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Visual Abstract, Highlights • AAV9 vectors can upregulate Rxfp1 mRNA in murine heart after intravenous injection. • RXFP1 upregulation sensitizes the left ventricle to relaxin-induced inotropy. • RXFP1 overexpression protects heart from ischemia-reperfusion injury., Summary Relaxin is a pleiotropic hormone shown to confer cardioprotection in several preclinical models of cardiac ischemia-reperfusion injury. In the present study, the effects of up-regulating relaxin family peptide receptor 1 (RXFP1) via adeno-associated virus serotype 9 (AAV9) vectors were investigated in a mouse model of myocardial infarction. AAV9-RXFP1 vectors were generated and injected in adult male CD1 mice. Up-regulation of Rxfp1 was confirmed via quantitative polymerase chain reaction, and overexpressing animals showed increased sensitivity to relaxin-induced ventricular inotropic response. Overexpressing animals also demonstrated reduced infarct size and preserved cardiac function 24 hours after ischemia-reperfusion. Up-regulation of RXFP1 via AAV9 vectors has potential therapeutic utility in preventing adverse remodeling after myocardial infarction.

Despite significant advances and the continuous development of novel, effective therapies to treat a variety of malignancies, cancer therapy-induced cardiotoxicity has been identified as a prominent cause of morbidity and mortality, closely competing with secondary malignancies. This unfortunate limitation has prompted the inception of the field of cardio-oncology with its purpose to provide the necessary knowledge and key information on mechanisms that support the use of the most efficacious cancer therapy with minimal or no interruption while paying close attention to preventing cardiovascular related morbidity and mortality. Several mechanisms that contribute to cancer therapy-induced cardiotoxicity have been proposed and studied. These mainly involve mitochondrial dysfunction and reactive oxygen species-induced oxidative stress, lysosomal damage, impaired autophagy, cell senescence, DNA damage, and sterile inflammation with the formation and activation of the NLRP3 inflammasome. In this review, we focus on describing the principal mechanisms for different classes of cancer therapies that lead to cardiotoxicity involving the NLRP3 inflammasome. We also summarize current evidence of cardio-protection with inflammasome inhibitors in the context of heart disease in general, and further highlight the potential application of this evidence for clinical translation in at risk patients for the purpose of preventing cancer therapy associated cardiovascular morbidity and mortality.

Visual Abstract, Highlights • Acute pericarditis is characterized by an intense inflammatory response involving the pericardium. Although mostly benign in its clinical course, 30% of patients may experience complications (recurrence, treatment failure, cardiac tamponade). • The pathogenesis of pericarditis is poorly understood. The scarcity of animal models might justify the limited understanding of this syndrome and the lack of targeted therapies. • Acute pericarditis is believed to represent a stereotypical response to an acute injury of the pericardium. The NLRP3 inflammasome, through its main product, IL-1β, could play a central role in the clinical manifestations. • A mouse model of acute pericarditis was developed through the intrapericardial injection of zymosan A, leading to the classical features of the inflamed pericardium: pericardial effusion, pericardial thickening, and increased expression of the NLRP3 inflammasome. By inhibiting the NLRP3 inflammasome or IL-1β, the pericardial effusion and thickening and the NLRP3 inflammasome expression were greatly reduced compared with vehicle. • Treatment with IL-1 trap, neutralizing both IL-1β and IL-1α, produced a powerful effect on pericardial inflammation in the experimental pericarditis model., Summary Human samples of patients with chronic pericarditis and appropriate control subjects were stained for the inflammasome components. A mouse model of pericarditis was developed through the intrapericardial injection of zymosan A. Different inflammasome blockers were tested in the mouse model. Patients with pericarditis presented an intensification of the inflammasome activation compared with control subjects. The experimental model showed the pathological features of pericarditis. Among inflammasome blockers, NLRP3 inflammasome inhibitor, anakinra, and interleukin-1 trap were found to significantly improve pericardial alterations. Colchicine partially improved the pericardial inflammation. An intense activation of the inflammasome in pericarditis was demonstrated both in humans and in mice.

Background Acute pericarditis is the most common presentation of pericardial diseases. Although generally benign, complications such as constrictive pericarditis, cardiac tamponade, and recurrence can occur. Research Question What are the clinical factors associated with adverse outcomes in acute pericarditis? Study Design and Methods We used an informatics-based search engine to search for International Classification of Diseases codes related to pericardial disease between January 1, 2009 and November 14, 2018 and then extracted clinical information, including only patients meeting the European Society of Cardiology criteria for acute pericarditis. We then evaluated the predictive value of clinical characteristics for adverse outcomes (cardiac tamponade, constrictive pericarditis, failure of therapy, recurrences, or death). Results We identified 240 patients with a first episode of pericarditis (51 [34-62] years, 56% males and 50% white). Pericarditis was determined to be idiopathic in 126 (53%) cases and related to cardiac injury in 79 (33%). During a median follow-up time of 179 (20-450) days, 82 (34%) patients experienced at least one adverse outcome. Subacute presentation was an independent predictor of adverse outcomes. Patients with postcardiac injury pericarditis had a lower incidence in the composite of failure of treatment and recurrence (13% vs 26%; P = .022) compared with patients with idiopathic pericarditis. Troponin I measurements were obtained in 167 patients (70%). Elevated troponin I levels were associated with lower incidence of recurrences (4% vs 17%; P = .024) and of the composite outcome (13% vs 36%; P = .004). Interpretation Acute pericarditis is associated with at least one adverse outcome in 34% of patients. Subacute presentation and idiopathic etiology are associated with higher incidence of adverse outcomes, whereas elevated troponin I levels identify a group with reduced risk of recurrences.

Highlights •Pericarditis is the most common disease of the pericardium. Generally self-limiting, pericarditis can be fraught by a significant risk of acute complications and of recurrences. •Prompt diagnosis and appropriate treatment of acute pericarditis may reduce the risk of acute complications and recurrences. •New therapies, such as IL-1 blockers, show promising results in patients with recurrent/refractory pericarditis. •Future studies are needed to deepen the knowledge about pericarditis pathophysiology and provide targeted therapies.

The quest of defeating cancer and improving prognosis in survivors has generated remarkable strides forward in research and have advanced the development of new antineoplastic therapies. These achievements, combined with rapid screening and early detection, have considerably extended the life expectancy of patients surviving multiple types of malignancies. Consequently, chemotherapy-related toxicity in several organ systems, especially the cardiovascular system, has surfaced as one of the leading causes of morbidity and mortality among cancer survivors. Recent evidence classifies chemotherapy-induced cardiotoxicity as the second-leading cause of morbidity and mortality, closely comparing with secondary cancer malignancies. While a certain degree of cardiotoxicity has been reported to accompany most chemotherapies, including anthracyclines, anti-metabolites, and alkylating agents, even the latest targeted cancer therapies such as immune checkpoint inhibitors and tyrosine kinase inhibitors have been associated with acute and chronic cardiac sequelae. In this chapter, we focus on describing the principal mechanism(s) for each class of chemotherapeutic agents that lead to cardiotoxicity and the innovative translational research approaches that are currently being explored to prevent or treat cancer therapy-induced cardiotoxicity and related cardiac complications.

Objectives: Because of its multifaceted cardioprotective effects, remote ischemic pre-conditioning (RIPC) was examined as a strategy to attenuate doxorubicin (DOX) cardiotoxicity. Background: The use of DOX is limited by dose-dependent cardiotoxicity and heart failure. Oxidative stress, mitochondrial dysfunction, inflammation, and autophagy modulation have been proposed as mediators of DOX cardiotoxicity. Methods: After baseline echocardiography, adult male CD1 mice were randomized to either sham or RIPC protocol (3 cycles of 5 min femoral artery occlusion followed by 5 min reperfusion) 1 h before receiving DOX (20 mg/kg, intraperitoneal). The mice were observed primarily for survival over 85 days (86 mice). An additional cohort of 50 mice was randomized to either sham or RIPC 1 h before DOX treatment and was followed for 25 days, at which time cardiac fibrosis, apoptosis, and mitochondrial oxidative phosphorylation were assessed, as well as the expression profiles of apoptosis and autophagy markers. Results: Survival was significantly improved in the RIPC cohort compared with the sham cohort (p = 0.007). DOX-induced cardiac fibrosis and apoptosis were significantly attenuated with RIPC compared with sham (p < 0.05 and p < 0.001, respectively). Although no mitochondrial dysfunction was detected at 25 days, there was a significant increase in autophagy markers with DOX that was attenuated with RIPC. Moreover, DOX caused a 49% decline in cardiac BCL2/BAX expression, which was restored with RIPC (p < 0.05 vs. DOX). DOX also resulted in a 17% reduction in left ventricular mass at 25 days, which was prevented with RIPC (p < 0.01), despite the lack of significant changes in left ventricular ejection fraction. Conclusions: Our preclinical results suggested that RIPC before DOX administration might be a promising approach for attenuating DOX cardiotoxicity.

Background: Aging is associated with metabolic and structural changes causing heart failure with preserved ejection fraction (HFpEF). Interleukin-1 (IL-1) is a pro-inflammatory cytokine involved in aging-related inflammation. We sought to determine whether IL-1 mediates aging-related changes in the heart as seen in HFpEF. Hypothesis: We hypothesize that mice with genetic deletion of IL-1RI, not responsive to IL-1β, would be protected from age-associated cardiac dysfunction. Methods: We studied age-matched young (4-month-old), middle-aged (14-month-old) and old (23-month-old) wild-type (WT) C57BL/6J and IL-1 receptor type I deficient (IL1RI-KO) male mice. Echocardiography was used to left ventricular (LV) dimensions and systolic/diastolic function, and a pressure-transducer was used to measure the LV end-diastolic pressure. Picrosirius red stain was used to assess for myocardial interstitial fibrosis (MIF) at pathology. Results: WT and IL-1RIKO mice showed a normal cardiac phenotype at young age, without any differences in between the two groups. With aging, the WT mice developed LV concentric hypertrophy (as measured by a significant increase in LV mass [+42%, P Conclusions: Genetically-modified mice lacking the IL-1RI receptor, not responsive to IL-1, are protected from aging-related LV hypertrophy, fibrosis and diastolic dysfunction. These data support a central role of IL-1 in the pathophysiology of aging-related HFpEF.

Aging is associated with metabolic and structural changes causing heart failure with preserved ejection fraction (HFpEF). Interleukin-1 (IL-1) is a pro-inflammatory cytokine involved in aging-related inflammation.We sought to determine whether IL-1 mediates aging-related changes in the heart, as seen in HFpEF.We studied age-matched young (4-month-old), middle-aged (14-month-old), and old (23-month-old) wild-type (WT) C57BL/6J and IL-1 receptor type I deficient (IL1RI-KO) male mice. Echocardiography was used to evaluate left ventricular (LV) dimensions and systolic/diastolic function, and a pressure transducer was used to measure the LV end-diastolic pressure. Picrosirius red stain was used to assess for myocardial interstitial fibrosis (MIF) at pathology.WT and IL-1RIKO mice showed a normal cardiac phenotype at young age, without any differences between the two groups. With aging, the WT mice developed LV concentric hypertrophy (as measured by a significant increase in LV mass [+42%, P 0.01] and relative wall thickness [+34%, P 0.01]), whereas the aging IL-1RI-KO mice did not. With aging, the WT mice also developed diastolic dysfunction (as measured by a significant increase in isovolumetric relaxation time [+148%, P 0.01] and a significantly higher LV end-diastolic pressure [+174%, P 0.01]), whereas the aging IL1RI-KO did not. Aged WT mice showed a significant increase in MIF (+124%, P 0.01) at cardiac pathology, whereas the aging IL-1RI-KO did not.Genetically-modified mice lacking the IL-1RI receptor, not responsive to IL-1, are protected from aging-related LV hypertrophy, fibrosis, and diastolic dysfunction. These data support a central role of IL-1 in the pathophysiology of aging-related HFpEF.

Interleukin-1β (IL-1β), a product of the NLRP3 inflammasome, modulates cardiac contractility and diastolic function. We proposed that OLT1177® (dapansutrile), a novel NLRP3 inhibitor, could preserve contractile reserve and diastolic function after myocardial infarction (MI). We used an experimental murine model of severe ischemic cardiomyopathy through the ligation of the left coronary artery without reperfusion, and after 7 days randomly assigned mice showing large anterior MI (&gt, 4 akinetic segments), increased left ventricular (LV) dimensions ([LVEDD] &gt, 4.4 mm), and reduced function (LV ejection fraction &lt, 40%) to a diet that was enriched with OLT1177® admixed with the chow in the diet at 3.75 g/kg (Group 1 [n = 10]) or 7.5 g/kg (Group 2 [n = 9]), or a standard diet as the no-treatment control group (Group 3 [n = 10]) for 9 weeks. We measured the cardiac function and contractile reserve with an isoproterenol challenge, and the diastolic function with cardiac catheterization at 10 weeks following the MI surgery. When compared with the control (Group 3), the mice treated with OLT1177 (Group 1 and 2) showed significantly greater preservation of their contractile reserve (the percent increase in the left ventricular ejection fraction [LVEF] after the isoproterenol challenge was +33 ± 11% and +40 ± 6% vs. +9 ± 7% in the standard diet, p &lt, 0.05 and p &lt, 0.005 for Group 1 and 2, respectively) and of diastolic function measured as the lower left ventricular end-diastolic pressure (3.2 ± 0.5 mmHg or 4.5 ± 0.5 mmHg vs. 10.0 ± 1.6 mmHg, 0.005 and p &lt, 0.009 respectively). No differences were noted between the resting LVEF of the MI groups. These effects were independent of the effects on the ventricular remodeling after MI. NLRP3 inflammasome inhibition with OLT1177® can preserve β-adrenergic responsiveness and prevent left ventricular diastolic dysfunction in a large non-reperfused anterior MI mouse model. OLT1177® could therefore be used to prevent the development of heart failure in patients with ischemic cardiomyopathy.

Acute myocardial infarction (AMI) is associated with the induction of a sterile inflammatory response that leads to further injury. The NACHT, leucine-rich repeat, and pyrin domain-containing protein 3 (NLRP3) inflammasome is a macromolecular structure responsible for the inflammatory response to injury or infection. NLRP3 can sense intracellular danger signals, such as ischemia and extracellular or intracellular alarmins during tissue injury. The NLRP3 inflammasome is primed and triggered by locally released damage-associated molecular patterns and amplifies the inflammatory response and cell death through caspase-1 activation. Here, we examine the scientific evidence supporting a role for NLRP3 in AMI and the available strategies to inhibit the effects of the inflammasome. Our focus is on the beneficial effects seen in experimental models of AMI in preclinical animal models and the initial results of clinical trials.

BACKGROUND Activation of the NLRP3 inflammasome is a primary driver of sterile inflammation in response to myocardial ischemia reperfusion. Pharmacologic inhibitors of the NLRP3 inflammasome are being developed. We proposed that OLT1177 (dapansutrile), a novel NLRP3 inflammasome inhibitor, could preserve myocardial function after ischemia reperfusion injury in the mouse. METHODS We used an experimental murine model of myocardial ischemia reperfusion injury through transient ligation of the left coronary artery and measured the effects of OLT1177 (6, 60, or 600 mg/kg intraperitoneal dose) on infarct size at pathology and on systolic cardiac function at echocardiography. To simulate a clinical scenario, we investigated the time window of therapeutic intervention with OLT1177 (60 mg/kg) administered 60, 120, or 180 minutes after reperfusion. RESULTS OLT1177 was rapidly detectable in the plasma following intraperitoneal injection and had no effect on cardiac function in healthy mice. OLT1177 treatment at reperfusion showed significant dose-dependent reduction in infarct size (-36%, -67%, and -62% for 6, 60, and 600 mg/kg, respectively; P < 0.001 for linear trend, P = 0.010 vs. vehicle for 6 mg/kg, and P < 0.001 vs. vehicle for 60 and 600 mg/kg) and preserved cardiac systolic function measured as left ventricular fractional shortening at 24 hours and 7 days after injury (P = 0.015 for 6 mg/kg and P < 0.01 for 60 and 600 mg/kg). OLT1177 reduced infarct size also when given after 60 minutes of reperfusion (-71%, P < 0.001 vs. vehicle). CONCLUSION OLT1177 (dapansutrile) limits infarct size and prevents left ventricular systolic dysfunction when given within 60 minutes following ischemia reperfusion injury in the mouse.

After radiation exposure, endothelium-dependent vasorelaxation is impaired due to impaired nitric oxide production. Endothelial dysfunction is characterized by uncoupled endothelial nitric oxide synthase activity, oxidation of the reduced cofactor tetrahydrobiopterin to dihydrobiopterin as one well recognized mechanism. Oral treatment with sepiapterin, a tetrahydrobiopterin precursor, decreased infiltrating inflammatory cells and cytokine levels in mice with colitis. We therefore tested whether a synthetic sepiapterin, PTC923, might mitigate radiation-induced cardiac and pulmonary injuries. C57L/J wild-type 6-8-week-old mice of both sexes received 5 Gy total-body irradiation (TBI), followed by a top-up dose of 6.5 Gy to the thorax (total thoracic dose of 11.5 Gy). Starting from 24 h postirradiation, mice were treated once daily with 1 mg/kg PTC923 for six days by oral gavage. Assessment of lung injury by breathing rate was measured every other week and echocardiography to assess heart function was performed at different time points (8, 30, 60, 90 and 180 days). Plasma proteins (fibrinogen, neutrophil elastase, C-reactive protein, and IL-6) were assessed as well. TBI induced a reduction in cardiac contractile reserve and an impairment in diastolic function restored by daily oral PTC923. Postirradiation lung injury was significantly delayed by PTC923. TBI mice treated with PTC923 experienced a longer survival compared to nonirradiated mice (71% vs. 40% of mice alive after 180 days). PTC923-treated mice showed a reduction in inflammatory mediators, especially IL-6 and IL-1b. In conclusion, these findings support the proposal that PTC923 is a potential mitigator of cardiac and lung injury caused by TBI.

Heart transplantation is a lifesaving procedure, which is limited by the availability of donor hearts. Using hearts from donors after circulatory death, which have sustained global ischemia, requires thorough studies on reliable and reproducible models that developing researchers may not have mastered. By combining the most recent literature and our recommendations based on observations and trials and errors, the methods here detail a sound in vivo heterotopic heart transplantation model for rats in which protective interventions on the ischemic heart can be studied, and thus allowing the scientific community to advance organ preservation research. Knowledge gathered from reproducible animal models allow for successful translation to clinical studies.

Objective The orf8b protein of the coronavirus SARS-CoV, analogous to SARS-CoV-2, triggers the NLRP3 inflammasome in macrophages in vitro. Deregulated inflammasome-mediated release of interleukin-1 family cytokines is important in hyper-inflammatory syndromes, like happens in SARS-CoV-2-mediated cytokine release syndrome. We propose that an intense inflammasome formation characterizes the lungs of patients with fatal COVID-19 disease due to pneumonia and acute respiratory distress syndrome (ARDS). Methods Samples from four patients with confirmed COVID-19 pneumonia who had been hospitalized at the Hospital of the University of Trieste (Italy) and died of ARDS and four lung samples from a historical repository from subjects who had died of cardiopulmonary arrest and had not been placed on mechanical ventilation and without evidence of pulmonary infection at postmortem examination were collected. Pathology samples had been fixed in formalin 10% at time of collection and subsequently embedded in paraffin. We conducted staining for ASC (Apoptosis-associated Speck-like protein containing a Caspase recruitment domain), NLRP3 (NACHT, LRR, and PYD domains-containing protein 3), and cleaved caspase-1. Results Intense expression of the inflammasome was detected, mainly in leukocytes, within the lungs of all patients with fatal COVID-19 in the areas of lung injury. The number of ASC inflammasome specks per high power fields was significantly higher in the lungs of patients with fatal COVID-19 as compared with the lungs of control subjects (52 ± 22 vs 6 ± 3, P = 0.0064). Conclusions These findings identify the presence of NLRP3 inflammasome aggregates in the lungs of fatal COVID-19 pneumonia thus providing the potential molecular link between viral infection and cytokine release syndrome. Electronic supplementary material The online version of this article (10.1007/s00011-020-01413-2) contains supplementary material, which is available to authorized users.

Significance The nucleotide-binding domain, leucine-rich containing family, pyrin domain-containing-3 (NLRP3) inflammasome, an intracellular complex that regulates maturation and release of interleukin (IL)-1β, is active in biopsies of metastatic melanoma. Here, we demonstrate that NLRP3 activation in melanoma cells drives tumor progression in mice. Subsequent to NLRP3 activation in melanoma cells, IL-1β induces melanoma-associated inflammation, resulting in immunosuppression. Oral administration of a single NLRP3 inhibitor (OLT1177) reduces melanoma growth and melanoma-associated myeloid-derived suppressor cell expansion. Inhibition of the NLRP3 signaling in combination with anti–PD-1 revealed augmented efficacy compared to monotherapy. These data propose that NLRP3 is a therapeutic target for human melanoma., Interleukin-1β (IL-1β)–mediated inflammation suppresses antitumor immunity, leading to the generation of a tumor-permissive environment, tumor growth, and progression. Here, we demonstrate that nucleotide-binding domain, leucine-rich containing family, pyrin domain-containing-3 (NLRP3) inflammasome activation in melanoma is linked to IL-1β production, inflammation, and immunosuppression. Analysis of cancer genome datasets (TCGA and GTEx) revealed greater NLRP3 and IL-1β expression in cutaneous melanoma samples (n = 469) compared to normal skin (n = 324), with a highly significant correlation between NLRP3 and IL-1β (P < 0.0001). We show the formation of the NLRP3 inflammasome in biopsies of metastatic melanoma using fluorescent resonance energy transfer analysis for NLRP3 and apoptosis-associated speck-like protein containing a CARD. In vivo, tumor-associated NLRP3/IL-1 signaling induced expansion of myeloid-derived suppressor cells (MDSCs), leading to reduced natural killer and CD8+ T cell activity concomitant with an increased presence of regulatory T (Treg) cells in the primary tumors. Either genetic or pharmacological inhibition of tumor-derived NLRP3 by dapansutrile (OLT1177) was sufficient to reduce MDSCs expansion and to enhance antitumor immunity, resulting in reduced tumor growth. Additionally, we observed that the combination of NLRP3 inhibition and anti–PD-1 treatment significantly increased the antitumor efficacy of the monotherapy by limiting MDSC-mediated T cell suppression and tumor progression. These data show that NLRP3 activation in melanoma cells is a protumor mechanism, which induces MDSCs expansion and immune evasion. We conclude that inhibition of NLRP3 can augment the efficacy of anti–PD-1 therapy.

INTRODUCTION Pericarditis is an inflammatory disease of the pericardium. Progress has been done in recent years in the understanding of its pathophysiology. In particular, preclinical and clinical studies have contributed to increasing our knowledge on the role of interleukin (IL)-1 and NLRP3 (NACHT, leucine-rich repeat, and pyrin domain-containing protein 3) inflammasome. EVIDENCE ACQUISITION Information for this study has been retrieved in original articles, reviews, systematic reviews, and meta-analyses identified through PubMed using the following search terms (or combination of terms): "pericarditis," "acute pericarditis," "recurrent pericarditis," "idiopathic recurrent acute pericarditis," "autoimmunity," "autoinflammation," "outcomes." Only articles published in English were included. Additional papers identified from the reference list of the retrieved articles were also considered. EVIDENCE SYNTHESIS Based on current evidence, pericarditis should be considered as an inflammatory reaction to various stimuli, including chemical/physical, infectious, or ischemic ones, with a viral infection being a common etiology. Interaction of pathogens or irritants with toll-like receptor (TLRs) and stimulation of IL-1 receptor by IL-1α and IL-1β leads to an increased transcription of proinflammatory genes, including those needed for NLRP3 inflammasome assembly. This pathway is confirmed indirectly by the beneficial effect of colchicine (an indirect NLRP3 inflammasome inhibitor) and IL-1 blockers in patients with recurrent pericarditis. More recently, a direct evidence of the NLRP3 inflammasome within the inflamed pericardium has been provided as well. It may, however, occur that self-antigens on the surface of mesothelial cells or microbial peptides may stimulate autoreactive T cells along with B cells producing antiheart antibodies, although less evidence is available on this. CONCLUSIONS Some uncertainties still remain about the role of neutrophils, neutrophil extracellular traps (NETs), and pericardial interstitial cells in recurrent and constrictive pericarditis. Unraveling these aspects might have a direct impact on the development of novel targeted therapies, especially considering the increasing number of drugs targeting NETs.

Background: The pleiotropic hormone relaxin has been shown to mediate physiologic responses during pregnancy by facilitating increases in vascular compliance and cardiac output. While relaxin signaling has been attributed to increases in atrial inotropy, the hormonal effects on ventricular contractility have not been explored in vivo . In this study, we investigated the dose-response effects of recombinant human relaxin (serelaxin) on ventricular mechanics. Methods and Results: Adult male CD1 mice were anesthetized and mechanically ventilated prior to Millar cardiac catheterization. A pressure-volume catheter was advanced into the left ventricle (LV), and baseline systolic and diastolic pressure/volume (PV) loops were recorded (Pre). Mice were injected with either saline, 10 μg/kg (low dose) or 1 mg/kg (high dose) of serelaxin, and PV loops were obtained after stabilization (Post) . Mice injected with 1 mg/kg demonstrated significantly higher LV contractility ( Fig A ) independent of preload ( Fig B ). These responses were not observed with low dose. In order to further examine the role of RXFP1 in modulating these responses, healthy CD1 were injected with AAV9-RXFP1 or empty vector as control and receptor overexpression was confirmed four weeks later via qPCR ( Fig C ). PV loop analysis showed an increased inotropic response [ΔP Max , Δ(dP/dt) Max ] and improved relaxation kinetics [Δ(dP/dt) Min , Δτ] in RXFP1-overexpressing mice at a lower dose of serelaxin, when compared to empty vector mice ( Fig D ). Conclusions: Our data show a novel, dose-dependent correlation between serelaxin and ventricular inotropic and lusitropic responses in mice. These responses were observed at a lower drug dose in cardiac RXFP1-overexpressing mice. We propose that further investigation of this phenomenon could advance the therapeutic, cardioprotective effects associated with relaxin therapy in different etiologies of heart failure that may exhibit changes in RXFP1 expression.

Background: Doxorubicin (DOX) is a first-line anticancer drug for the treatment of triple negative breast cancer (TNBC). However, its dose-dependent delayed and progressive cardiotoxicity limits its therapeutic application. NovoMedix (NM922) is a novel dual mTOR inhibitor/AMPK activator that was shown to attenuate adverse cardiac remodeling and fibrosis in a pressure-overload mouse model of heart failure. We investigated whether combination therapy with DOX and NM922 exhibits synergistic chemotherapeutic effect while mitigating DOX cardiotoxicity. Methods & Results: Tumors were generated in athymic female BALB/cAnNCr-nu/nu mice by implanting MDA-MB-231 cells into the rear right flank. Mice with tumors (volume≈200mm 3 ) were randomized into 6 groups and treated as follows: 1) Control (n=10); 2) DOX (3 mg/kg; i.p. twice weekly, total 15 mg/kg; n=10); 3) NM922 (25 mg/kg/d; p.o. n=5); 4) DOX+NM922 (25 mg/kg/d; p.o. n=15); 5) NM922 (100 mg/kg/d; p.o. n=5); 6) DOX+NM922 (100 mg/kg/d; p.o. n=15). Tumor size, body weight and cardiac function were assessed throughout the study. DOX alone, and to a significant extent when in combination with NM922 (25 mg/kg) reduced tumor growth compared to control. NM922 (100 mg/kg) with/without DOX significantly reduced tumor growth as compared to DOX alone (Fig A). DOX caused reduction in body weight and survival of tumor-bearing mice. NM922 did not prevent DOX-induced cachexia, but significantly improved survival in DOX-treated mice (Fig B). DOX treatment caused a significant decline in left ventricular ejection fraction compared to control over 3 weeks, which was ameliorated with NM922 (100 mg/kg) co-treatment (Fig C&D). Conclusion: Our results suggest that NM922 may potentiate the chemotherapeutic efficacy of DOX in TNBC, while mitigating its cardiotoxicity. Moreover, these findings advocate the potential efficacy of utilizing lower DOX dosages when combined with NM922, which would have significant clinical implications.

Aims: Hydrogen sulfide (H2S) protects against ischemic and inflammatory injury following myocardial ischemia via induction of microRNA (miR)-21. We sought to determine whether H2S attenuates ischemic heart failure with reduced ejection fraction (HFrEF) and interrogate the role of cofilin-2, a target of miR-21, in this protective process. Methods and Results: Adult male mice underwent myocardial infarction (MI) by coronary artery ligation after baseline echocardiography. Following MI, mice were treated with Na2S (100 μg/kg/day; intraperitoneal [IP]) or saline up to 28 days. End-diastolic pressure, measured by Millar catheter, was significantly increased ( P < .05 vs sham) at 3 days post-MI in the saline group, which was attenuated with Na2S. Left ventricular (LV) fractional shortening decreased significantly at 28 days post-MI in the saline group but was preserved with Na2S and LV infarct scar size was smaller in Na2S group as compared to control. Apoptotic signaling, measured by Bcl-2/Bax ratio, was significantly increased in the saline group but was mitigated with Na2S. Survival rate was 2-fold higher in Na2S group compared to saline control ( P < .05). Proteomic analysis and Matrix-Assisted Laser Desorption/Ionization-Time of Flight (TOF)/TOF tandem mass spectrometry identified significant changes in proapoptotic cofilin-2 expression, a specific target of miR-21, between saline- and sodium sulfide -treated mice at 28 days post-MI. Western blot analysis confirmed a significant increase in cofilin-2 after MI, which was suppressed with Na2S treatment. Chronic Na2S treatment also attenuated inflammasome formation and activation leading to reduction of maladaptive signaling. Conclusion: Na2S treatment after MI preserves LV function and improves survival through attenuation of inflammasome-mediated adverse remodeling. We propose H2S donors as promising therapeutic tools for ischemic HFrEF.

A longitudinal molecular model of the development and progression of nonalcoholic fatty liver disease (NAFLD) over time is lacking. We have recently validated a high fat/sugar water-induced animal (an isogenic strain of C57BL/6 J:129S1/SvImJ mice) model of NAFLD that closely mimics most aspects of human disease. The hepatic transcriptome of such mice with fatty liver (8 weeks), steatohepatitis with early fibrosis (16-24 weeks) and advanced fibrosis (52 weeks) after initiation of the diet was evaluated and compared to mice on chow diet. Fatty liver development was associated with transcriptional activation of lipogenesis, FXR-RXR, PPAR-α mediated lipid oxidation and oxidative stress pathways. With progression to steatohepatitis, metabolic pathway activation persisted with additional activation of IL-1/inhibition of RXR, granulocyte diapedesis/adhesion, Fc macrophage activation, prothrombin activation and hepatic stellate cell activation. Progression to advanced fibrosis was associated with dampening of metabolic, oxidative stress and cell stress related pathway activation but with further Fc macrophage activation, cell death and turnover and activation of cancer-related networks. The molecular progression of NAFLD involves a metabolic perturbation which triggers subsequent cell stress and inflammation driving cell death and turnover. Over time, inflammation and fibrogenic pathways become dominant while in advanced disease an inflammatory-oncogenic profile dominates.

BACKGROUND: Despite the benefits of reperfusion in limiting myocardial injury, the infarct size continues to expand after reperfusion because of secondary inflammatory injury. Plasma-derived alpha-1 antitrypsin (AAT) inhibits the inflammatory injury in myocardial ischemia and reperfusion. To explore the effects of plasma-derived AAT on the inflammatory response to ischemia-reperfusion injury, we analyzed time-to-reperfusion and enzymatic infarct size estimates in a post hoc analysis of the VCU-a1RT clinical trial (clinicaltrials.gov NCT01936896). METHODS: Ten patients with ST-segment elevation acute myocardial infarction (STEMI) were enrolled in an open-label, single-arm treatment study of Prolastin C, plasma-derived AAT, at 60 mg/kg infused intravenously within 12 hours of reperfusion. Biomarkers were measured serially over the first 72 hours, and patients were followed clinically for the occurrence of new-onset heart failure, recurrent MI, or death. Twenty patients with STEMI who had been enrolled in previous randomized trials with identical inclusion/ exclusion criteria and had been assigned to placebo served as historical controls. RESULTS: Time to percutaneous coronary intervention and time to drug did not significantly differ between groups. AAT-treated patients had a significantly shorter time-to-peak creatine kinase myocardial band (CK-MB) values (525 [480–735] vs. 789 [664– 959] minute, P = 0.005) and CK-MB area under the curve (from 1204 [758–2728] vs. 2418 [1551–4289] U$day, P = 0.035), despite no differences in peak CK-MB (123 [30–196] vs. 123 [71–213] U/ mL, P = 0.71). CONCLUSIONS: A single administration of Prolastin C given hours after reperfusion in patients with STEMI led to a significant shorter time to peak and area under the curve for CK-MB, despite similar peak CK-MB values. These preliminary data support the hypothesis that Prolastin C shortens the duration of the ischemia-reperfusion injury in patients with STEMI.

Visual Abstract, Highlights • The effects of UFA on CRF in patients with HFpEF are unknown. • In obese HFpEF patients, UFA consumption analyzed with a validated 24-h dietary recall was positively associated with improved body composition, cardiac diastolic function and greater CRF, measured as peak VO2 at maximal cardiopulmonary exercise testing. Conversely, sugars consumption was associated with worse CRF. • In mice, an isocaloric high-fat diet, high in UFA and low in saturated fat prevented cardiac diastolic dysfunction measured with echocardiography and body weight gain in a model of cardiac dysfunction and obesity induced by Western diet, despite similar total caloric intake. • A high-UFA diet is associated with preservation of CRF in patients with HFpEF and cardiac function in the mouse., Summary Heart failure with preserved ejection fraction (HFpEF) is associated with obesity and, indirectly, with unhealthy diet. The role of dietary components in HFpEF is, however, largely unknown. In this study, the authors showed that in obese HFpEF patients, consumption of unsaturated fatty acids (UFA), was associated with better cardiorespiratory fitness, and UFA consumption correlated with better diastolic function and with greater fat-free mass. Similarly, mice fed with a high-fat diet rich in UFA and low in sugars had preserved myocardial function and reduced weight gain. Randomized clinical trials increasing dietary UFA consumption and reducing sugar consumption are warranted to confirm and expand our findings.

We measured peak oxygen consumption (VO(2)) in previous recipients of thoracic radiotherapy and assessed the determinants of cardiorespiratory fitness with an emphasis on cardiac and pulmonary function. Cancer survivors who have received thoracic radiotherapy with incidental cardiac involvement often experience impaired cardiorespiratory fitness, as measured by reduced peak VO(2), a marker of impaired cardiovascular reserve. We enrolled 25 subjects 1.8 (0.1 to 8.2) years following completion of thoracic radiotherapy with significant heart exposure (at least 10% of heart volume receiving at least 5 Gray). All subjects underwent cardiopulmonary exercise testing, Doppler echocardiography, and circulating biomarkers assessment. The cohort included 16 Caucasians (64%), 15 women (60%) with a median age of 63 (59 to 66) years. The peak VO(2) was 16.8 (13.5 to 21.9) ml·kg(−1)·min(−1) or moderately reduced at 62% (50% to 93%) of predicted. The mean cardiac radiation dose was 5.4 (3.7 to 14.7) Gray, and it significantly correlated inversely with peak VO(2) (R = −0.445, p = 0.02). Multivariate regression analysis revealed the diastolic functional reserve index and the N-terminal pro-brain natriuretic peptide (NTproBNP) serum levels were independent predictors of peak VO(2) ( = +0.813, p

Background: B7-33 is a peptide derived from the B-chain of human relaxin, and is shown to elicit biased signaling at relaxin receptor 1 (RXFP1) in human cell lines. The influence of B7-33 on myocardial ischemia-reperfusion (IR) injury and associated cardiac adverse remodeling is unknown. Methods and Results: Primary cardiomyocytes and cardiac fibroblasts were isolated from adult CD1 mice and subjected to simulated ischemia-reoxygenation (SIRO). Myocytes were placed in 1% O 2 chamber for 40 min, followed by 1 h of reoxygenation with control (myocyte media) or B7-33 (10, 50 or 100 nM)-infused media. Trypan blue staining showed a significant decrease in cell death with B7-33 concentrations of 50 nM and 100 nM ( Fig. A ).Treating myocytes with B7-33 for 15 min exhibited a dose-dependent increase in Erk1/2 phosphorylation, which reached statistical significance at 100 nM ( Fig. B ). Fibroblasts subjected to 4 h of hypoxia and 15 h of reoxygenation had increased viability (MTT assay) with B7-33 at all concentrations ( Fig. C ). In vivo , CD1 mice were subjected to IR injury via coronary artery ligation for 30 min, followed by 24 h of reperfusion. Vehicle (saline) or B7-33 (10 μg/kg) was injected i.p. at the onset of reperfusion. After 24 h, B7-33-treated mice demonstrated decreased infarct size (TTC stain) ( Fig. D ) and preserved fractional shortening (M-mode echo, Fig. E ) compared to vehicle-treated mice. Conclusion: Reperfusion therapy with B7-33 reduces infarct size post-MI, preserves cardiac function and protects cardiomyocytes and fibroblasts against SIRO. We propose that B7-33 might be effective against acute MI and a possible alternative to recombinant relaxin for clinical utility.

Background: Relaxin signaling confers cardioprotection against acute myocardial infarction (MI) and mitigates adverse cardiac remodeling. We investigated the role of relaxin receptor (RXFP1) overexpression via cardiotropic AAV9 vectors in attenuating MI-associated adverse remodeling. Methods and Results: Murine RXFP1 cDNA with CMV promoter was introduced into AAV cis plasmids for generation of AAV9-RXFP1 viral vectors. 1.0 x 10 11 viral genomes in 100μL saline were administered i.v. to 8-week-old CD1 male mice. Four weeks later, overexpression was confirmed via qPCR of RXFP1 mRNA in whole cardiac tissue lysates ( Fig A ) and primary cardiomyocytes ( Fig B ) versus control. Overexpressing (AAV9-RXFP1) mice were subjected to MI (30 minutes of coronary artery ligation followed by reperfusion for 24h or 7d). Immunohistochemical staining of cardiac sections 7d post-MI shows increased RXFP1 expression in AAV9-RXFP1 mice, as shown in Fig C . Infarct size was reduced at 24h post-MI (TTC staining, Fig D ) and fractional shortening (M mode echo) was preserved at 24h and 7d post-MI in AAV9-RXFP1 mice ( Fig E ); endocardial strain rate assessment (speckle tracking analysis) showed preserved function in the affected ventricular segments of AAV9-RXFP1 mice 7d post-MI ( Fig F ). Picrosirius staining showed reduced LV fibrosis at 7d post-MI in AAV9-RXFP1 mice ( Fig G ). Conclusion: Administration of AAV9-RXFP1 successfully upregulates RXFP1 in cardiac tissue and primary cardiomyocytes, reduces infarct size and fibrosis, and preserves cardiac function post MI. Targeting cardiac RXFP1 via gene therapy could be a novel strategy to prevent MI-related adverse remodeling.

Acute myocardial infarction (AMI) leads to myocardial cell death and ensuing sterile inflammatory response, which represents an attempt to clear cellular debris and promote cardiac repair. However, an overwhelming, unopposed or unresolved inflammatory response following AMI leads to further injury, worse remodeling and heart failure (HF). Additional therapies are therefore warranted to blunt the inflammatory response associated with ischemia and reperfusion and prevent long-term adverse events. Low-density lipoprotein receptor-related protein 1 (LRP1) is a ubiquitous endocytic cell surface receptor with the ability to recognize a wide range of structurally and functionally diverse ligands. LRP1 transduces multiple intracellular signal pathways regulating the inflammatory reaction, tissue remodeling and cell survival after organ injury. In preclinical studies, activation of LRP1-mediated signaling in the heart with non-selective and selective LRP1 agonists is linked with a powerful cardioprotective effect, reducing infarct size and cardiac dysfunction after AMI. The data from early phase clinical studies with plasma-derived α1-antitrypsin (AAT), an endogenous LRP1 agonist, and SP16 peptide, a synthetic LRP1 agonist, support the translational value of LRP1 as a novel therapeutic target in AMI. In this review, we will summarize the cellular and molecular bases of LRP1 functions in modulating the inflammatory reaction and the reparative process after injury in various peripheral tissues, and discuss recent evidences implicating LRP1 in myocardial inflammation and infarct healing.

Introduction: Indolent B-Cell Non-Hodgkin's Lymphomas (NHL) represent a heterogeneous group of lymphoproliferative malignancies, that remain largely incurable. Marginal zone lymphomas (MZL) are the second most common subtype of indolent NHL, and lack a unique cytogenetic identifying abnormality. The B-cell receptor signaling pathway is activated in B-cell malignancy and mediates its activity mainly through the Phosphoinositide 3-kinase (PI3K) pathway. Furthermore, novel PI3K inhibitors, such as copanlisib and parsaclisib, have shown impressive clinical activity in several indolent lymphomas including MZL. This further supports the important role of the PI3K pathway in the pathogenesis of this tumor. Therefore, we hypothesized that the PI3K-mTOR pathway is sufficient for driving the pathogenesis of MZL. Methods: In order to test our hypothesis, we generated a genetically engineered mouse model carrying heterozygous global knockout alleles of both the tumor suppressor genes Phosphatase and Tensin Homolog (PTEN) and Liver Kinase B1 (LKB1). This led to over-activation of the PI3K-mTOR pathway in all mouse tissues. We closely monitored these mice for tumor formation via weekly physical examinations for several months. Upon tumor detection, the mouse was sacrificed, and tumors were sectioned for histological characterization. In order to generate a more specific model of B cells, and more accurately mimic the underlying human disease, we used the Cre-LoxP system to create the CD19-Cre-PTENfl/fl-LKB1fl/fl. Results: Thirty mice of global KO PTEN+/- LKB1 +/- died or were sacrificed due to disease progression, defined as either lymph node enlargement and/or splenomegaly. All mice showed either abnormal lymphadenopathy or splenomegaly. By Kaplan-Meier analysis, we saw a steady decrease in both tumor-free and overall survival after 3 months of age. Utilizing the product limit method, the median survival time was 6 months (95% CI: 6, 8). A total of 51 lymph nodes were sent for immunohistochemistry and pathological characterization. Of the 51 nodes, 61.5% (N=32) showed indolent Non-Hodgkin's Lymphoma, 25% (N=13) were atypical, and 11.5% (N=6) were reactive. All lymph nodes with indolent NHL were of MZL subtype. Compared to wild type (n=3), the new CD19-Cre-PTENfl/fl -LKB1fl/fl (n=3) showed an overall increase in spleen mass (120 vs 196 mg, p=0.0564), % B1 cells (4% vs 59%, p= 0.0075), % MZ cells (5% vs 30%, p=0.0547), % plasma cells (1% vs 12%, p=0.0729), and decrease in % FO cells (80% vs 12%, p=0.0003) by flowcytometry. Further characterization of the new model is currently underway. Conclusion: Marginal zone lymphoma remains an incurable lymphoma that lacks reliable preclinical models. Our data provides, for the first time, a proof of concept on the role of the PI3K-mTOR pathway in the pathogenesis of marginal zone lymphoma and paves the way for future studies understanding the biology of this disease, and developing rational therapies for this incurable malignancy. Disclosures Yazbeck: Celgene: Consultancy; AstraZeneca: Consultancy; Gilead: Research Funding; Seattle Genetics: Consultancy; Verastem: Speakers Bureau.

Introduction: Indolent B-cell non-Hodgkin’s lymphomas (NHL) represent a heterogeneous group of lymphoproliferative malignancies that remain largely incurable. Marginal zone lymphomas (MZL) are the second most common subtype of indolent NHL and lack a unique cytogenetic identifying abnormality. The B-cell receptor signaling pathway is activated in B-cell malignancy and mediates its activity mainly through the phosphoinositide 3-kinase (PI3K) pathway. Furthermore, novel PI3K inhibitors, such as copanlisib and parsaclisib, have shown impressive clinical activity in several indolent lymphomas, including MZL. This further supports the important role of the PI3K pathway in the pathogenesis of this tumor. Therefore, we hypothesized that the PI3K-mTOR pathway is sufficient for driving the pathogenesis of MZL. Methods: In order to test our hypothesis, we generated a genetically engineered mouse model carrying heterozygous global knockout alleles of both the tumor suppressor genes Phosphatase and Tensin Homolog (PTEN) and Liver Kinase B1 (LKB1). This led to overactivation of the PI3K-mTOR pathway in all mouse tissues. We closely monitored these mice for tumor formation via weekly physical examinations for several months. Upon tumor detection, the mouse was sacrificed and tumors were sectioned for histologic characterization. In order to generate a more specific model of B cells, and more accurately mimic the underlying human disease, we used the Cre-LoxP system to create the CD19-Cre-PTENfl/fl-LKB1fl/fl. Results: Thirty mice of global KO PTEN+/- LKB1 +/- died or were sacrificed due to disease progression, defined as either lymph node enlargement and/or splenomegaly. All mice showed either abnormal lymphadenopathy or splenomegaly. By Kaplan-Meier analysis, we saw a steady decrease in both tumor-free and overall survival after 3 months of age. Utilizing the product limit method, the median survival time was 6 months (95% CI: 6, 8). A total of 51 lymph nodes were sent for immunohistochemistry and pathologic characterization. Of the 51 nodes, 61.5% (N=32) showed indolent non-Hodgkin’s lymphoma, 25% (N=13) were atypical, and 11.5% (N=6) were reactive. All lymph nodes with indolent NHL were of MZL subtype. Compared to wild-type (n=3), the new CD19-Cre-PTENfl/fl-LKB1fl/fl (n=3) showed an overall increase in spleen mass (120 vs. 196 mg, p=0.0564), % B1 cells (4% vs. 59%, p= 0.0075), % MZ cells (5% vs. 30%, p=0.0547), % plasma cells (1% vs. 12%, p=0.0729), and a decrease in % FO cells (80% vs. 12%, p=0.0003) by flow cytometry. Further characterization of the new model is currently under way. Conclusion: Marginal zone lymphoma remains an incurable lymphoma that lacks reliable preclinical models. Our data provide, for the first time, a proof of concept on the role of the PI3K-mTOR pathway in the pathogenesis of marginal zone lymphoma and pave the way for future studies understanding the biology of this disease and developing rational therapies for this incurable malignancy. Citation Format: Victor Yazbeck, Ian McConnell, Emily Harris, Joseph Lownick, Ariel Sindel, Roy Sabo, Alden Chesney, Guanhua Lai, Adolfo Mauro, Chad Cain, Fadi Salloum, Steven Grant, Jamal Zweit, Jolene Windle, Steven Grossman. Modeling marginal zone lymphomagenesis [abstract]. In: Proceedings of the AACR Special Conference on the Evolving Landscape of Cancer Modeling; 2020 Mar 2-5; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2020;80(11 Suppl):Abstract nr B41.

BACKGROUND: A diet rich in saturated fat and sugars (Western diet, WD) induces myocardial expression of the NLRP3 inflammasome and dysfunction in mice. We therefore hypothesized that a diet enriched with an orally available NLRP3 inflammasome inhibitor could prevent WD-induced cardiac dysfunction in mice. METHODS: Ten-week-old CD-1 male mice were fed WD or standard diet (SD) for 8 weeks. The compound 16673-34-0, an orally active NLRP3 inhibitor, was added to the diet at a concentration of 100 mg/Kg. The plasmatic levels of the NLRP3 inflammasome inhibitor were measured. Food intake, body weight, and glucose tolerance were assessed. Cardiac systolic and diastolic functions were measured by Doppler echocardiography at baseline, 4 weeks, and 8 weeks. RESULTS: WD induced a significant increase in body weight (+14%, P = 0.02), impaired glucose tolerance (+34%, P = 0.03), and a significant increase in isovolumetric relaxation time (+129%, P = 0.03) and reduction in left ventricular ejection fraction (−10%, P = 0.03), as compared to standard chow diet (SD). The treatment with NLRP3 inhibitor in the diet prevented cardiac systolic and diastolic dysfunction (P < 0.05 for left ventricular ejection fraction, isovolumetric relaxation time, and myocardial performance index in WD with drug vs. WD without drug), without significant changes in heart rate and metabolic parameters. CONCLUSIONS: An orally available NLRP3 inhibitor prevented WD-induced cardiac dysfunction in obese mice.

Background Sacubitril/valsartan (SAC/VAL) is approved by the U.S. Food and Drug Administration for heart failure with reduced ejection fraction (HFrEF). Objectives This study investigated the effects of SAC/VAL on acute myocardial infarction (MI) and cardiac remodeling in a translational rabbit model of MI. Methods New Zealand White rabbits were sedated and underwent conscious MI (45-min ischemia) by balloon inflation (previously implanted surgically) followed by 72 h (acute protocol) or 10 weeks (chronic protocols) of reperfusion. “Infarct-sparing” protocol: SAC/VAL, VAL, or placebo were randomly allocated and administered at reperfusion. “HFrEF-treatment” protocol: rabbits were randomized, and treatment commenced after echocardiography-confirmed left ventricular ejection fraction (LVEF) ≤40%. “HFrEF-prevention” protocol: treatment started at reperfusion and continued daily throughout the study. Results Compared with placebo, SAC/VAL and VAL significantly reduced infarct size (TTC staining) and plasma troponin levels; however, only SAC/VAL preserved LVEF at 72 h post-MI. In the HFrEF-treatment protocol, LVEF improvement was observed with SAC/VAL compared with both placebo and VAL starting 2 weeks post-treatment, a benefit that persisted throughout study duration. In the HFrEF-prevention protocol, SAC/VAL and VAL attenuated the decline in LVEF post-MI, although SAC/VAL offered better functional protection. The functional improvement observed in both treatment protocols was paralleled by significant reduction in left ventricular (LV) scar size (Picrosirius red staining) in the SAC/VAL groups. Conclusions Reperfusion therapy with SAC/VAL or VAL offers robust acute infarct-sparing benefits; however, SAC/VAL treatment offered superior short-term and long-term benefits in preventing MI-induced LV dysfunction compared with VAL. SAC/VAL also significantly attenuated LV scar size following MI compared with placebo, whereas VAL did not reach statistical significance in scar reduction.

We read with great interest the state-of-the-art review from Valero-Munoz et al. [(1)][1] in a recent issue of JACC: Basic to Translational Science . The authors described the more commonly used murine models of heart failure with preserved ejection fraction (HFpEF). They emphasize the importance

Heart failure (HF) with preserved ejection fraction (HFpEF) is a clinical syndrome of HF symptoms associated with impaired diastolic function. Although it represents ∼50% of patients with HF, the mechanisms of disease are poorly understood, and therapies are generally ineffective in reducing HF progression. Animal models of HFpEF not due to pressure or volume overload are lacking, therefore limiting in-depth understanding of the pathophysiological mechanisms and the development of novel therapies. We hypothesize that a continuous infusion of low-dose angiotensin II (ATII) is sufficient to induce left ventricular (LV) diastolic dysfunction and HFpEF, without increasing blood pressure or inducing LV hypertrophy or dilatation. Osmotic pumps were implanted subcutaneously in 8-wk-old male mice assigned to the ATII (0.2 mg·kg−1·day−1) or volume-matched vehicle ( N = 8/group) for 4 wk. We measured systolic and diastolic arterial blood pressures through a tail-cuff transducer, LV dimensions and ejection fraction through echocardiography, and LV relaxation through pulsed-wave Doppler and LV catheterization. Myocardial fibrosis and cardiomyocyte cross-sectional area were measured. ATII infusion had no effects on systemic arterial blood pressure. ATII induced significant impairment in LV diastolic function, as measured by an increase (worsening) in LV isovolumetric relaxation time, myocardial performance index, isovolumetric relaxation time constant, and LV end-diastolic pressure without altering LV dimensions, mass, or ejection fraction. Chronic infusion of low-dose ATII recapitulates the HFpEF phenotype in the mouse, without increasing systemic arterial blood pressure. This mouse model may provide insight into the mechanisms of HFpEF.

Significance: An inflammatory response follows an injury of any nature, and while such a response is an attempt to promote healing, it may, itself, result in further injury. Recent Advances: The inflammasome is a macromolecular structure recently recognized as a central mediator in the acute inflammatory response. The inflammasome senses the injury and it amplifies the response by leading to the release of powerful pro-inflammatory cytokines, interleukin-1β (IL-1β) and IL-18. Critical Issues: The activation of the inflammasome in the heart during ischemic and nonischemic injury represents an exaggerated response to sterile injury and promotes adverse cardiac remodeling and failure. Future Directions: Pilot clinical trials have explored blockade of the inflammasome-derived IL-1β and have shown beneficial effects on cardiac function. Additional clinical studies testing this approach are warranted. Moreover, specific inflammasome inhibitors that are ready for clinical use are currently lacking. Anti...

Thoracic X-ray therapy (XRT), used in cancer treatment, is associated with increased risk of heart failure. XRT-mediated injury to the heart induces an inflammatory response leading to cardiomyopathy. The aim of this study was to determine the role of inter-leukin (IL)-1 in response to XRT injury to the heart and on the cardiomyopathy development in the mouse. Female mice with genetic deletion of the IL-1 receptor type I (IL-1R1 knockout mice [IL-1R1 KO]) and treatment with recombinant human IL-1 receptor antagonist anakinra, 10 mg/kg twice daily for 7 d, were used as independent approaches to determine the role of IL-1. Wild-type (wt) or IL-1R1 KO mice were treated with a single session of XRT (20 or 14 gray [Gy]). Echocardiography (before and after isoproterenol challenge) and left ventricular (LV) catheterization were performed to evaluate changes in LV dimensions and function. Masson’s trichrome was used to assess myocardial fibrosis and pericardial thickening. After 20 Gy, the contractile reserve was impaired in wt mice at d 3, and the LV ejection fraction (EF) was reduced after 4 months when compared with sham-XRT. IL-1R1 KO mice had preserved contractile reserve at 3 d and 4 months and LVEF at 4 months after XRT. Anakinra treatment for 1 d before and 7 d after XRT prevented the impairment in contractile reserve. A significant increase in LV end-diastolic pressure, associated with increased myocardial interstitial fibrosis and pericardial thickening, was observed in wt mice, as well as in IL-1R1 KO-or anakinra-treated mice. In conclusion, induction of IL-1 by XRT mediates the development of some, such as the contractile impairment, but not all aspects of the XRT-induced cardiomyopathy, such as myocardial fibrosis or pericardial thickening.

Background: Sacubitril/Valsartan (SAC/VAL), a drug combining a neprilysin inhibitor and an angiotensin receptor blocker, was shown to reduce myocardial infarct size and left ventricular (LV) dysfunction in preclinical models of myocardial infarction (MI). In the PARADIGM-HF trial, SAC/VAL prevented the clinical progression of patients with heart failure (HF) more effectively than enalapril. Whether SAC/VAL attenuates cardiac fibrosis and improves LV function in a rabbit model of MI-induced HF with reduced ejection fraction (rEF) is unknown. Methods: Anesthetized adult male NZW rabbits (~2.5kg) underwent left thoracotomy and the left anterior descending (LAD) coronary artery was identified and occluded for 45 min followed by reperfusion. Weekly echocardiography was performed to confirm reduced EF (~40%), which was uniformly achieved at 5 weeks post MI. Subsequently, rabbits were randomized to orally receive placebo (volume-matched water, BID), SAC/VAL (10 mg/kg, BID) or VAL (9.1mg/kg/day) starting on week 6. At 10 weeks post MI, rabbits were sacrificed and hearts were harvested, fixed with 10% formalin and embedded in paraffin to assess myocardial fibrosis (Picrosirius red staining). Operators performing echocardiography, Picrosirius red staining and analysis were blinded to treatment allocation. Results: Two weeks after treatment initiation, a significant improvement in LVEF was observed in the SAC/VAL group compared to both placebo and VAL, a benefit that lasted throughout the entire study (Fig. A). The functional improvement observed was associated with a significant reduction in LV scar size compared to placebo at week 10 (Fig. B). However, when compared to VAL, the decrease in scar size did not reach statistical significance despite a clear trend. Conclusion: Our results suggest that SAC/VAL may offer superior benefits compared to equivalent dose of stand-alone VAL in attenuating LV scar size and improving LVEF in a rabbit model of ischemic HFrEF.

Donation after circulatory death donors (DCD) have the potential to increase the number of heart transplants. The DCD hearts undergo an extended period of warm ischemia, which mandates the use of machine perfusion preservation if they are to be successfully recovered for transplantation. Because the minimum coronary artery flow needed to meet the basal oxygen demand (DCRIT) of a DCD heart during machine perfusion preservation is critical and yet unknown, we studied this in a DCD rat heart model. Adult male rats were anesthetized, intubated, heparinized, and paralyzed with vecuronium. The DCD hearts (n = 9) were recovered 30 minutes after circulatory death whereas non-DCD control hearts (n = 12) were recovered without circulatory death. Hearts were perfused through the aorta with an oxygenated Belzer Modified Machine Perfusion Solution (A3-Bridge to Life Ltd. Columbia, SC) at 15°C or 22°C starting at a flow index of 300 ml/100 g/min and decreasing by 40 ml/100 g/min every 10 minutes. Inflow (aortic) and outflow (inferior vena cava) perfusate samples were collected serially to assess the myocardial oxygen consumption index (MVO2) and O2 extraction ratio. The DCRIT is the minimum coronary flow below which the MVO2 becomes flow dependent. The MVO2, DCRIT, and oxygen extraction ratios were higher in DCD hearts compared with control hearts. The DCRIT for DCD hearts was achieved only at 15°C and was significantly higher (131.6 ± 7 ml/100 g/min) compared with control hearts (107.7 ± 8.4 ml/100 gm/min). The DCD hearts sustain warm ischemic damage and manifest higher metabolic needs during machine perfusion. Establishing adequate coronary perfusion is critical to preserving organ function for potential heart transplantation.