Galectin-3 Is Associated with Cardiac Fibrosis and an Increased Risk of Sudden Death.

BACKGROUND: Myocardial fibrosis is a common postmortem finding among individuals with Sudden Cardiac Death (SCD). Numerous in vivo and in vitro studies have shown that increased galectin-3 (gal3) expression into the myocardium is associated with higher incidence of fibrosis. Although elevated gal3 expression is linked with myocardial fibrosis, its role in predicting the risk of SCD is unknown. METHODS: We reviewed the clinical datasets and post-mortem examination of 221 subjects who had died suddenly. We examined myocardial pathology including the extent of cardiac hypertrophy, fibrosis, and the degree of coronary atherosclerosis in these subjects. In a select group of SCD subjects, we studied myocardial gal3 and periostin expression using immunohistochemistry. To further examine if a higher level of circulating gal3 can be detected preceding sudden death, we measured serum gal3 in a porcine model of subtotal coronary artery ligation which shows an increased tendency to develop lethal cardiac arrhythmias, including ventricular tachycardia or fibrillation. RESULTS: Of the total 1314 human subjects screened, 12.7% had SCD. Comparison of age-matched SCD with non-SCD subjects showed that SCD groups had excessive myocardial fibrosis involving both the left ventricular free wall and interventricular septum. In pigs with subtotal coronary artery ligation and SCD, we detected significantly elevated circulating gal3 levels approximately 10 days preceding the SCD event. Immunohistochemistry showed increased myocardial gal3 and periostin expression in pigs that died suddenly, compared to the controls. CONCLUSION: Our study shows that increased gal3 is associated with a higher risk of myocardial fibrosis and the risk of SCD. This supports the importance of larger translational studies to target gal3 to prevent cardiac fibrosis and attenuate the risk of SCD.

Widespread intracoronary allogeneic cardiosphere-derived cell therapy with and without cyclosporine in reperfused myocardial infarction.

Allogeneic cardiosphere-derived cell (CDC) therapy has been demonstrated to improve myocardial function when administered to reperfused myocardial infarcts. We previously pretreated animals with low-dose cyclosporine immunosuppression to limit allogeneic CDC rejection, but whether it is necessary and, if so, can be initiated at the time of reperfusion remains uncertain. Closed-chest swine (n = 29 animals) were subjected to a 90-min left anterior descending (LAD) coronary artery occlusion. Using a three-way blinded design, we randomized two groups to receive global intracoronary infusions of 20 × 106 CDCs 30 min after reperfusion. A third control group was treated with saline. One CDC group received cyclosporine 10 min before reperfusion (2.5 mg/kg iv and 100 mg/day po), whereas the other groups received placebos. After 1 mo, neither chronic infarct size relative to area at risk (saline control, 46.2 ± 4.0%; CDCs, 46.4 ± 2.1%; and CDCs + cyclosporine, 49.2 ± 3.1%; P = 0.79) nor ejection fraction (saline control, 51 ± 2%; CDCs, 51 ± 2%; and CDC + cyclosporine, 48 ± 2%; P = 0.42) were different among treatment groups. Multiple histological measures of cellular remodeling, myocyte proliferation, and apoptosis were also not different among treatment groups. In contrast to previous studies, we were unable to reproduce the cardioprotective effects demonstrated by allogeneic CDCs without cyclosporine. Furthermore, initiation of intravenous cyclosporine at the time of reperfusion followed by oral therapy was not sufficient to elicit the functional improvement observed in studies where cyclosporine was started 72 h before CDC therapy. This suggests that oral cyclosporine pretreatment may be necessary to effect cardiac repair with allogeneic CDCs.NEW & NOTEWORTHY In a three-way blinded, randomized design, we determined whether allogeneic CDCs administered at reperfusion improved myocardial function and whether intravenous cyclosporine enhanced their efficacy. In contrast to prior studies using oral cyclosporine, CDCs with or without intravenous cyclosporine had no effect on function or infarct size. This indicates that CDCs may be most efficacious for treating chronic LV dysfunction where cyclosporine can be initiated at least 72 h before cell therapy.

Myocardial injury, troponin release, and cardiomyocyte death in brief ischemia, failure, and ventricular remodeling.

Troponin released from irreversibly injured myocytes is the gold standard biomarker for the rapid identification of an acute coronary syndrome. In acute myocardial infarction, necrotic cell death is characterized by sarcolemmal disruption in response to a critical level of energy depletion after more than 15 min of ischemia. Although troponin I and T are highly specific for cardiomyocyte death, high-sensitivity assays have demonstrated that measurable circulating levels of troponin are present in many normal subjects. In addition, transient as well as chronic elevations have been demonstrated in many disease states not clearly associated with myocardial ischemia. The latter observations have given rise to the clinical concept of myocardial injury. This review will summarize evidence supporting the notion that circulating troponin levels parallel the extent of myocyte apoptosis in normal ventricular remodeling and in pathophysiological conditions not associated with infarction or necrosis. It will review the evidence that myocyte apoptosis can be accelerated by diastolic strain from elevated ventricular preload and systolic strain from dyskinesis after brief episodes of ischemia too short to cause a critical level of myocyte energy depletion. We then show how chronic, low rates of myocyte apoptosis from endogenous myocyte turnover, repetitive ischemia, or repetitive elevations in left ventricular diastolic pressure can lead to significant myocyte loss in the absence of neurohormonal stimulation. Finally, we posit that the differential response to strain-induced injury in heart failure may determine whether progressive myocyte loss and heart failure with reduced ejection fraction or interstitial fibrosis and heart failure with preserved ejection fraction become the heart failure phenotype.

Does quantification of [11C]meta-hydroxyephedrine and [13N]ammonia kinetics improve risk stratification in ischemic cardiomyopathy.

BACKGROUND: In ischemic cardiomyopathy patients, cardiac sympathetic nervous system dysfunction is a predictor of sudden cardiac arrest (SCA). This study compared abnormal innervation and perfusion measured by [11C]meta-hydroxyephedrine (HED) vs [13N]ammonia (NH3), conventional uptake vs parametric tracer analysis, and their SCA risk discrimination. METHODS: This is a sub-study analysis of the prospective PAREPET trial, which followed ischemic cardiomyopathy patients with reduced left ventricular ejection fraction (LVEF ≤ 35%) for events of SCA. Using n = 174 paired dynamic HED and NH3 positron emission tomography (PET) scans, regional defect scores (%LV extent × severity) were calculated using HED and NH3 uptake, as well as HED distribution volume and NH3 myocardial blood flow by kinetic modeling. RESULTS: During 4.1 years follow-up, there were 27 SCA events. HED defects were larger than NH3, especially in the lowest tertile of perfusion abnormality (P < .001). Parametric defects were larger than their respective tracer uptake defects (P < .001). SCA risk discrimination was not significantly improved with parametric or uptake mismatch (AUC = 0.73 or 0.70) compared to HED uptake defect scores (AUC = 0.67). CONCLUSION: Quantification of HED distribution volume and NH3 myocardial blood flow produced larger defects than their respective measures of tracer uptake, but did not lead to improved SCA risk stratification vs HED uptake alone.

Neutrophils aid cellular therapeutics by enhancing glycoengineered stem cell recruitment and retention at sites of inflammation.

The efficacy of cell-based therapies relies on targeted payload delivery and enhanced cell retention. In vitro and in vivo studies suggest that the glycoengineering of mesenchymal and cardiosphere-derived cells (CDCs) may enhance such recruitment at sites of injury. We evaluated the role of blood cells in amplifying this recruitment. Thus, the human α(1,3)fucosyltransferase FUT7 was stably expressed in CDCs, sometimes with P-selectin glycoprotein ligand-1 (PSGL-1/CD162). Such FUT7 over-expression resulted in cell-surface sialyl Lewis-X (sLeX) expression, at levels comparable to blood neutrophils. Whereas FUT7 was sufficient for CDC recruitment on substrates bearing E-selectin under flow, PSGL-1 co-expression was necessary for P-/L-selectin binding. In both cone-plate viscometer and flow chamber studies, chemokine driven neutrophil activation promoted the adhesion of glycoengineered-CDCs to blood cells. Here, blood neutrophils activated upon contact with IL-1ß stimulated endothelial cells, amplified glycoengineered-CDC recruitment. In vivo, local inflammation in a mouse ear elicited both glycoengineered-CDC and peripheral blood neutrophil homing to the inflamed site. Glycoengineering CDCs also resulted in enhanced (~16%) cell retention at 24 h in a murine myocardial infarction model, with CDCs often co-localized with blood neutrophils. Overall, peripheral blood neutrophils, activated at sites of injury, may enhance recruitment of glycoengineered cellular therapeutics via secondary capture mechanisms.

Positron Emission Tomography Imaging of Regional Versus Global Myocardial Sympathetic Activity to Improve Risk Stratification in Patients With Ischemic Cardiomyopathy.

BACKGROUND: Current risk assessment approaches fail to identify the majority of patients at risk of sudden cardiac arrest (SCA). Noninvasive imaging of the cardiac sympathetic nervous system using single-photon emission computed tomography and positron emission tomography offers the potential for refining SCA risk assessment. While various [11C]meta-hydroxyephedrine quantification parameters have been proposed, it is currently unknown whether regional denervation or global innervation yields greater SCA risk discrimination. The aim of the study was to determine whether the global innervation parameters yield any independent and additive prognostic value over the regional denervation alone. METHODS: In a post hoc competing-risks analysis of the PAREPET trial (Prediction of Arrhythmic Events With Positron Emission Tomography), we compared global innervation and regional denervation parameters using the norepinephrine analog [11C]meta-hydroxyephedrine for SCA risk discrimination. Patients with ischemic cardiomyopathy (n=174) eligible for an implantable cardioverter-defibrillator for the primary prevention of SCA were recruited into the trial. [11C]meta-hydroxyephedrine uptake and clearance rates were measured to assess global (left ventricle mean) retention index and volume of distribution. Regional defects were quantified as the percentage of the left ventricle having values <75% of the maximum. RESULTS: During a median follow-up of 4.2 years, there were 56 cardiac-related deaths, of which 26 were SCAs. For any given regional denervation volume, there was substantial heterogeneity in global innervation scores. Global retention index and distribution volume did not decrease until regional defects exceeded 40% left ventricle. Global scale parameters, retention index, and distribution volume (area under the curve=0.61, P=0.034, P=0.046, respectively), yielded inferior SCA risk discrimination compared to regional heterogeneity (area under the curve=0.74). CONCLUSIONS: Regional denervation volume has superior cause-specific mortality prediction for SCA versus global parameters of sympathetic innervation. These results have widespread implications for future cardiac sympathetic imaging, which will greatly simplify innervation analysis. Registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT01400334.

Gaining Efficiency in Clinical Trials With Cardiac Biomarkers: JACC Review Topic of the Week.

The momentum of cardiovascular drug development has slowed dramatically. Use of validated cardiac biomarkers in clinical trials could accelerate development of much-needed therapies, but biomarkers have been used less for cardiovascular drug development than in therapeutic areas such as oncology. Moreover, there are inconsistences in biomarker use in clinical trials, such as sample type, collection times, analytical methods, and storage for future research. With these needs in mind, participants in a Cardiac Safety Research Consortium Think Tank proposed the development of international guidance in this area, together with improved quality assurance and analytical methods, to determine what biomarkers can reliably show. Participants recommended the development of systematic methods for sample collection, and the archiving of samples in all cardiovascular clinical trials (including creation of a biobank or repository). The academic and regulatory communities also agreed to work together to ensure that published information is fully and clearly expressed.

Changes in Field Termination of Resuscitation and Survival Rates After an Educational Intervention to Promote on Scene Resuscitation for Out-of-Hospital Cardiac Arrest.

BACKGROUND: Emergency medical services (EMS) agencies with higher field termination-of-resuscitation (TOR) rates tend to have higher survival rates from out-of-hospital cardiac arrest (OHCA). Whether EMS agencies can improve survival rates through efforts to focus on resuscitation on scene and optimize TOR rates is unknown. OBJECTIVE: The goal of this study was to determine if an EMS agency's efforts to enhance on-scene resuscitation were associated with increased TOR and OHCA survival with favorable neurologic outcome. METHODS: A single-city, retrospective analysis of prospectively collected 2017 quality assurance data was conducted. Patient demographics, process, and outcome measures were compared before and after an educational intervention to increase field TOR. The primary outcome measure was survival to hospital discharge with favorable neurologic status. RESULTS: There were 320 cases that met inclusion criteria. No differences in age, gender, location, witnessed arrest, bystander cardiopulmonary resuscitation, initial shockable rhythm, or presumed cardiac etiology were found. After the intervention, overall TOR rate increased from 39.6% to 51.1% (p = 0.06). Among subjects transported without return of spontaneous circulation (ROSC), average time on scene increased from 26.4 to 34.2 min (p = 0.02). Rates of sustained ROSC and survival to hospital admission were similar between periods. After intervention, there was a trend toward increased survival to hospital discharge rate (relative risk [RR] 2.09; 95% confidence interval [CI] 0.74-5.91) and an increase in survival with favorable neurologic status rate (RR 5.96; 95% CI 0.80-44.47). CONCLUSION: This study described the association between an educational intervention focusing on optimization of resuscitation on scene and OHCA process and outcome measures. Field termination has the potential to serve as a surrogate marker for aggressively treating OHCA patients on scene.

Reproducible Quantification of Regional Sympathetic Denervation with [11C]meta-Hydroxyephedrine PET Imaging.

BACKGROUND: Regional cardiac sympathetic denervation is predictive of sudden cardiac arrest in patients with ischemic cardiomyopathy. The reproducibility of denervation scores between automated software programs has not been evaluated. This study seeks to (1) compare the inter-rater reliability of regional denervation measurements using two analysis programs: FlowQuant® and Corridor4DM®; (2) evaluate test-retest repeatability of regional denervation scores. METHODS: N = 190 dynamic [11C]meta-hydroxyephedrine (HED) PET scans were reviewed from the PAREPET trial in ischemic cardiomyopathy patients with reduced left ventricular ejection fraction(LVEF ≤ 35%). N = 12 scans were excluded due to non-diagnostic quality. N = 178 scans were analyzed using FlowQuant and Corridor4DM software, each by two observers. Test-retest scans from N = 20 patients with stable heart failure were utilized for test-retest analysis. Denervation scores were defined as extent × severity of relative uptake defects in LV regions with < 75% of maximal uptake. Results were evaluated using intraclass correlation coefficient (ICC) and Bland-Altman coefficient of repeatability (RPC). RESULTS: Inter-observer, inter-software, and test-retest ICC values were excellent (ICC = 94% to 99%) and measurement variability was small (RPC < 11%). Mean differences between observers ranged .2% to 1.1% for Corridor4DM (P = .28), FlowQuant (P < .001), and between software programs (P < .001). Kaplan-Meier analysis demonstrated HED scores from both programs were predictive of SCA. CONCLUSION: Inter-rater reliability for both analysis programs was excellent and test-retest repeatability was consistent. The minimal difference in scores between FlowQuant and Corridor4DM supports their use in future trials.

MiR-101a loaded extracellular nanovesicles as bioactive carriers for cardiac repair.

Myocardial infarction (MI) remains a major cause of mortality worldwide. Despite significant advances in MI treatment, many who survive the acute event are at high risk of chronic cardiac morbidity. Here we developed a cell-free therapeutic that capitalizes on the antifibrotic effects of micro(mi)RNA-101a and exploits the multi-faceted regenerative activity of mesenchymal stem cell (MSC) extracellular nanovesicles (eNVs). While the majority of MSC eNVs require local delivery via intramyocardial injection to exert therapeutic efficacy, we have developed MSC eNVs that can be administered in a minimally invasive manner, all while remaining therapeutically active. When loaded with miR-101a, MSC eNVs substantially decreased infarct size (9.2 ±â€¯1.7% vs. 20.0 ±â€¯6.5%) and increased ejection fraction (53.6 ±â€¯7.6% vs. 40.3 ±â€¯6.0%) and fractional shortening (23.6 ±â€¯4.3% vs. 16.6 ±â€¯3.0%) compared to control. These findings are significant as they represent an advance in the development of minimally invasive cardio-therapies.

Quantitative proteomic and phosphoproteomic profiling of ischemic myocardial stunning in swine.

Despite decades of research on the pathophysiology of myocardial stunning, protein changes and/or phosphorylation status underlying alterations in cardiac function/structure remain inadequately understood. Here, we utilized comprehensive and quantitative proteomic and phosphoproteomic approaches to explore molecular mechanisms of myocardial stunning in swine. The closed-chest swine (n = 5 pigs) were subjected to a 10-min left anterior descending coronary artery (LAD) occlusion producing regional myocardial stunning. Tissues from the ischemic LAD region and a remote nonischemic area of the left ventricle were collected 1 h after reperfusion. Ion current-based proteomics (IonStar) and quantitative phosphoproteomics were employed in parallel to identify alterations in protein level and site-specific phosphorylation changes. A novel swine heart protein database exhibiting high accuracy and low redundancy was developed here to facilitate comprehensive study. Further informatic investigations identified potential protein-protein interactions in stunned myocardium. In total, we quantified 2,099 protein groups and 4,699 phosphorylation sites with only 0.4% missing values. Proteomic analyses revealed downregulation of contractile function and extracellular matrix remodeling. Meanwhile, alterations in phosphorylation linked with contractile dysfunction and apoptotic cell death were uncovered. NetworKIN/STRING analysis predicted regulatory kinases responsible for altered phosphosites, such as protein kinase C-mediated phosphorylation of cardiac troponin I-S199 and CaMKII-mediated phosphorylation of phospholamban-T17. In summary, the ion current-based proteomics and phosphoproteomics reliably identified novel alterations in protein content and phosphorylation contributing to contractile dysfunction, extracellular matrix (ECM) damage, and programmed cell death in stunned myocardium, which corroborate well with our physiological observations. Moreover, this work developed a comprehensive database of the swine heart proteome, a highly valuable resource for future translational research in porcine models with cardiovascular diseases.NEW & NOTEWORTHY We first used ion current-based proteomics and phosphoproteomics to reliably identify novel alterations in protein expression and phosphorylation contributing to contractile dysfunction, extracellular matrix (ECM) damage, and programmed cell death in stunned myocardium and developed a comprehensive swine heart-specific proteome database, which provides a valuable resource for future research in porcine models of cardiovascular diseases.

Transmural variation in microvascular remodeling following percutaneous revascularization of a chronic coronary stenosis in swine.

Remodeling of the coronary microcirculation is known to occur distal to a chronic coronary stenosis, but the reversibility of these changes and their functional significance on maximum myocardial perfusion before and after revascularization is unknown. Accordingly, swine instrumented with a chronic silastic stenosis on the left anterior descending coronary artery to produce hibernating myocardium underwent percutaneous coronary intervention (PCI; n = 8) and were compared with animals with a persistent stenosis (n = 8), as well as sham controls (n = 6). Stenotic animals demonstrated an increased subendocardial arteriolar wall thickness-to-lumen ratio (37.8 ± 3.3 vs. 28.3 ± 1.3% in sham, P = 0.04), reduced lumen area per arteriole (597 ± 88 vs. 927 ± 113 µm2, P = 0.04), and a compensatory increase in arteriolar density (9.4 ± 1.0 vs. 5.3 ± 0.4 arterioles/mm2, P < 0.01). As a result, vasodilated flow immediately after PCI was similar to normally perfused remote regions (5.1 ± 1.0 vs. 4.8 ± 0.9 ml·min-1·g-1, P = 0.87). When assessed 1-mo after PCI, increases in wall thickness-to-lumen diameter (42.2 ± 3.3%) and reductions in lumen area per arteriole (638 ± 59 µm2) remained unchanged, but arteriolar density returned to normal (5.2 ± 0.5 arterioles/mm2). As a result, maximum subendocardial flow during adenosine declined and was lower than remote regions (2.6 ± 0.3 vs. 5.9 ± 1.1 ml·min-1·g-1, P = 0.01). There was no microvascular remodeling in subepicardial arterioles, and maximum perfusion remained unchanged. These data demonstrate that subendocardial microvascular remodeling occurs distal to a chronic epicardial stenosis. The regression of arteriolar density without increases in luminal area may precipitate stress-induced subendocardial ischemia in the absence of a physiologically significant stenosis.NEW & NOTEWORTHY Swine with a chronic coronary stenosis exhibit subendocardial microvascular remodeling distal to a critical stenosis characterized by an increase in arteriolar wall thickness and reduction in lumen area with a compensatory increase in arteriolar density. The present study is the first to demonstrate that subendocardial arteriolar density normalizes 1-mo after revascularization, but the lumen area of individual arterioles remains reduced. This leads to a reduction in maximal subendocardial perfusion at this time point despite initial normalization of vasodilator reserve after revascularization. This pattern of chronic microvascular structural remodeling could contribute to recurrent subendocardial ischemia in the absence of coronary restenosis during tachycardia and increases in myocardial oxygen demand.

Structural and Physiological Imaging to Predict the Risk of Lethal Ventricular Arrhythmias and Sudden Death.

Identifying patients at risk of sudden cardiac death remains a major challenge in cardiovascular medicine. Advances in cardiovascular imaging have identified several anatomic and functional variables that can be quantified as continuous variables to predict the risk of developing lethal ventricular tachyarrhythmias in patients with depressed left ventricular (LV) systolic function. Some, such as LV mass, volume, and the dyssynchrony of contraction, can be derived from currently available echocardiographic and nuclear imaging modalities. Others require advanced cardiac imaging modalities with quantification of myocardial scar with gadolinium-enhanced cardiac magnetic resonance and myocardial sympathetic denervation using norepinephrine analogs and positron emission tomography or single-photon emission computed tomography offering the most promise. There is an immediate need to develop a sequential cost-effective approach that capitalizes on readily available clinical information complemented with advanced imaging modalities in selected patients to improve risk stratification for arrhythmic death beyond LV ejection fraction.

Adaptive Reductions in Left Ventricular Diastolic Compliance Protect the Heart From Stretch-Induced Stunning.

Swine subjected to 2 weeks of repetitive pressure overload (RPO) exhibited significant myocyte loss, but left ventricular (LV) systolic function was preserved, and chamber dilatation did not occur. Instead, myocardial remodeling characterized by myocyte hypertrophy and interstitial fibrosis led to a marked reduction in LV diastolic compliance, which protected the heart from stretch-induced myocyte injury and preserved LV ejection fraction without anatomic LV hypertrophy. These results support a novel paradigm that links cardiac adaptations to RPO with the pathogenesis of reduced LV diastolic compliance and may explain how LV stiffening can occur in the absence of sustained hypertension or anatomic hypertrophy.