for Cardiac Research, Oslo, Norway (R.A., W.E.L., A.C.) Department of Pathology (R.A.), School of Medicine, University of Mississippi Medical Center, Jackson *R. Ghali and R. Altara contributed equally to this work as first authors. Search for more papers by this author , William E. LouchWilliam E. Louch Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, Norway (R.A., W.E.L., A.C.) KG Jebsen Center for Cardiac Research, Oslo, Norway (R.A., W.E.L., A.C.) Search for more papers by this author , Alessandro CataliottiAlessandro Cataliotti Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, Norway (R.A., W.E.L., A.C.) KG Jebsen Center for Cardiac Research, Oslo, Norway (R.A., W.E.L., A.C.) Search for more papers by this author , Ziad MallatZiad Mallat Division of Cardiovascular Medicine, Department of Medicine, University of Cambridge, United Kingdom (Z.M.) Institut National de la Sante et de la Recherche Medicale (Inserm), Unit 970, Paris Cardiovascular Research Center, France (Z.M.). Search for more papers by this author , Abdullah KaplanAbdullah Kaplan From the Department of Pharmacology and Toxicology, American University of Beirut Medicine Center, Lebanon (R.G., A.K., F.A.Z.) Search for more papers by this author , Fouad A. ZoueinFouad A. Zouein Correspondence to Fouad A. Zouein, Department of Pharmacology and Toxicology, American University of Beirut Medical Center, Riad El-Solh, Beirut 1107 2020, Beirut-Lebanon. E-mail E-mail Address: [email protected] From the Department of Pharmacology and Toxicology, American University of Beirut Medicine Center, Lebanon (R.G., A.K., F.A.Z.) Search for more papers by this author and George W. BoozGeorge W. Booz Department of Pharmacology and Toxicology (G.W.B.), School of Medicine, University of Mississippi Medical Center, Jackson Search for more papers by this author Originally published27 Aug 2018https://doi.org/10.1161/HYPERTENSIONAHA.118.11157Hypertension. 2018;72:818–828is corrected byCorrection to: IL-33 (Interleukin 33)/sST2 Axis in Hypertension and Heart FailureOther version(s) of this articleYou are viewing the most recent version of this article. Previous versions: September 12, 2018: Previous Version of Record August 27, 2018: Ahead of Print IL-33 (interleukin 33) is type of cytokine designated an alarmin, which is released under stress conditions or by cell death to induce protective measures in neighboring cells.1 It signals via the ST2 receptor, which is also expressed as a truncated and soluble form that acts as a decoy receptor soluble ST2 (sST2).2 Substantial evidence supports the use of sST2 measurement as an indicator of cardiac stress and remodeling in hypertension, heart failure, cardiomyopathies, and aortic stenosis (AS). However, as discussed here, there are gaps in our understanding of the IL-33/sST2 axis, including the source of the components, stimuli for their expression and release, what it actually assesses, and the role of immune cells. Moreover, as discussed, the fundamental assumption that Il-33 has beneficial actions that are thwarted by sST2 requires further scrutiny.Background, Signaling, and ExpressionIL-33 is constitutively expressed and mainly present in nuclei of epithelial and endothelial cells.3,4 IL-33 was first discovered in 1999 as a nuclear protein of unknown function encoded by a gene upregulated in canine vasospastic cerebral vasculature after subarachnoid hemorrhage and termed DVS27.5 In 2003, the same nuclear protein was found to be abundantly expressed in high endothelial venules that allow lymphocytes access into lymphoid organs and lymph nodes6,7 and accordingly named nuclear factor from high endothelial venules.8 Schmitz et al9 determined 2 years later that DVS27 and nuclear factor from high endothelial venules were the same protein and noticed that its C-terminal domain contains a β-sheet trefoil fold structure similar to IL-1 family members. Indeed, the protein was subsequently observed to function as a cytokine that induces T helper (Th) type 2 responses through ST2, an orphan receptor of the IL-1 receptor superfamily. This novel IL-1 family member was then named IL-33.9The ST2 receptor exists as 2 isoforms, sST2 and transmembrane (ST2L), encoded by the same gene (IL1RL1) under a proximal and distal promoter, respectively. Oxidation and extracellular proteases terminate IL-33 biological actions.10Because of its expression at barrier tissues, IL-33 plays a key role in maintaining barrier tissue defense.11 As an alarmin, IL-33 activates various cell types of the innate and adaptive immune system. Certain proteases, such as neutrophil and mast cell serine proteases, cathepsin G, chymase, and elastase, cleave IL-33 extracellularly and enhance its activity.12–14 Resident immune cells constitutively expressing ST2L are a main target for IL-33 and include group 2 innate lymphoid cells (ILC2s),15,16 mast cells,17–19 and tissue-resident regulatory T cells.20,21 Variations in ST2L expression affect the type of immune response that IL-33 produces. For instance, while expression of ST2L was decreased in ILC2 in a murine model of chronic obstructive pulmonary disease, it was induced in macrophages (with M2 polarization) and natural killer (NK) cells.22 Other targets of IL-33 include Th2 cells, B cells, basophils, eosinophils, dendritic cells, and natural killer T (NKT) cells.1 Th1 and CD8+ T cells also show induced expression of ST2L in certain cases.23,24 Depending on context, IL-33 is therefore able to activate and perpetuate inflammation (eg, via Th2 or mast cells)25 or reduce and resolve inflammatory responses (eg, via tissue-resident regulatory T cells or ILC2).26,27 By activating Th2 cells, IL-33 elicits a type 2 immune response, which if exuberant may lead to tissue damage likely through activation of mast cells or eosinophils and development of pathological fibrosis.28 In this way, IL-33 plays a role in the pathophysiology of several proinflammatory and autoimmune diseases.29 In certain cells, IL-33 may induce expression of ST2L (eg, ILC2, eosinophils, and tissue-resident regulatory T cells) or sST2 (eg, mast cells).1Figure 1 summarizes some notable actions of IL-33 on immune cells that are potentially relevant to the heart.Download figureDownload PowerPointFigure 1. Principal actions of IL (interleukin)-33 on certain immune cells that are relevant for heart function and remodeling.1 CCL17 indicates chemokine (C-C motif) ligand 17; ILC2, group 2 innate lymphoid cells; IFN, interferon; LPS, lipopolysaccharides; MCP, monocyte chemoattractant protein; TNF, tumor necrosis factor; and Treg, tissue-resident regulatory T cells.Expression of IL-33 in nonimmune cells occurs normally throughout the human body. Endothelial cells constitute the main cell type that exhibits elevated levels of IL-33 at barrier surfaces.3 IL-33 is present also in adipose tissue30 and atherosclerotic plaques31 and is constitutively present in secondary lymphoid organs.3 Il-33 is constitutively expressed in endothelial cells of the vasculature tree of normal tissues, but not the microcirculation of glomeruli and brain.3 Keratinocytes, epithelial cells, fibroblasts, and smooth muscle cells also express IL-33.3,9,32,33 Under pathological conditions, increased IL-33 levels are detected in various adenocarcinomas,3 and elevated levels are found in the circulation and tissues during several inflammatory diseases including rheumatoid arthritis, inflammatory bowel syndrome, asthma, and skin diseases.34–37 Although IL-33 is primarily expressed by nonhematopoietic cells, activated dendritic cells and M2 macrophages may also express and secrete IL-33.9,38Expression of IL-33 at epithelial surfaces occurs in both humans and mice.3,9,35 Unlike in humans, IL-33 is not normally present along the mouse vascular tree, although expression has been observed in a few vascular beds (ovaries, adipose tissue, and liver).4,39–41 However, IL-33 expression was observed to be increased in endothelial (and interstitial) cells of the mouse heart after transverse aortic constriction.42In the human heart, IL-33 is mainly present in endothelial cells of the vasculature, although cardiomyocytes and fibroblasts of adults also constitutively express the protein. Proinflammatory cytokines like IL-1β, TNF (tumor necrosis factor)-α, and IFN (interferon)-γ increase IL-33 in these cells, as well as in human coronary artery smooth muscle cells.43 Very low levels of sST2 and ST2L mRNA are detected in human cardiomyocytes, vascular smooth muscle cells, and cardiac fibroblasts, whereas ST2 is widely expressed in endothelial cells of the cardiac vasculature, and both cardiac macro- and microvascular endothelial cells secrete sST2.43 Expression of both IL-33 and sST2 is induced in rat neonatal cardiomyocytes and (more so) in cardiac fibroblasts on mechanical stress.33,44Hypertension/Pulmonary HypertensionCirculating sST2 levels may have diagnostic and prognostic value in hypertension.45,46 Increased sST2 serum levels are associated with alterations of left ventricular (LV) geometry46 and increased systolic blood pressure.47,48 Among Framingham Heart Study participants, elevated blood levels of sST2 were associated with hypertension and diabetes mellitus48; however, in healthy individuals from the general population, sST2 had little predictive value for cardiovascular events.49In pulmonary arterial hypertension (PAH), the role of the IL-33/sST2 axis would seem straightforward. Because sST2 release is potentiated during myocardial stretch, it is hypothesized that increased pulmonary arterial blood pressure and right ventricular (RV) afterload increase transmural wall stress, hence increasing myocardial stretch and sST2 release. Therefore, sST2 could be considered a cardiac biomarker of mechanical strain and act as a useful biomarker gauging RV dysfunction in PAH.50 Indeed, sST2 expression reflects the size and function of the RV and independently predicts 1-year mortality in PAH patients.51,52 Carlomagno et al53 found that elevated sST2 expression in patients with PAH was associated with systolic dysfunction and RV dilatation. Elevated sST2 levels were also associated with poor prognosis in childhood PAH with sST2/NT-proBNP (N-terminal pro-B-type natriuretic peptide) biomarker combination predicting clinical condition and outcome.54 Moreover, increased sST2 levels predicted disease severity in idiopathic PAH patients and correlated with hemodynamic parameters and other biomarkers.50 In dyspneic patients, increased sST2 levels directly correlated with RV dysfunction and predicted worsened prognosis.55 In patients with secondary pulmonary hypertension because of chronic obstructive pulmonary disease, increases in both sST2 and NT-proBNP correlated with RV dysfunction and possessed prognostic and diagnostic value.51However, the assumption that RV myocytes release sST2 when stretched is based on work with LV myocytes (see below), which have a different embryological origin. Alternatively, loss of nuclear IL-33 in pulmonary endothelial cells might explain elevated sST2 serum levels in idiopathic PAH patients. Nuclear IL-33 was found to be markedly reduced in endothelial cells of pulmonary arterioles of patients with idiopathic PAH, which has no cure and is associated with RV failure and death.56 Circulating levels of IL-33 were unchanged, although accurate assessment of circulating IL-33 is fraught with challenges.10 Further investigation revealed that nuclear IL-33 in human endothelial cells formed a complex with a transcriptional repressor and bound multiple putative homeodomain protein binding motifs in the proximal and distal promoters of ST2 genes to repress their expression. Nuclear IL-33 was also shown to function as a transcriptional repressor of other inflammatory genes, such as IL-6, in human endothelial cells. In addition, mast cells contribute to different types of PAH,57 and active mast cells were found to accumulate in the RV of a mouse model of PAH in association with increased RV hypertrophy and fibrosis.58 Mast cells constitutively express ST2L, and sST2 is strongly induced by IL-33.1Cardiac HypertrophyWith hypertension, the LV undergoes hypertrophy, a risk factor for cardiovascular mortality.59 Pressure overload of the mouse heart by transverse aortic constriction was found to induce IL-33 expression, and greater LV hypertrophy and adverse cardiac remodeling were observed in ST2−/− mice, lacking both sST2 and ST2L.33 Using cultured neonatal rat cardiomyocytes, IL-33 was demonstrated to antagonize agonist-induced cardiac hypertrophy, an action blocked by sST2 or an antibody to ST2L.33 Moreover, treatment with recombinant IL-33 attenuated hypertrophy and cardiac remodeling in wild-type, but not ST2-knockout mice. Although IL-33 transiently activated NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) in cardiac myocytes, its antihypertrophic actions were partially attributed to attenuation of NF-κB activation by hypertrophic agonists, secondary to suppression of ROS (reactive oxygen species) generation.Systemic ablation of IL-33 exacerbated LV hypertrophy in mice in response to transverse aortic constriction and aggravated cardiac fibrosis and inflammation, including increased myocardial expression of Th1 cytokines, such as TNF-α.60 However, subsequent findings on mice showed that pressure overload increases IL-33 expression in cardiac endothelial cells, and IL-33 released from endothelial cells of the heart (but not cardiac myocytes) was responsible not only for attenuating cardiac hypertrophy and fibrotic gene expression via ST2L but also paradoxically for inducing systemic inflammation.42 Cardiac myocyte-derived sST2 was proposed to act locally to block the antihypertrophic actions of IL-33, whereas circulating levels of sST2 were postulated to arise from endothelial cells of the heart.These preclinical findings support a model in which IL-33 is a mechanically induced protein in the heart that acts as a brake on cardiac hypertrophy and fibrosis. Thus, a correlation may occur between levels of IL-33 (negative) or sST2 (positive) and the degree of hypertension or extent of cardiac hypertrophy and fibrosis, and indeed, in vivo and in vitro evidence indicates that IL-33 has direct antihypertrophic actions on adult and neonatal rodent cardiomyocytes.33,42 Whether the same is true in the adult human heart remains uncertain (Figure 2). Human coronary artery and heart microvascular endothelial cells from cardiomyopathy patients were found to express sST2, as well as IL-33 and ST2L. However, human cardiac myocytes, fibroblasts, and vascular smooth muscle cells were observed to express IL-33, but only minor amounts of ST2L or sST2 mRNA and not to secrete sST2 or be responsive to IL-33.43 Explanted human cardiomyopathic hearts exhibited weak or no ST2L staining by cardiac myocytes or fibroblasts, respectively. In addition, no myocardial sST2 protein gradient was detected in heart failure patients.61 Still, the biomechanical or inflammatory stress associated with myocardial infarction or pressure/volume overload of the heart might induce sST2 release from cardiac myocytes. Mechanical strain, phorbol ester, IL-1β, and endothelin 1 were reported to increase production of sST2 by neonatal cardiac myocytes or fibroblasts.33,44,62Download figureDownload PowerPointFigure 2. Controversies surrounding the IL (interleukin)-33/sST2 axis in the heart. Although multiple studies highlight the use of circulating sST2 levels in cardiac diseases, there are many unresolved issues concerning the sources and stimuli for release of IL-33 and sST2, as well as the cellular targets of IL-33. Whether IL-33 is beneficial or harmful is also not known. In the (human) heart, endothelial cells are likely the major source for IL-33 and sST2 release, with a lesser contribution from fibroblasts and myocytes. Although undoubtedly important, the contribution of immune cells is unexplored. The primary target of IL-33 in the heart would seem to be endothelial cells. A direct effect via ST2L on fibroblasts and myocytes in the adult human heart is less certain. However, in these cells, loss or increase in nuclear IL-33 levels may impact on inflammatory gene expression (inset). Although likely targeted, the role of immune cells in IL-33 actions on the heart awaits investigation. Dendritic cells and possibly other immune cells may internalize IL-33, but the cytoplasmic or nuclear consequences of this are not known. Finally, the contribution of extracardiac sources of sST2 and IL-33, such as adipose tissue, liver, large vessels, and lungs, remains a possibility. CBM indicates chromatin binding motif; H2A and H2B, core histones; IL1AcP, IL-1 receptor accessory protein; ILC2, group 2 innate lymphoid cells; Treg, tissue-resident regulatory T cells; and VSMC, vascular smooth muscle cells.Notably, in human LV hypertrophy because of AS and congestive heart failure of undefined cause, serum sST2 levels were found to correlate with diastolic load. However, in contrast to BNP, no correlation was observed with systolic wall stress, which may explain why sST2 in some cases may offer additional predictive value.61 In heart failure, sST2 levels also correlate with serum levels of norepinephrine, natriuretic peptides, and CRP (C-reactive protein), and release of sST2 from human coronary artery endothelial cells is induced by certain inflammatory cytokines (IL-1β and TNF-α).61 Thus, circulating sST2 levels may function as a measure of hemodynamic and inflammatory stress on the heart as primarily gauged by vascular endothelial cells. The importance of immune cells in contributing to the IL-33/sST2 axis in the stressed heart is not defined. Although adult cardiac myocytes and fibroblasts express IL-33, its role in those cells may be largely intracellular. Extracellular IL-33/sST2, on the other hand, may enable crosstalk between endothelial and various immune cells that indirectly leads to cardiac remodeling. Treatment of human coronary artery endothelial cells with IL-33 was found to induce expression of adhesion molecules, produce the chemokine MCP1 (monocyte chemoattractant protein 1), and promote adhesion of human leukocytes.31Hypertensive patients with LV concentric hypertrophy have higher circulating sST2 levels than patients with normal heart geometry.46 Another study by the same group reported that overall sST2 levels were higher in hypertensive patients with LV hypertrophy and correlated with both systolic and diastolic function, as well as RV remodeling.45,46 However, although increased sST2 was associated with increased likelihood of LV hypertrophy in patients with metabolic syndrome, no association was noted with diastolic dysfunction.63 These disparate conclusions could reflect disease stage, relative contributions of systemic versus cardiac inflammation, as well as extent and type of mechanical stress on the heart.64 Such considerations may help explain why both correlations between circulating sST2 and cardiac hypertrophy and between sST2 and cardiac fibrosis (or diastolic dysfunction) need not be observed concurrently. For instance, in patients with hypertrophic cardiomyopathy, a correlation of sST2 with LV hypertrophy, but not LV fibrosis, was recently reported.65Heart FailureCirculating sST2 levels are elevated in chronic heart failure and strongly associated with heart failure severity and adverse outcomes, such as cardiovascular events, cardiac hospitalization, increased risk of death or cardiac transplantation, and sudden cardiac death.66 In contrast to levels of natriuretic peptides, sST2 levels are reported to be unaffected by body mass index, renal function, or age (although there are some reports of an association with age or body mass index).48,67–72Substantial evidence has shown that sST2 is a powerful prognostic biomarker in chronic heart failure with reduced ejection fraction (HFrEF). Blood sST2 levels have use to improve risk stratification in heart failure when added to established clinical variables or biomarkers, such as NT-proBNP. A review in mid-2014 of clinical trials on mostly HFrEF reported that sST2 levels, where assessed separately, offered incremental value as a predictor variable for primary outcome in 18 studies.66 In 10 of these studies, sST2 offered independent prognostic information, and in 8 provided additive value to clinical variables and biomarkers for risk stratification. A recent meta-analysis assessing 6 of these studies, plus 1 from 2016, showed that sST2 is a predictor of both all-cause and cardiovascular death in heart failure patients.73 The use of sST2 to predict clinical course of heart failure, all-cause and cardiovascular mortality, was observed in patients with diabetes mellitus as well.74 Although shortcomings of these trials, such as limited multivariate analysis and narrow patient selection have been noted,67 sST2 was included in the 2013 American College of Cardiology/American Heart Association Guidelines for management of patients with heart failure by additive risk stratification.Other recent studies have supported the use of serial sST2 assessment75–78 or a multi-biomarker profiling approach68,79 in risk stratification of heart failure patients. In both cases, the lower biological variability for sST2 may be advantageous,70,80 if properly combined with other biomarkers that are not redundant for the same pathophysiological process.81 In the large (n=1449) CORONA study (Controlled Rosuvastatin Multinational Trial in Heart Failure), sST2 was independently associated with worsening heart failure.82 In addition, clinical improvement in New York Heart Association (NYHA) III and IV heart failure was associated with a significant reduction in sST2 levels.69 Thus, sST2 may provide insight into disease progression in heart failure comparable to NT-proBNP levels,68 with added value over natriuretic peptides for accurately assessing disease management for monitoring or adjusting drug therapy.83 sST2 was also demonstrated to have value for predicting clinical improvement in systolic heart failure patients. The ST2-R2 score, a composite of clinical variables plus serum concentrations of sST2 and complementary biomarkers, was demonstrated to predict reverse LV remodeling and functional improvement in systolic heart failure patients and mortality up to 4 years.84,85 Interestingly, sST2 was the only studied biomarker independently associated with reverse remodeling. In the PARADIGM-HF trial (Prospective Comparison of ARNI With ACEI to Determine Impact on Global Mortality and Morbidity in Heart Failure), baseline sST2 level was an independent predictor of clinical outcomes even after adjusting for other predictors, including high-sensitivity troponin T (hs-TnT) and NT-proBNP. The association between baseline sST2 and outcomes was linear.86 Notably, increases in sST2 at 1 month were associated with subsequent worse outcomes and decreased with better outcomes. Compared with enalapril, sacubitril/valsartan led to more reductions and fewer increases in sST2 levels over time.More than half the patients with heart failure experience impaired cardiac relaxation, that is, diastolic dysfunction, in the face of a normal ejection fraction, a condition termed heart failure with preserved ejection fraction (HFpEF).87,88 HFpEF, which is more prevalent among the elderly, is characterized by the presence of several comorbidities, such as obesity, type 2 diabetes mellitus, or hypertension. As there are no effective treatments for HFpEF based on the outcomes of clinical trials, HFpEF represents one of the largest unmet needs in medicine. Systemic vascular inflammation and coronary microvascular endothelial dysfunction are thought to be the basis for HFpEF.89 In 2 studies of patients with stable or compensated HFpEF, sST2 blood levels were elevated compared with controls and roughly comparable to those of patients with HFrEF.90,91 In 1 study, slightly higher sST2 levels for HFpEF (n=112; HFrEF, n=458) combined with a different biomarker profile, improved discrimination between the 2 HF subtypes in multivariable analysis.91 However, no difference in sST2 levels was found between HFpEF (n=50) or HFrEF (n=51) patients and a control group (n=50) of randomly sampled free-living adults after adjusting for sex, age, clinical risk factors, and medications.90 In contrast, multivariate analysis showed that elevated sST2 was independently associated with HFpEF in hypertensive patients (n=107), although levels provided no additional insight into the extent of diastolic dysfunction or heart failure functional class.92 Nonetheless, evidence was reported that sST2 levels may have prognostic use in HFpEF, as well as HFrEF, for overall survival and heart failure hospitalization-free survival based on patient populations of predominantly NYHA class ≥III.91A correlation between sST2 and severity of disease, as assessed by blood NT-proBNP levels, E/E′ (a measure of diastolic dysfunction), left atrial (LA) volume, and galectin 3 (a measure of tissue fibrosis) was reported in HFpEF patients, although in a multivariable model after adjusting for confounders, only female sex, NYHA class, and LA volume were associated with higher sST2.93 Increased LV diastolic stiffness in HFpEF might cause increased LA volume. Among HFpEF patients (n=174) enrolled in the RELAX trial (Phosphdiesterase-5 Inhibition to Improve Clinical Status and Exercise Capacity in Diastolic Heart Failure), sST2 was not associated with LV structure or LV systolic or diastolic function; however, higher sST2 levels were associated with the presence of diabetes mellitus, biomarkers of systemic inflammation and fibrosis, neurohumoral activation, myocardial necrosis, atrial fibrillation, systemic congestion, renal dysfunction, RV pressure overload (because of LV diastolic dysfunction [LVDD] and pulmonary congestion) and dysfunction, and exercise intolerance.94 Interestingly, although type 2 diabetes mellitus was associated with elevated levels of sST2 (n=140 versus 18 patients without diabetes mellitus) and a correlation between glycemic control and sST2 was observed, patients with diabetes mellitus with LVDD (n=50) exhibited even higher sST2 levels.95 The rather poor association of sST2 levels with ventricular function in the context of HFpEF may reflect its lack of cardiac specificity. The presence of multiple comorbidities, which may become more prominent with age, extracardiac sources of sST2, and its elevation in many other medical conditions likely limit its cardiac-related prognostic use in HFpEF.A persuasive case can be made that clinical HFpEF is more reflective of adverse LA remodeling than LV concentric hypertrophy and fibrosis. HFpEF is characterized by greater LA stiffness compared with greater eccentric remodeling seen in HFrEF.96,97 Preclinical and clinical evidence indicates that maladaptive LA remodeling occurs early during HFpEF development.96,98,99 At early stages of HFpEF, LA dysfunction is associated with a higher risk of heart failure hospitalization independent of potential clinical confounders, although not independent of LV strain and filling pressure, suggesting a common link between LA and LV remodeling.96 With more advanced-stage heart failure, LA dysfunction is a predictor of mortality in HFpEF.97,100 Limited evidence implicates fibrosis in LA stiffness,98,101 and increased fibrosis and wall stress could contribute to atrial arrhythmias, which is a common comorbidity of HFpEF.97,101 Impaired LA function may explain the pulmonary congestion, shortness of breath, and exercise intolerance associated with HFpEF and could lead to right heart failure,97,102 a leading cause of death in patients with HFpEF accounting for >50% of deaths.103 Consistent with a primary role of LA stiffness in HFpEF are recent findings reported on 86 HFpEF patients from the prospective, multicenter KaRen substudy (Karolinska-Rennes).104 Plasma sST2 levels were (inversely) associated with LA strain, but not LA size, nor LV geometry or systolic/diastolic function. Moreover, heart failure symptoms and outcome were independently predicted by reduced LA strain and not LV functional parameters. As discussed,104 increased LA fibrosis (and sST2) in these HFpEF patients would seem to be reflective of inflammation, rather than attributable to increased hemodynamic load.Although the prognostic use of sST2 is established in chronic heart failure, HFrEF or HFpEF, its predictive value in asymptomatic patients is uncertain. In a longitudinal observational study of 3915 community-dwelling elderly persons without established heart failure, the predictive value of sST2 for heart failure of any subtype or cardiovascular death was found to be modest compared with the existing risk models, despite strong cross-sectional a

Cardiac autonomic neuropathy (CAN) is an early cardiovascular complication of diabetes occurring before metabolic derangement is evident. The cause of CAN remains elusive and cannot be directly linked to hyperglycemia. Recent clinical data report cardioprotective effects of some antidiabetic drugs independent of their hypoglycemic action. Here, we used a rat model receiving limited daily increase in calories from fat (HC diet) to assess whether mild metabolic challenge led to CAN in absence of interfering effects of hyperglycemia, glucose intolerance, or obesity. Rats receiving HC diet for 12 weeks showed reduction in baroreceptor sensitivity and heart rate variability despite lack of change in baseline hemodynamic and cardiovascular structural parameters. Impairment of cardiac autonomic control was accompanied with perivascular adipose inflammation observed as an increased inflammatory cytokine expression, together with increased cardiac oxidative stress, and signaling derangement characteristic of diabetic cardiomyopathy. Two-week treatment with metformin or pioglitazone rectified the autonomic derangement and corrected the molecular changes. Switching rats to normal chow but not to isocaloric amounts of HC for two weeks reversed CAN. As such, we conclude that adipose inflammation due to increased fat intake might underlie development of CAN and, hence, the beneficial effects of metformin and pioglitazone.