AimAn echocardiographic algorithm derived by machine learning (e ' VM) characterizes pre-clinical individuals with different cardiac structure and function, biomarkers, and long-term risk of heart failure (HF). Our aim was the external validation of the e ' VM algorithm and to explore whether it may identify subgroups who benefit from spironolactone. Methods and resultsThe HOMAGE (Heart OMics in AGEing) trial enrolled participants at high risk of developing HF randomly assigned to spironolactone or placebo over 9 months. The e ' VM algorithm was applied to 416 participants (mean age 74 +/- 7 years, 25% women) with available echocardiographic variables (i.e. e ' mean, left ventricular end-diastolic volume and mass indexed by body surface area [LVMi]). The effects of spironolactone on changes in echocardiographic and biomarker variables were assessed across e ' VM phenotypes. A majority (>80%) had either a 'diastolic changes' (D), or 'diastolic changes with structural remodelling' (D/S) phenotype. The D/S phenotype had the highest LVMi, left atrial volume, E/e', natriuretic peptide and troponin levels (all p < 0.05). Spironolactone significantly reduced E/e' and B-type natriuretic peptide (BNP) levels in the D/S phenotype (p < 0.01), but not in other phenotypes (p > 0.10; p(interaction) <0.05 for both). These interactions were not observed when considering guideline-recommended echocardiographic structural and functional abnormalities. The magnitude of effects of spironolactone on LVMi, left atrial volume and a type I collagen marker was numerically higher in the D/S phenotype than the D phenotype but the interaction test did not reach significance. ConclusionsIn the HOMAGE trial, the e ' VM algorithm identified echocardiographic phenotypes with distinct responses to spironolactone as assessed by changes in E/e' and BNP.

BACKGROUND: Cardiac fibroblasts have crucial roles in the heart. In particular, fibroblasts differentiate into myofibroblasts in the damaged myocardium, contributing to scar formation and interstitial fibrosis. Fibrosis is associated with heart dysfunction and failure. Myofibroblasts therefore represent attractive therapeutic targets. However, the lack of myofibroblast-specific markers has precluded the development of targeted therapies. In this context, most of the noncoding genome is transcribed into long noncoding RNAs (lncRNAs). A number of lncRNAs have pivotal functions in the cardiovascular system. lncRNAs are globally more cell-specific than protein-coding genes, supporting their importance as key determinants of cell identity.METHODS: In this study, we evaluated the value of the lncRNA transcriptome in very deep single-cell RNA sequencing. We profiled the lncRNA transcriptome in cardiac nonmyocyte cells after infarction and probed heterogeneity in the fibroblast and myofibroblast populations. In addition, we searched for subpopulation-specific markers that can constitute novel targets in therapy for heart disease.RESULTS: We demonstrated that cardiac cell identity can be defined by the sole expression of lncRNAs in single-cell experiments. In this analysis, we identified lncRNAs enriched in relevant myofibroblast subpopulations. Selecting 1 candidate we named FIXER (fibrogenic LOX-locus enhancer RNA), we showed that its silencing limits fibrosis and improves heart function after infarction. Mechanitically, FIXER interacts with CBX4, an E3 SUMO protein ligase and transcription factor, guiding CBX4 to the promoter of the transcription factor RUNX1 to control its expression and, consequently, the expression of a fibrogenic gene program.. FIXER is conserved in humans, supporting its translational value.CONCLUSIONS: Our results demonstrated that lncRNA expression is sufficient to identify the various cell types composing the mammalian heart. Focusing on cardiac fibroblasts and their derivatives, we identified lncRNAs uniquely expressed in myofibroblasts. In particular, the lncRNA FIXER represents a novel therapeutic target for cardiac fibrosis.

Background: Collagen cross-linking is a fundamental process in dilated cardiomyopathy (DCM) and occurs when collagen deposition exceeds degradation, leading to impaired prognosis. This study investigated the associations of collagen-metabolism biomarkers with left ventricular function and prognosis in DCM. Methods: DCM patients who underwent endomyocardial biopsy, blood sampling, and cardiac MRI were included. The primary endpoint included death, heart failure hospitalization, or life-threatening arrhythmias, with a follow-up of 6 years (5-8). Results: In total, 209 DCM patients were included (aged 54 & PLUSMN; 13 years, 65% male). No associations were observed between collagen volume fraction, circulating carboxy-terminal propeptide of procollagen type-I (PICP), or collagen type I carboxy-terminal telopeptide [CITP] and matrix metalloproteinase [MMP]-1 ratio and cardiac function parameters. However, CITP:MMP-1 was significantly correlated with global longitudinal strain (GLS) in the total study sample (R = -0.40, p < 0.0001; lower CITP:MMP-1 ratio was associated with impaired GLS), with even stronger correlations in patients with LVEF > 40% (R = -0.70, p < 0.0001). Forty-seven (22%) patients reached the primary endpoint. Higher MMP-1 levels were associated with a worse outcome, even after adjustment for clinical and imaging predictors (1.026, 95% CI 1.002-1.051, p = 0.037), but CITP and CITP:MMP-1 were not. Combining MMP-1 and PICP improved the goodness-of-fit (LHR36.67, p = 0.004). Conclusion: The degree of myocardial cross-linking (CITP:MMP-1) is associated with myocardial longitudinal contraction, and MMP-1 is an independent predictor of outcome in DCM patients.

Introduction: Microvascular rarefaction, the functional reduction in perfused microvessels and structural reduction of microvascular density, seems to be an important mechanism in the pathophysiology of small blood vessel related disorders including vascular cognitive impairment (VCI) due to cerebral small vessel disease and heart failure with preserved ejection fraction (HFpEF). Both diseases share common risk factors including hypertension, diabetes mellitus, obesity, and ageing; in turn, these co-morbidities are associated with microvascular rarefaction. Our consortium aims to investigate novel non-invasive tools to quantify microvascular health and rarefaction in both organs, as well as surrogate biomarkers for cerebral and/or cardiac rarefaction (via sublingual capillary health, vascular density of the retina, and RNA content of circulating extracellular vesicles), and to determine whether microvascular density relates to disease severity.Methods/design: The clinical research program of CRUCIAL consists of four observational cohort studies. We aim to recruit 75 VCI patients, 60 HFpEF patients, 60 patients with severe aortic stenosis (AS) undergoing surgical aortic valve replacement as a pressure overload HFpEF model, and 200 elderly participants with mixed comorbidities to serve as controls. Data collected will include medical history, physical examination, cognitive testing, advanced brain and cardiac MRI, ECG, echocardiography, sublingual capillary health, optical coherence tomography angiography (OCTa), extracellular vesicles RNA analysis and myocardial remodelling-related serum biomarkers. The AS cohort undergoing surgery will also have myocardial biopsy for histological microvascular assessment. Discussion: CRUCIAL will examine the pathophysiological role of microvascular rarefaction in VCI and HFpEF using advanced brain and cardiac MRI techniques. Furthermore, we will investigate surrogate biomarkers for non-invasive, faster, easier, and cheaper assessment of microvascular density since these are more likely to be disseminated into widespread clinical practice. If microvascular rarefaction is an early marker of developing small vessel diseases, then measuring rarefaction may allow pre-clinical diagnosis, with implications for screening, risk stratification, and prevention. Further knowledge of the relevance of microvascular rarefaction and its underlying mechanisms may provide new avenues for research and therapeutic targets.

BACKGROUND Myocardial fibrosis may increase vulnerability to poor prognosis in patients with heart failure (HF), even in those patients exhibiting left ventricular reverse remodeling (LVRR) after guideline-based therapies.OBJECTIVES This study sought to characterize fibrosis at baseline in patients with HF with left ventricular ejection fraction (LVEF) <50% by determining serum collagen type I-derived peptides (procollagen type I C-terminal propeptide [PICP] and ratio of collagen type I C-terminal telopeptide to matrix metalloproteinase-1) and to evaluate their association with LVRR and prognosis.METHODS Peptides were determined in 1,034 patients with HF at baseline. One-year echocardiography was available in 665 patients. Associations of peptides with 1-year changes in echocardiographic variables were analyzed by multivariable linear mixed models. LVEF was considered improved if it increased by & GE;15% or to & GE;50% or if it increased by & GE;10% to >40% in patients with LVEF & LE;40%. Cardiovascular death and HF-related outcomes were analyzed in all patients ran-domized to derivation (n = 648) and validation (n = 386) cohorts.RESULTS Continuous associations with echocardiographic changes were observed only for PICP. Compared with high-PICP (& GE;108.1 ng/mL) patients, low-PICP (<108.1 ng/mL) patients exhibited enhanced LVRR and a lower risk of HF-related outcomes (P & LE; 0.018), with women and nonischemic patients with HF showing a stronger LVEF increase (interaction P & LE; 0.010). LVEF increase was associated with a better prognosis, particularly in low-PICP patients (interaction P & LE; 0.029). Only patients with both low PICP and improved LVEF exhibited a better clinical evolution than patients with nonimproved LVEF (P < 0.001).CONCLUSIONS Phenotyping with PICP, a peptide associated with myocardial fibrosis, may be useful to differentiate patients with HF who are more likely to experience clinical myocardial recovery from those with partial myocardial improvement. (J Am Coll Cardiol HF 2023;11:58-72) & COPY; 2023 by the American College of Cardiology Foundation.

Background: We sought to identify protein biomarkers of new-onset heart failure (HF) in 3 independent cohorts (HOMAGE cohort [Heart Omics and Ageing], ARIC study [Atherosclerosis Risk in Communities], and FHS [Framingham Heart Study]) and assess if and to what extent they improve HF risk prediction compared to clinical risk factors alone. Methods: A nested case-control design was used with cases (incident HF) and controls (without HF) matched on age and sex within each cohort. Plasma concentrations of 276 proteins were measured at baseline in ARIC (250 cases/250 controls), FHS (191/191), and HOMAGE cohort (562/871). Results: In single protein analysis, after adjusting for matching variables and clinical risk factors (and correcting for multiple testing), 62 proteins were associated with incident HF in ARIC, 16 in FHS, and 116 in HOMAGE cohort. Proteins associated with incident HF in all cohorts were BNP (brain natriuretic peptide), NT-proBNP (N-terminal pro-B-type natriuretic peptide), eukaryotic translation initiation factor 4E-BP1 (4E-binding protein 1), hepatocyte growth factor (HGF), Gal-9 (galectin-9), TGF-alpha (transforming growth factor alpha), THBS2 (thrombospondin-2), and U-PAR (urokinase plasminogen activator surface receptor). The increment in C-index for incident HF based on a multiprotein biomarker approach, in addition to clinical risk factors and NT-proBNP, was 11.1% (7.5%-14.7%) in ARIC, 5.9% (2.6%-9.2%) in FHS, and 7.5% (5.4%-9.5%) in HOMAGE cohort, all P<0.001), each of which was a larger increase than that for NT-proBNP on top of clinical risk factors. Complex network analysis revealed a number of overrepresented pathways related to inflammation (eg, tumor necrosis factor and interleukin) and remodeling (eg, extracellular matrix and apoptosis). Conclusions: A multiprotein biomarker approach improves prediction of incident HF when added to natriuretic peptides and clinical risk factors.

Background: Spironolactone might improve the prognosis of patients with heart failure with preserved left ventricular ejection fraction (HFpEF), but the mechanisms by which it acts are uncertain. Serum concentrations of procollagen type I carboxy-terminal propeptide (PICP) reflect the synthesis of type I collagen and correlate well with histologically proven cardiac fibrosis.Aims: To investigate the effect of spironolactone on serum PICP concentration in patients with stage B and C HFpEF across three trials (HOMAGE, ALDO-DHF, and TOPCAT) for which measurements of serum PICP were available.Methods: Random-effects meta-analysis.Results: A total of 1038 patients with PICP measurements available both at baseline and 9-12 months were included in this analysis: 488 (47.0%) from HOMAGE, 386 (37.2%) from ALDO-DHF, and 164 (15.8%) from TOPCAT. The median (percentile25-75) serum PICP was 98 (76-128) ng/mL. Compared to placebo or usual care, administration of spironolactone for 9 to 12 months reduced serum PICP by-7.4 ng/mL, 95%CI-13.9 to-0.9, P -value =0.02. The effect was moderately heterogeneous (I2 = 64%) with the most pronounced effect seen in TOPCAT where PICP was reduced by-27.0 ng/mL, followed by HOMAGE where PICP was reduced by-8.1 ng/ mL, and was least marked in ALDO-DHF where PICP changed by-2.9 ng/mL. The association between spi-ronolactone and serum PICP was not mediated substantially by blood pressure.Conclusions: Spironolactone reduced serum concentrations of PICP in patients with HFpEF with different severity and stages of disease. These findings are consistent with spironolactone having an anti-fibrotic effect.

Simple Summary Left ventricular dysfunction (LVD) induced by anthracycline-based cancer chemotherapy (ACC) is becoming an urgent healthcare concern. Myocardial fibrosis (MF) may contribute to LVD after ACC. We show that elevated circulating levels of procollagen type I C-terminal propeptide (PICP, biomarker of MF) are associated with early subclinical LVD and predict later development of cardiotoxicity in patients treated with ACC. In addition, an association between PICP and LVD in patients with ACC-induced heart failure is observed. These results provide novel insights into MF as a mechanism underlying LVD after ACC, with PICP emerging as a promising tool to monitor cardiotoxicity in patients treated with ACC. Anthracycline-based cancer chemotherapy (ACC) causes myocardial fibrosis, a lesion contributing to left ventricular dysfunction (LVD). We investigated whether the procollagen-derived type-I C-terminal-propeptide (PICP): (1) associates with subclinical LVD (sLVD) at 3-months after ACC (3m-post-ACC); (2) predicts cardiotoxicity 1-year after ACC (12m-post-ACC) in breast cancer patients (BC-patients); and (3) associates with LVD in ACC-induced heart failure patients (ACC-HF-patients). Echocardiography, serum PICP and biomarkers of cardiomyocyte damage were assessed in two independent cohorts of BC-patients: CUN (n = 87) at baseline, post-ACC, and 3m and 12m (n = 65)-post-ACC; and HULAFE (n = 70) at baseline, 3m and 12m-post-ACC. Thirty-seven ACC-HF-patients were also studied. Global longitudinal strain (GLS)-based sLVD (3m-post-ACC) and LV ejection fraction (LVEF)-based cardiotoxicity (12m-post-ACC) were defined according to guidelines. BC-patients: all biomarkers increased at 3m-post-ACC versus baseline. PICP was particularly increased in patients with sLVD (interaction-p < 0.001) and was associated with GLS (p < 0.001). PICP increase at 3m-post-ACC predicted cardiotoxicity at 12m-post-ACC (odds-ratio >= 2.95 per doubling PICP, p <= 0.025) in both BC-cohorts, adding prognostic value to the early assessment of GLS and LVEF. ACC-HF-patients: PICP was inversely associated with LVEF (p = 0.004). In ACC-treated BC-patients, an early increase in PICP is associated with early sLVD and predicts cardiotoxicity 1 year after ACC. PICP is also associated with LVD in ACC-HF-patients.

Heart failure is a major cause of morbidity and mortality worldwide, and can result from pressure overload, where cardiac remodelling is characterized by cardiomyocyte hypertrophy and death, fibrosis, and inflammation. In failing hearts, transforming growth factor (TGF)beta drives cardiac fibroblast (CFB) to myofibroblast differentiation causing excessive extracellular matrix production and cardiac remodelling. New strategies to target pathological TGF beta signalling in heart failure are needed. Here we show that the secreted glycoprotein ADAMTSL3 regulates TGF beta in the heart. We found that Adamtsl3 knock-out mice develop exacerbated cardiac dysfunction and dilatation with increased mortality, and hearts show increased TGF beta activity and CFB activation after pressure overload by aortic banding. Further, ADAMTSL3 overexpression in cultured CFBs inhibits TGF beta signalling, myofibroblast differentiation and collagen synthesis, suggesting a cardioprotective role for ADAMTSL3 by regulating TGF beta activity and CFB phenotype. These results warrant future investigation of the potential beneficial effects of ADAMTSL3 in heart failure.

Aims: The HOMAGE randomized trial found that spironolactone reduced left atrial volume index (LAVI), E:A ratio, and a marker of collagen type I synthesis (procollagen type I C-terminal propeptide) in patients at risk of heart failure (HF). Previous trials showed that patients with HF, preserved ejection fraction and low serum collagen type I C-terminal telopeptide to matrix metalloproteinase-1 ratio (CITP:MMP-1), associated with high collagen cross-linking, had less improvement in diastolic function with spironolactone. We evaluated the interaction between serum CITP:MMP-1 and spironolactone on cardiac function in the HOMAGE trial. Methods and results: Patients at risk of HF were randomized to spironolactone (n = 260) or not (n = 255). Blood sampling and echocardiography were done at baseline, one and nine months. CITP:MMP-1 was used as an indirect measure of collagen cross-linking. Higher baseline CITP:MMP-1 (i.e. lower collagen cross-linking) was associated with greater reductions in LAVI with spironolactone at both one (p = 0.003) and nine (p = 0.01) months, but no interaction was observed for E:A ratio. Spironolactone reduced LAVI after one and nine months only for those patients in the third tertile of CITP:MMP-1 (estimated lowest collagen cross-linking) [mean differencesspiro/control : -1.77 (95% confidence interval, CI -2.94 to -0.59) and -2.52 (95% CI -4.46 to -0.58) mL/m2 ; interaction pacross-tertiles = 0.005; interaction pthird tertile = 0.008] with a similar trend for N-terminal pro-B-type natriuretic peptide which was consistently reduced by spironolactone only in the lowest collagen cross-linking tertile [mean differencesspiro/control : -0.47 (95% CI -0.66 to -0.28) and -0.31 (95% CI -0.59 to -0.04) ng/L; interaction pacross-tertiles = 0.09; interaction pthird tertile < 0.001]. Conclusions: These findings suggest that, for patients at risk of HF, the effects of spironolactone on left atrial remodelling may be more prominent in patients with less collagen cross-linking (indirectly assessed by serum CITP:MMP-1).