Circulation: Genomic and Precision Medicine

BackgroundPediatric dilated cardiomyopathy (DCM) is a rare, progressive heart disease with variable outcomes that range from recovery to heart transplantation. To date, there are no prognostic biomarkers for children with DCM. Identifying circulating biomarkers that are associated with clinical outcomes is critical for personalized management. MethodsmiRNAs were identified by RNA-seq, whereas proteins were identified by SomaScan(R). Machine learning methodologies were used to explore the predictive ability of circulating factors identified from serum samples collected at the time of presentation with acute heart failure. ResultsThirty patients experienced poor outcomes (cardiac transplantation, mechanical circulatory support, or death) and 19 patients recovered left ventricular function. Distinct miRNA and protein signatures differentiated outcomes groups. Top candidate proteins (COL2A1, CXCL12, and ADGRF5) and miRNAs (miR-874-3p, miR-335-3p, miR-323a-3p) demonstrated strong discriminatory performance within the study cohort (recovered vs poor outcomes; Area Under the Curve of 0.92). Ingenuity Pathway Analysis implicates cardiac remodeling, fibrosis, and inflammatory signaling as central pathways differentiating patient outcomes. ConclusionsCirculating miRNA and protein signatures at presentation identify a circulating molecular signature associated with divergent clinical trajectories in pediatric DCM. These findings support the potential utility of multi-omic biomarkers for early risk stratification and provide insight into mechanisms underlying divergent outcomes. CLINICAL PERSPECTIVEWhat Is New? O_LICirculating miRNA and protein profiles measured at presentation distinguish children with pediatric DCM who recover from those who progress to advanced heart failure. C_LIO_LIA combined multi-omic biomarker demonstrated strong discriminatory performance in this cohort (AUC 0.92). C_LIO_LIPathway analysis implicates extracellular matrix remodeling, fibrosis, and inflammatory signaling in children with adverse clinical trajectories. C_LI What Are the Clinical Implications? O_LISerum-based molecular biomarkers may enable earlier risk stratification in children presenting with dilated cardiomyopathy. C_LIO_LIMulti-omic integration may improve identification of pediatric patients at risk for transplantation, mechanical circulatory support, or death. C_LIO_LIThese findings support further validation of circulating biomarker panels to guide personalized management in this rare disease. C_LI RESEARCH PERSPECTIVEWhat New Question Does This Study Raise? O_LICan integrated circulating miRNA-protein signatures identify biologically distinct trajectories of recovery versus progression in children with dilated cardiomyopathy? C_LIO_LIDo circulating molecular profiles reflect underlying disease mechanisms that determine divergent clinical outcomes in pediatric DCM? C_LI What Question Should Be Addressed Next? O_LIDo the pathways identified by integrated miRNA-protein analysis (fibrosis, remodeling, and inflammation) play causal roles in determining recovery versus progression? C_LIO_LICan multi-omic biomarkers be incorporated into prospective studies to improve early risk stratification and guide clinical management? C_LI

Background: Andersen-Tawil syndrome type 1 (ATS1) is caused by loss-of-function mutations in KCNJ2, which encodes the inward rectifier K+ channel Kir2.1, a key determinant of IK1. Impaired Kir2.1 destabilizes membrane excitability and predisposes to ventricular arrhythmias. Most ATS1 variants disrupt channel regulation by phosphatidylinositol 4,5-bisphosphate (PIP2), but whether specific mutations confer differential arrhythmic risk remains unclear. Objective: To determine whether ATS1 variants disrupting Kir2.1-PIP2 interactions define distinct arrhythmic risk profiles and establish a mechanistically informed framework for risk stratification. Methods: We performed a pooled patient-level analysis of 225 ATS1 patients carrying KCNJ2 variants impairing Kir2.1-PIP2 interaction. Inclusion of 22 clinical and electrocardiographic variables were used to identify mutation-specific risk profiles and predictors for arrhythmia risk. The approach was validated in a multicenter cohort of 20 ATS1 patients. Functional validation was performed using patient-derived iPSC-CMs, cardiac-targeted mouse models, and structural in silico analyses. Results: ATS1 variants segregated into three discrete clusters corresponding to high-, intermediate-, and low-risk arrhythmic phenotypes, establishing a mutation-dependent hierarchy of arrhythmic risk. Regression analyses identified six variables independently associated with severe arrhythmic outcomes. Patient-derived iPSC-CM demonstrated graded impairment of electrical propagation and arrhythmia susceptibility, with a hierarchy in conduction velocity, CV:Control > R82W > R218W > G215D). Cardiac-targeted ATS1 mouse models reproduced the clinical risk stratification. Structural modeling showed that high-risk variants localize near the channel pore and disrupt Kir2.1-PIP2 interactions through mutation-specific mechanisms. Conclusions: ATS1 caused by Kir2.1-PIP2-disrupting variants is not a uniform disorder but comprises biologically distinct subgroups with predictable differences in arrhythmic severity. Integrating genetics, functional phenotyping, and structural modeling provides a mechanistically grounded framework for ATS1 risk stratification and precision therapy development.

BackgroundMetabolic dysfunction-associated steatohepatitis (MASH) is emerging as a risk factor of cardiometabolic diseases, including the atrial fibrillation (AF) - the most common sustained arrhythmia. Given that the liver is a major source of inflammatory mediators, lipids, and hepatokines under metabolic stress, we hypothesized that hepatocyte-derived factors in MASH may accelerate atrial remodeling and arrhythmogenesis. MethodsAnalysis of the Atherosclerosis Risk in Communities (ARIC) visit 5 cohort was performed to determine the association between the FIB-4 index - a classic indicator of liver fibrosis, and AF risk, with multivariable adjustment for common comorbidities. A murine model of MASH was induced using the GAN (Gubra-Amylin NASH) diet. Programmed intracardiac stimulation and echocardiography were performed to assess AF susceptibility and cardiac function. Calcium imaging, histology, flow cytometry, plasma proteomics, and single-nucleus RNA sequencing (snRNA-seq) analyses were employed to elucidate the role of recruited inflammatory macrophages via hepatocyte-derived osteopontin (OPN) in MASH-induced atrial remodeling. ResultsAnalysis of the ARIC cohort confirmed a higher cumulative incidence of AF and an elevated adjusted hazard ratio (HR) in patients with intermediate and high FIB-4 indices compared to individuals with low FIB-4 scores. MASH mice exhibited increased susceptibility to pacing-induced AF, accompanied by enhanced proarrhythmic calcium release events, atrial enlargement, and fibrosis, independent of ventricular dysfunction. Proteomics and snRNA-seq revealed that the hepatocyte-secreted OPN under MASH conditions promoted the differentiation and recruitment of TGFBR1+ inflammatory macrophages to the atria, leading to gasdermin D (GSDMD) activation - an effector of inflammasome signaling and consequent proarrhythmic atrial remodeling. Activation of the monocyte-derived pro-inflammatory TGFBR1+ macrophages was dependent on the OPN receptor CD44. Furthermore, the MASH-induced atrial fibroinflammatory milieu and enhanced AF susceptibility were mitigated through several strategies, including hepatocyte-specific Spp1 (encoding OPN) deletion, neutralization of circulating OPN, ablation of CD44 or GSDMD. ConclusionsThese findings establish a pathogenic role of the hepatokine osteopontin in driving activation and recruitment of TGFBR1+ inflammatory macrophages into the atria, leading to proarrhythmic atrial remodeling under MASH. Osteopontin-targeted therapy or GSDMD inhibition prevents AF, indicating a novel therapeutic strategy for liver disease-related atrial arrhythmogenesis. Clinical PerspectiveO_ST_ABSWhat is new?C_ST_ABSO_LIIn the ARIC cohort, metabolic dysfunction-associated steatohepatitis (MASH) is associated with increased risk of atrial fibrillation (AF) after adjusting for common comorbidities. Elevated levels of circulating osteopontin (encoded by SPP1) predict an increased risk of AF in patients with MASH-induced liver fibrosis. C_LIO_LIMASH enhances hepatocyte secretion of osteopontin, leading to expansion of myeloid cells and recruitment of inflammatory macrophages into atria. This liver-to-atrial inflammatory circuit promotes the development of a substrate conducive to AF, which can be attenuated by hepatocyte-specific Spp1 deletion or neutralizing anti-anti-osteopontin antibody treatment to eliminate the mediator, or ablation of inflammasome effector gasdermin D to correct the atrial response. C_LI What are the clinical implications?O_LIOsteopontin may serve as a biomarker for AF in MASH cohorts. C_LIO_LIAnti-osteopontin therapy through neutralizing antibodies may serve as a novel therapeutic strategy for liver disease-related atrial arrhythmia. C_LI

Background: Non-ischemic cardiomyopathy (NICM) represents a major cause of heart failure with limited tools for early risk stratification. Atrial fibrillation (AF) is a well-established contributor to cardiomyopathy but is often clinically silent in its early stages. The atrial fibrillation polygenic risk score (AF-PRS) reflects genetic susceptibility to AF and may identify individuals at risk for AF-related cardiomyopathy. We hypothesized that higher AF-PRS is associated with greater risk of NICM. Methods: This was a retrospective cohort study of 16,801 individuals of European ancestry from the Sanford Biobank and Imagenetics program with genetic sequencing and longitudinal electronic health record data. AF-PRS was calculated using 315 genome-wide significant single-nucleotide polymorphisms with standard quality control. NICM was defined by International Classification of Diseases, 10th Revision, Clinical Modification codes, excluding ischemic etiologies. Cox regression models evaluated the association between AF-PRS and incident NICM, adjusting for age, sex, smoking status, body mass index (BMI), hypertension, and diabetes. AF-PRS was analyzed both as a quasi-continuous variable (15% quartile increments) and dichotomized at the 85th percentile. Sensitivity analyses assessed associations with all-cause cardiomyopathy and ischemic cardiomyopathy. Survival analysis was used to model time-to-event outcomes. Results: Among all participants, 418 (2.5%) had NICM. 99% were Caucasian. NICM cases were older and more often male (both p<0.001) than those without a diagnosis. After multivariable adjustment for sex, smoking status, BMI, and hypertension, a linear AF-PRS (15% increments) was specifically predictive of increased hazard risk of NICM (HR = 1.09 [1.03, 1.15], p < 0.001). Conclusion: These findings complement recent evidence of bidirectional genetic relationships between cardiomyopathy and AF, supporting comprehensive genetic risk assessment in cardiovascular disease prevention. Clinical implementation requires validation in diverse populations and prospective evaluation. Future research should investigate the mechanistic pathways linking AF-associated genetic variants to cardiomyopathy development and evaluate whether AF-PRS-guided screening improves clinical outcomes.

Background. Hypertrophic cardiomyopathy (HCM) is a clinically variable disease in terms of onset and progression. Pathogenic MYBPC3 variants account for a substantial proportion of HCM diagnoses. This study sought to identify protein biomarkers associated with HCM severity. Methods. Olink-assayed plasma proteins of 144 MYBPC3 pathogenic variant carriers were tested for associations with HCM severity based on HCM diagnostic criteria (unaffected, mildly, or severely affected). The UK Biobank was used to replicate the identified proteins through considering time to onset of HCM (67 cases), cardiomyopathy (156 cases),and associations with cardiac MRI derived left ventricular maximum wall thickness (6,492 participants). Replicated proteins were further prioritised based on cardiac tissue expression and druggability, and annotated using pathway enrichment and association with onset of: heart failure (HF), dilated cardiomyopathy (DCM), sudden cardiac arrest (SCA), and ventricular arrhythmias (VA). Results. Among pathogenic MYBPC3 variant carriers, we identified 27 proteins associated with HCM severity. We independently replicated 21 proteins in the UK Biobank. Of the five prioritised proteins (NT-proBNP, GDF-15, FGF-23, ADM, and NCAM1), all but NT-proBNP were targeted by drugs with repurposing potential. The replicated proteins additionally associated with the incidence of HF (n=5), DCM (n=4), SCA (n=4), and VA (n=4). Conclusion. This study replicated 21 and prioritised five proteins associated with HCM severity in pathogenic MYBPC3 variant carriers. Replication in unselected HCM suggests the prioritised proteins are associated with HCM independent of genotype, providing important leads for plasma-based markers for diagnoses, disease monitoring, and drug targets.

Background and aimsCurrent FH VCEP specifications of ACMG/AMP guidelines for familial hypercholesterolemia (FH) variant interpretation assign a higher evidence weight to functional data obtained with flow cytometry than microscopy assays, due to lack of existing evidence. This restricts the use of microscopy-derived functional data for variant classification. We aimed to systematically compare functional data of LDLR variants obtained by high-content microscopy and flow cytometry to determine their concordance and assess whether microscopy-based assays could support a higher evidence level. MethodsFifty LDLR variants with available flow cytometry and high-content microscopy data were compared for LDL uptake activity, including 21 newly characterized variants by microscopy in this study. Variants were grouped by FH VCEP functional thresholds (<70% activity, abnormal function; >90% activity, normal function) and results were integrated with UK Biobank data to assess associations with lipid traits. ResultsFirst, we validated our scalable microscopy assay with FH VCEP-classified control variants. Then we compared functional activity measured by microscopy and flow cytometry assays for 50 variants, which showed significant correlation (r = 0.66, p<0.0001) and a close average agreement (Bland-Altman bias = -0.05). Applying FH VCEP functional classification thresholds yielded broadly consistent classification in both methods, with minor shifts among categories. Integration with UK Biobank data showed that carriers of variants with reduced LDLR activity (<70% and <50%) had higher LDL-C, total cholesterol and ApoB levels compared to those with normal activity (>90%) for both microscopy and flow cytometry assays, with more pronounced differences observed at the <50% LDLR activity threshold. ConclusionHigh-content microscopy provides reliable and scalable measurements of LDLR function, showing high concordance with flow cytometry and consistent associations with lipid phenotypes. These findings support reconsideration of the evidence weight assigned to validated microscopy assays within FH VCEP variant classification frameworks, namely to Strong (Level 1).

BackgroundSpontaneous coronary artery dissection (SCAD) is a well-recognised cause of acute coronary syndrome particularly among women without conventional cardiovascular risk factors. Increasing evidence indicates a genetic contribution; however, the underlying genetic architecture of SCAD remains insufficiently understood. ObjectiveThe aim of this study was to assess the prevalence of rare variants in previously reported SCAD associated genes and to explore the potential presence of novel genetic alterations in well-characterised Swedish patients with SCAD. MethodsThe study comprised 201 patients enrolled in SweSCAD, a national project examining the clinical characteristics, aetiology, and outcomes of SCAD. All individuals had a confirmed diagnosis based on invasive coronary angiography. Comprehensive exome sequencing was performed to identify rare variants contributing to disease susceptibility. ResultsGenetic variants that have been associated with SCAD according to current clinical genetics practice for variant reporting were identified in approximately 4 % of patients. In addition, rare potentially relevant variants were detected in almost 60 % of patients in genes associated with vascular integrity and vascular remodelling. ConclusionThis study supports SCAD as a genetically complex arteriopathy, driven by rare high-impact variants together with broader polygenic susceptibility. Variants in collagen, vascular extracellular matrix, and oestrogen-responsive pathways provide biologically plausible links to female-predominant disease. Although the diagnostic yield of clearly actionable variants is modest, these findings support broader genomic evaluation beyond overt syndromic presentations and highlight the need for larger integrative genomic and functional studies to refine risk stratification and management.

Background and aimsSpontaneous coronary artery dissection (SCAD) is a non-atherosclerotic cause of acute myocardial infarction (MI) that predominantly affects young women. As an under-recognized cause of MI, large genome-wide association studies (GWAS) remain challenging. We aimed to leverage SCAD shared genetic basis with related vascular diseases to uncover genetically determined biological mechanisms. MethodsSummary statistics for SCAD GWAS (1,917 cases, 9,293 controls) was harmonised with seven related vascular traits: fibromuscular dysplasia, intracranial aneurysm, cervical artery dissection, migraine, coronary artery disease, abdominal aortic aneurysm, and thoracic aortic aneurysm/dissection. We applied Multi-Trait Analysis of GWAS (MTAG). We integrated coronary-artery regulatory annotations, cis-eQTL mapping, and colocalization to prioritize candidate genes. Gene-based testing (LDAK-GBAT) was applied to SCAD dataset. ResultsMTAG identified 40 independent SCAD loci, including 24 that were novel. Candidate variants were enriched in open chromatin from coronary smooth muscle cells and fibroblasts and in vascular regulatory regions. LDAK-GBAT identified 46 significant genes, including 12 outside MTAG loci. Integrated functional annotation prioritized 56 genes linked to arterial integrity, vasoactive tone, haemostasis, and coagulation. Extracellular matrix organization was confirmed as a key pathway, with additional enrichment in bone mineralization and TGF-{beta} related terms. ConclusionsIntegrating multi-trait GWAS, gene-based testing, epigenetic and transcriptomic data substantially expanded the SCAD genetic landscape. Our findings implicate key arterial-wall pathways beyond extracellular matrix organization, and point at relevant biological mechanisms in non-atherosclerotic dissection. These findings nominate tractable targets for experimental follow-up and support future efforts toward SCAD risk stratification in women.

Heart failure with preserved ejection fraction (HFpEF) is a heterogeneous cardiometabolic syndrome in which the molecular programs linking metabolic stress to myocardial remodeling and diastolic dysfunction remain incompletely defined. We integrated ventricular RNA sequencing with pathway activity profiling, transcription factor inference, cell-type enrichment, phenotype association, elastic-net severity modeling, cross-lab murine validation, and human proteomic comparison to define the systems-level architecture of remodeling in the db/db + aldosterone mouse model of cardiometabolic HFpEF. HFpEF hearts exhibited a distinct transcriptomic state characterized by coordinated upregulation of collagen organization, TGF{beta} signaling, inflammatory response, and NF{kappa}B signaling, with reduced ion-channel activity and smaller shifts in oxidative phosphorylation, excitation-contraction coupling, and mechanotransduction. These pathway programs were linked to left ventricular hypertrophy and diastolic dysfunction and were accompanied by enrichment of fibroblast, myofibroblast, and macrophage signatures that tracked the same disease dimensions. Gene-level prioritization identified extracellular matrix, inflammatory, and mechanotransduction-associated candidates linked to disease severity, while transcription factor analysis revealed a broader multi-regulator architecture associated with fibrotic, inflammatory, and stress-responsive remodeling. Elastic-net modeling further showed that phenotype-derived remodeling severity was captured in an exploratory nested cross-validation framework primarily by transcription factor and fibro-inflammatory cell-program features, whereas pathway-summary scores added little incremental predictive information. In an independent HFD+L-NAME cohort, pathway remodeling showed selective reproducibility, and cross-species comparison demonstrated that concordance with human HFpEF proteomic subgroups was pathway selective rather than global. Together, these findings define a multilevel systems architecture of cardiometabolic HFpEF remodeling and support mechanistic prioritization and pathway-matched preclinical model selection.

BackgroundHypertrophic cardiomyopathy (HCM) is a heritable trait with marked variability in expression and outcomes. Our aims were to discover new genetic loci associated with HCM and to test the effect of a new polygenic risk score (PRS) on incidence, phenotype and outcomes, stratified by sarcomere genotype status. MethodsA discovery genome-wide association study (GWAS) was performed on 2,284 HCM cases and 4,525 controls. Two fixed-effects meta-analyses combined our discovery GWAS with single-trait and multi-trait results from a published study. Discovered loci underwent comprehensive bioinformatic analysis including functional and druggability annotations. A PRS using loci from the two meta-analyses was evaluated for association with: HCM diagnosis in 411,213 individuals from UK Biobank (UKBB); imaging phenotypes in individuals without HCM; a composite endpoint (including all-cause mortality and transplantation) and sudden cardiac death (SCD) in 1,756 HCM cases. PRS analyses were stratified by sarcomere genotype status. ResultsThree loci were found in the discovery GWAS (BAG3, FHOD3 and novel locus PPP1R3A). In the meta-analyses, 70 unique loci were identified, four novel (MYPN, YWHAE, NOS1AP and OBSCN). Bioinformatic analyses identified NOS1AP as a candidate HCM gene. A new PRS was significantly associated with HCM diagnosis (hazard ratio [HR] 3.19, 95% CI:2.46-4.14, for top 5% vs lower 95%; HR 1.88, 95% CI:1.72-2.06, per SD increase). Significant associations were found between PRS and greater left ventricular (LV) wall thickness and higher LV ejection fraction in UKBB participants without HCM. Sarcomere-negative HCM cases in the top 20% of the PRS distribution had an increased risk of SCD (HR 2.72, 95% CI:1.03-7.17). ConclusionsWe report novel HCM loci. A new PRS predicted the risk of HCM development and associated imaging characteristics in the UKBB and outcomes in an HCM cohort.

Background and AimsDespite treatment, patients with established atherosclerotic cardiovascular disease (ASCVD) are at high risk of recurrent events. Existing clinical risk scores for recurrence provide only moderate predictive performance and rely largely on the same conventional risk factors used to predict disease onset. Proteomics is a promising source of new biomarkers but the technologies need focused use cases in order to achieve utility and implementation. We aimed to determine whether plasma proteomics improves prediction of recurrent cardiovascular events beyond established clinical risk models in secondary prevention in a population-scale cohort. MethodsPlasma proteomic profiles from [~]9,300 participants in the UK Biobank with established ASCVD at baseline were analysed using machine learning methods to derive and evaluate proteomic predictors of recurrent cardiovascular events. The top performing model comprised proteins with non-zero weights (full protein score). Predictive performance of the proteomic predictors, an established clinical risk score (SMART2), and their combination was evaluated across six pre-defined testing datasets representing multiple ethnic and geographic groups. A parsimonious set of proteins with existing clinical-grade enzyme-linked immunosorbent assays (ELISAs) available was then derived. ResultsThe full protein score achieved higher performance for recurrent ASCVD than the SMART2 risk score across all ethnic and geographic subgroups (mean C-index 0.743 vs 0.653). Adding the full protein score to SMART2 improved discrimination, with the largest increase in White Irish participants ({Delta}C-index, 0.140; 95% CI, 0.074-0.205; P<0.001). However, adding SMART2 to the protein score provided minimal additional value. The parsimonious score preserved most of the discrimination of the full protein model with C-indices of the recurrent ASCVD risk model comprising age, sex and the parsimonious protein score being nearly identical to the full protein model in the largest testing set (0.723 vs 0.728 for White British in England and Wales). The parsimonious protein score showed a marked gradient of risk with the top, middle and bottom quintiles showing 10-year recurrent ASCVD rates of [~]27.4%, [~]9.6% and [~]2.4%, respectively. ConclusionsIn patients with established ASCVD, plasma protein measurements substantially improved prediction of recurrent events beyond conventional clinical risk factors, supporting their potential as a complementary tool to guide secondary prevention of cardiovascular disease.

Background: Familial hypercholesterolemia (FH) is most frequently caused by pathogenic variants in LDLR, but phenotypic variability suggests the influence of genetic modifiers. Methods: We investigated a large multigenerational family with FH, combining clinical data, lipid profiles, and genetic analysis with functional studies. LDLR and PCSK9 variants were characterized according to ACMG/ClinGen guidelines. Functional assays in CHO-ldlA7 cells assessed LDLR activity, while plasma PCSK9 levels were quantified by ELISA. Results: LDLR c.2479G>A variant was associated with FH in the family. The presence of loss of function c.137G>T and c.2023del variants at PCSK9 appears to mitigate the effect of the LDLR variant. Conclusions: This study provides evidence that PCSK9 variants can counteract the deletereous effect of a LDLR variant associated with FH. These findings highlight the importance of gene/gene interactions in the clinical variability of FH and their potential implications for precision medicine.

Rationale: Heart failure with preserved ejection fraction (HFpEF) is a heterogeneous syndrome with substantial unmet diagnostic and therapeutic needs. Circulating lipid metabolism is increasingly implicated in HFpEF pathophysiology but has not been systematically leveraged for molecular stratification. Objective: To determine whether plasma lipidomics can identify molecular phenogroups of HFpEF associated with distinct clinical characteristics and outcomes. Methods and Results: Untargeted plasma lipidomics was performed in non-HF subjects and HFpEF patients from a primary Belgian cohort and an independent Canadian cohort (n=177 in each cohort). In the Belgian cohort, 235 unique lipids spanning 19 subclasses were annotated, including 96 significantly associated with HFpEF (q<0.02). Unsupervised analyses revealed marked lipidomic heterogeneity, with a distinct HFpEF subgroup separable from non-HF subjects. Hierarchical clustering identified three phenogroups with divergent lipid profiles and clinical features. One phenogroup exhibited severe atrial dysfunction, congestion-related biomarkers, elevated indices of cardiac and liver fibrosis, and markedly reduced survival, a second was characterized by prominent metabolic syndrome features, and a third by preserved renal function. Cross-cohort comparison using a supervised classifier trained on 158 shared lipids confirmed analogous lower-risk phenogroups in the Canadian cohort, while the high-risk phenotype was underrepresented. A signature of 10 lipids across six subclasses, including long-chain acylcarnitines, ether phosphatidylcholines, and oxidized sphingomyelins, discriminated the high-risk group and correlated with markers of disease severity. Conclusion: Our findings demonstrate that HFpEF comprises metabolically distinct patient subgroups across cohorts, revealing specific lipidomic dysfunctions that deepen our understanding of HFpEF heterogeneity and underlying pathophysiology.

Background: Coronary artery calcium (CAC) scores inform subclinical atherosclerotic cardiovascular disease (ASCVD) burden, helping guide preventative treatments. However, prediction of cardiovascular (CV) events by CAC is largely limited to ASCVD outcomes. This study investigated whether a previously validated proteomic test for predicting a broad composite of four-year CV events could enhance the prognostic utility of CAC. Methods: We used a 27-protein CV risk score (Prot-CVR), derived from ~5,000 SomaScan? Assay plasma protein measurements, to predict four-year risk of a composite CV and mortality outcome (myocardial infarction, stroke/TIA, heart failure hospitalization, death) in 2,122 participants with ?1 CV risk factors from the Multi-Ethnic Study of Atherosclerosis (MESA) observational cohort at exam 5 and compared predictions to CAC Agatston scores. Discriminatory performance was assessed using C-Index and 4-year area under the curve (AUC). Cox Proportional Hazard (CoxPH) ratios were calculated for the composite outcome, ASCVD outcome (myocardial infarction, resuscitated cardiac arrest, stroke, coronary heart disease death), and individual events. Changes in Prot-CVR and CAC scores from baseline to MESA exam 5 (+10-years) in CV event versus event-free participants were assessed using 2-tailed paired t-tests. CoxPH regression models of CV event status distributed by Prot-CVR, CAC, and relevant co-variates were evaluated for performance relative to individual models. Results: Individual Prot-CVR and CAC models predicting the composite outcome had comparable 4-year AUCs, but Prot-CVR had a higher C-index (0.68 (0.65-0.70) versus 0.63 (0.60-0.65), p=0.001) and greater hazard ratios for the composite outcome (p<0.001), death (p<0.001), and heart failure (p=0.015). A combined CoxPH model of Prot-CVR + CAC + Age had a higher 4-year AUC (0.72, p<0.05) and C-Index (0.71, p<0.05) than Prot-CVR or CAC alone. Both Prot-CVR and CAC scores detected an increase in risk prior to an approaching CV event in ~10-year sensitivity-to-change analysis. For 49.6% of MESA population with CAC=0 at baseline, Prot-CVR was greater in composite event versus event free participants at 4 years (0.23 versus 0.15, p=0.006) and full follow-up (0.18 versus 0.13, p<0.001). Conclusion: Protein testing complements CAC for CV risk assessment although the improvement is modest. Prot-CVR may resolve which patients with CAC=0 are at heightened CV risk.

Background: The clinical significance of sarcomere variants of uncertain significance (VUS) in hypertrophic cardiomyopathy (HCM) remains unclear, and VUS are currently regarded as clinically non-actionable despite their increasing prevalence. This study aimed to evaluate genotype?phenotype and genotype?outcome associations according to variant pathogenicity in patients with HCM, with a particular focus on the clinical relevance of sarcomere VUS. Methods: This multicenter retrospective cohort study included 438 patients with HCM who underwent next-generation sequencing-based genetic testing at two tertiary hospitals. Patients were classified into three groups: pathogenic or likely pathogenic (P/LP) variants, VUS, and no sarcomere mutations. Clinical characteristics, imaging phenotypes, and outcomes were compared across groups. The primary endpoint was a composite of cardiovascular death, aborted sudden cardiac death, appropriate implantable cardioverter-defibrillator therapy, and heart transplantation. Time-to-event analyses were performed using Kaplan-Meier methods and Cox proportional hazards models with Firth's penalized partial likelihood approach. Results: P/LP variants were identified in 171 patients (39.0%) and sarcomere VUS in 159 patients (36.3%). Patients with VUS demonstrated intermediate clinical and phenotypic features between P/LP carriers and genotype-negative patients. Kaplan?Meier analysis showed a graded difference in event-free survival across variant classifications. While VUS were not independently associated with adverse outcomes when modeled as a categorical variable, increasing pathogenicity from genotype-negative to VUS and P/LP variants was associated with a stepwise increase in risk of the primary endpoint (hazard ratio 2.05, 95% confidence interval 1.11?4.16 p=0.019). Identified VUS were preferentially enriched in Z-disc and giant sarcomere scaffolding proteins. Conclusion: Sarcomere VUS represent intermediate characteristics along a continuum of sarcomere dysfunction, associated with distinct phenotypic features and clinical outcomes compared with both P/LP variants and the absence of sarcomere mutations. These findings suggest that sarcomere VUS may not be entirely clinically neutral and should be interpreted within a broader genetic and structural context in patients with HCM.

Background: Insulin resistance (IR) and obesity are key drivers of atrial fibrillation (AF). However, the comparative predictive value of the Triglyceride-Glucose (TyG) index versus composite indices combining IR and anthropometric measures such as TyG-BMI, TyG-Waist Circumference (TyG-WC), and Waist-to-Height Ratio (WHtR) remains undefined. We aimed to evaluate these associations and the modifying effect of genetic susceptibility. Methods: We analyzed 293,318 UK Biobank participants free of AF at baseline. Hazard ratios (HRs) were estimated using Cox proportional hazards models, and non-linearity was assessed using restricted cubic splines. Incremental predictive value was evaluated via Net Reclassification Improvement (NRI). Interactions with AF Polygenic Risk Scores (PRS) were examined. Results: During follow-up, 22,707 incident AF cases occurred. While the TyG index was associated with AF in unadjusted models, this association was nullified after full adjustment. In contrast, composite indices (TyG-BMI, TyG-WC) and WHtR showed robust, positive associations (WHtR HR per SD: 1.30, 95% CI 1.28-1.32). Spline analysis identified non-linear threshold effects (e.g., WHtR inflection at 0.556). Adding WHtR or TyG-BMI to baseline models significantly improved risk reclassification (NRI ~10.3-11.8%, P<0.001), whereas TyG alone did not (P=0.73). Elevated metabolic risk increased AF incidence across all genetic categories, with significant interactions suggesting greater relative impact in low-genetic risk groups. Conclusions: Composite indices integrating central obesity and insulin resistance are superior to the TyG index alone in predicting incident AF. The identification of specific risk thresholds and genetic interactions highlights "metabolic health" as a crucial, modifiable target for AF prevention.

Bicuspid aortic valve (BAV) is one of the most common congenital heart defects but its genetic basis remains incompletely defined. Extracellular matrix components play key roles in outflow tract (OFT) and valve development, but their contribution to BAV is not fully established. Following the analysis of a cohort of BAV patients, we identified a family harbouring a rare human ELASTIN (ELN) variant (p.Gln691X). To assess its pathogenicity, we generated a zebrafish elna/b double knockout (KO) using an RNAless CRISPR Cas9 strategy to avoid genetic compensation. This mutant exhibited cardiovascular defects including OFT anomalies, reduced stroke volume and dysmorphic aortic valves, highlighting Elastins critical role in cardiac development. We then used this model to test the ELN variant identified in the BAV family. We found that wild-type ELN mRNA was able to restore normal cardiac function and morphology, whereas the variant ELN mRNA failed to do so. This study establishes a robust in vivo model to assess ELN variant pathogenicity and provides evidence linking ELASTIN to BAV, opening new avenues for uncovering the genetic mechanisms underlying BAV.

BackgroundPathogenic desmin (DES) variants have been implicated in early-onset atrial disease, yet the mechanisms by which desmin dysfunction alters atrial structure and function remain unclear. Desmin anchors the cytoskeleton to the nuclear envelope (NE) through the linker of nucleoskeleton and cytoskeleton (LINC) complex, suggesting that defects in this network may drive atrial cardiomyopathy. MethodsHuman desmin wild-type (WT) and the pathogenic variants p.S13F, p.N342D, and p.R454W were stably expressed in HL-1 atrial cardiomyocytes. Desmin organization, nuclear morphology, LINC-complex integrity (nesprin-3, lamin A/C), and DNA leakage, assessed by cyclic GMP-AMP synthase (cGAS), were analyzed by confocal microscopy. Action potential duration (APD) and calcium transients (CaT) were measured optically. Human myocardium samples from DES variant carriers were analyzed for validation. Data-independent acquisition (DIA) mass spectrometry profiled atrial proteomes from desmin-network (DN) and titin variant carriers and controls. The heat-shock proteins (HSPs) inducer geranylgeranylacetone (GGA) was evaluated for rescue effects. Resultsp.N342D caused severe filament-assembly defects with prominent perinuclear aggregates, whereas p.S13F showed mixed phenotypes with frequent perinuclear aggregates, and p.R454W largely preserved filamentous networks. p.N342D and p.S13F induced nuclear deformation with disrupted nesprin-3 and lamin A/C distribution. In p.N342D and p.S13F, desmin aggregates drove focal lamin A/C accumulation, nuclear envelope (NE) rupture, DNA leakage, and increased cGAS activation. DES variants significantly shortened APD20/90 and reduced CaT amplitude, indicating pro-arrhythmic electrical remodeling. Atrial proteomics revealed a DN-specific signature enriched for cytoskeletal, NE, intermediate filament, and chaperone pathways, consistent with the structural injury observed in vitro. GGA prevented desmin aggregation and nuclear morphology changes, and mitigated APD shortening in p.N342D-expressing cardiomyocytes. Human myocardium from DES variant carriers showed concordant desmin aggregation and polarized lamin A/C distribution. ConclusionsDES variants induce a desmin-dependent atrial cardiomyopathy characterized by cytoskeletal disorganization, disruption of LINC-complex, NE rupture with DNA leakage, and pro-arrhythmic electrophysiological remodeling. These findings provide mechanistic insight into how DN variants promote atrial disease. HSPs induction by GGA partially restores structural and functional integrity, identifying a potential therapeutic approach for desmin-related atrial cardiomyopathy. Clinical perspectiveWhat is new? O_LIPathogenic DES variants induce a previously unrecognized atrial cardiomyopathy characterized by desmin aggregation, and desmin-network (DN) collapse, disruption of the linker of nucleoskeleton and cytoskeleton (LINC) complex, and nuclear envelope rupture with DNA leakage. C_LIO_LIVariants that lead to desmin aggregation (e.g., p.N342D) cause focal lamin A/C polarization, cyclic GMP-AMP synthase (cGAS) activation, and structural injury at the nuclear envelope. C_LIO_LIDES variants produce pro-arrhythmic electrical remodeling, including action potential duration shortening and impaired Ca{superscript 2} handling in HL-1 atrial cardiomyocytes. C_LIO_LIAtrial proteomics from DN variant carriers reveals enrichment of pathways related to cytoskeletal, nuclear envelope, intermediate filament, and chaperone, supporting a desmin-dependent remodeling program. C_LIO_LIThe heat-shock protein inducer geranylgeranylacetone (GGA) prevents desmin aggregation, restores nuclear morphology, and mitigates electrical and Ca{superscript 2} handling remodeling. C_LI What are the clinical implications? O_LIThese findings establish DN dysfunction as a distinct cause of atrial cardiomyopathy, providing a mechanistic basis for the association between pathogenic DES variants and atrial arrhythmias, including atrial fibrillation. C_LIO_LINuclear envelope rupture and cytosolic DNA leakage represent new mechanistic evidence which links cytoskeletal injury and atrial arrhythmogenesis. C_LIO_LIIdentifying structural vulnerability in DES variant carriers fosters awareness of genetic counseling for atrial disease, enabling early detection and risk stratification. C_LIO_LIThe protective effects of GGA suggest that restoring proteostasis may be a therapeutic strategy for desmin-related atrial cardiomyopathy and potentially other genetic atrial diseases. C_LI Novelty and significance statementO_ST_ABSNoveltyC_ST_ABSThis study identifies a desmin-dependent atrial cardiomyopathy driven by cytoskeletal aggregation, LINC-complex disruption, and nuclear envelope rupture with DNA leakage. We show that pathogenic DES variants are associated with pro-arrhythmic molecular remodeling and that human atrial proteomics confirm nuclear envelope and cytoskeletal injury as core features. Importantly, the heat-shock protein-inducer GGA rescues structural, molecular, and electrophysiological defects, revealing a modifiable pathway in desmin-mediated atrial disease. SignificanceThese findings provide the first integrated mechanistic explanation linking DN variants to atrial cardiomyopathy. By uncovering nuclear envelope rupture and cGAS activation as key drivers of atrial cardiomyopathy, this work expands the molecular framework for inherited atrial disease and highlights proteostasis enhancement as a potential therapeutic strategy for patients carrying DES and related cytoskeletal variants. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=166 HEIGHT=200 SRC="FIGDIR/small/26348559v1_ufig1.gif" ALT="Figure 1"> View larger version (51K): org.highwire.dtl.DTLVardef@34b48forg.highwire.dtl.DTLVardef@3a2be1org.highwire.dtl.DTLVardef@116e8e6org.highwire.dtl.DTLVardef@1147b94_HPS_FORMAT_FIGEXP M_FIG C_FIG

IntroductionSpontaneous coronary artery dissection (SCAD) is frequently accompanied by persistent symptoms of unknown pathogenesis after the index event. Autonomic dysfunction is a plausible mechanism for these but has not been systematically characterized. We quantified antecedent and contemporary autonomic symptoms in survivors of SCAD and examined their associations with cardiac and extra-cardiac symptoms and health-related quality of life. MethodsThis cross-sectional study recruited 227 volunteers from multiple countries with a self-reported history of SCAD. Participants completed validated patient-reported measures, including the Composite Autonomic Symptom Score-31 (COMPASS-31), Anxiety Sensitivity Index-3 (ASI-3), and EuroQol-5 Dimension-5L (EQ-5D-5L). They also completed an internally derived retrospective autonomic predisposition score assessing symptoms during adolescence and early adulthood. ResultsParticipants were predominantly female (97.8%), median age 53 (47-58) years, and were surveyed a median of 3 (1-5) years after their index SCAD event. 21.6% reported SCAD recurrence. Moderate autonomic symptom burden (COMPASS-31 [&ge;]20) was present in 56.4% and severe burden ([&ge;]40) in 16.3%. History of antecedent autonomic symptoms was the strongest independent predictor of contemporary autonomic symptom burden after adjustment for demographic and clinical covariates ({beta}=0.514; P <0.001). Greater autonomic symptom burden independently predicted lower EQ-5D health utility ({beta}=-0.150; P=0.029) and was associated with the ASI-3 physical concerns ({beta}=0.232; P <0.001), but not social concerns domain. Autonomic symptoms were not associated with SCAD recurrence. ConclusionSymptoms of autonomic dysregulation are common in survivors of SCAD and are associated with reduced quality of life. Their association with antecedent dysautonomic features during adolescence and early adulthood suggests a longstanding predisposition, the significance of which warrants further evaluation. Clinical PerspectiveO_ST_ABSWhat Is New?C_ST_ABSO_LISelf-reported antecedent and current autonomic symptoms are common in survivors of spontaneous coronary artery dissection and are associated with poorer health-related quality of life, greater fatigue, and greater psychological distress. C_LI What Are the Clinical Implications?O_LIAutonomic symptoms warrant clinical recognition in patients with prior spontaneous coronary artery dissection, not only as a post-event complaint but also as a potential marker of pre-existing autonomic vulnerability that may influence recovery experience. C_LIO_LIGreater awareness of autonomic symptom burden may support more personalized follow-up, patient counseling, and rehabilitation planning to help patients return more safely and confidently to daily activities, work, and family life. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=135 SRC="FIGDIR/small/26351434v1_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@1589559org.highwire.dtl.DTLVardef@b5423forg.highwire.dtl.DTLVardef@103b97org.highwire.dtl.DTLVardef@1b8378f_HPS_FORMAT_FIGEXP M_FIG C_FIG

BackgroundAnomalous aortic origin of a coronary artery (AAOCA) can cause myocardial ischemia and sudden cardiac death. The optimal stress-testing strategy and impact of coronary morphology on ischemia remain unclear. We assessed the effect of coronary morphology on stress-test completion and results across multiple test modalities. MethodsThis retrospective cohort study included 531 adults with AAOCA at our institution (7/2015 - 3/2023). Coronary morphology was characterized by the anomalous coronary (right [RCA], left main [LMCA], left anterior descending [LAD], left circumflex) and the course type (intramural, interarterial-only, transseptal, and other [prepulmonic and retroaortic]). Exercise and pharmacologic stress tests were positive if ischemia included the territory of the anomalous coronary. A mixed-effect logistic regression modeled the odds of a positive test based on morphology, comorbidities, and modality. A random forest regression analyzed the stress iFR as a continuous outcome. ResultsStress test results were available for 396 (75%) of patients (age 50 {+/-} 17 years; 42% female). Stress testing included 699 ECGs, 198 echocardiograms, 288 SPECTs, 133 PETs, and 103 dobutamine iFR studies. Completion of invasive dobutamine iFR (versus noninvasive-only) stress testing was associated with high-risk coronary morphology, p<0.001. Coronary morphology that trended toward higher adjusted odds of ischemia included the anomalous LMCA (OR: 2.1, p=0.054) and intramural course (OR: 1.9, p=0.14). Compared to ECG, iFR had higher adjusted odds of a positive result (OR: 27, p<0.001), followed by PET (OR: 9.0, p<0.001). In the random forest regression, stress iFR value was lowest for LAD (0.75) compared to LMCA (0.83) and RCA (0.84). For course type, transseptal (strongly correlated with the anomalous LAD) had the lowest stress iFR (0.77), followed by intramural (0.83), and interarterial (0.88). ConclusionsIn our adult AAOCA cohort, high-risk coronary morphology demonstrated a borderline association with ischemia on stress testing, whereas stress test modality was the strongest determinant of ischemia detection. Invasive stress testing was reserved for higher-risk coronary morphology. These findings underscore that effective risk stratification in AAOCA integrates clinical symptoms, coronary morphology, and stress test modality. Long-term follow-up is needed to determine the optimal strategy for ischemia evaluation. Clinical PerspectivesO_ST_ABSWhat is new?C_ST_ABSO_LIIn this single-center registry of adults with anomalous aortic origin of a coronary artery (AAOCA), 75% of patients had stress testing, enabling the largest analysis of how anomalous coronary morphology impacts stress testing practices and the presence of ischemia. C_LIO_LIConsistent with published data in younger AAOCA cohorts, our adult population (mean age >50 years) trended toward increased risk of ischemia with an anomalous left coronary and intramural course. C_LIO_LIComparing various stress test modalities for AAOCA, instantaneous wave-free, followed by positron emission tomography and single-photon emission computed tomography, have higher odds of being positive for ischemia than electrocardiogram and echocardiograms. C_LI What are the clinical implications?O_LIAdults with AAOCA remain at risk for ischemia and require careful risk stratification, with coronary morphology and clinical symptoms informing stress test selection and result interpretation. C_LIO_LIFor higher risk morphologic variants of AAOCA, consider further risk stratification with invasive coronary provocative studies to detect inducible ischemia. C_LI