Nature Cardiovascular Research

Amyloid light-chain cardiomyopathy (ALCA) is an infiltrative disorder marked by misfolded immunoglobulin light-chain deposition in the myocardium, ultimately leading to cardiac dysfunction. Despite its clinical severity, the underlying mechanisms remain poorly understood. Here, we integrated multi-omics analyses of human cardiac samples to construct a comprehensive cellular and spatial atlas of the ALCA heart. We observed a marked expansion of PTX3+ fibroblasts (FB), which undergo a distinct phenotypic transition into pro-fibrotic POSTN+ FB regulated by EGR1.Concurrently, SPP1+ macrophages (Mac) emerged as major drivers of fibrosis, interacting robustly with PTX3+ FB via APP-CD74, GAS6-MERTK, and TGF-{beta}1-TGF-{beta}1/2 pathways. Endothelial cell (EC) profiling revealed substantial vascular remodeling characterized by the emergence of specialized capillary-like immune endothelial cells expressing chemokines CXCL1, CXCL3, and CCL2, alongside depletion of functional capillary EC. Immunologically, elevated cytotoxic CD8+ T cells and reduced NK cells contributed to an imbalanced inflammatory milieu, with NF{kappa}B2 orchestrating both fibrotic and immune pathways across multiple cell types. These findings highlight the pivotal role of fibroblast-immune crosstalk, particularly the SPP1+ Mac-PTX3+ FB axis, in driving ALCA pathogenesis. Targeting these pathological cellular interactions may offer a promising therapeutic avenue to mitigate fibrosis, restore immune homeostasis, and improve cardiac function in ALCA.

Aortic aneurysm and dissection (AAD) are life-threatening conditions for which there is a lack of effective pharmacological therapies. Hypertension is a known risk factor for AAD, leading to the common prescription of antihypertensive medications for AAD patients. The impaired contractility of vascular smooth muscle cells (VSMCs) is strongly linked to AAD, yet the role of calcium channel blockers (CCBs), which directly inhibit VSMC contractility, in AAD onset of hypertensive patients remains unclear. We thus analyzed data from 501,878 initially AAD-free participants and reported that CCB use had greater risks of AAD (adjusted HR=1.31, 95% CI: 1.17-1.48), TAAD (adjusted HR=1.23, 95% CI: 1.00-1.50), and AAA (adjusted HR=1.32, 95% CI: 1.13-1.53), than hypertensive patients not receiving antihypertensive medication. during a median follow-up of 13.5 years (P<0.001). In mouse models induced by angiotensin II, elastase, and {beta}-aminopropionitrile (BAPN), various subtypes of CCBs significantly increased aortic stiffness and the risk of AAD. Of 95 patients with type B aortic dissection included after endovascular repair surgery in the secondary data analysis, CCBs (69 patients) limited aortic aneurysm/dissection regression compared with that associated with other antihypertensives (26 patients). Moreover, the silencing of protein kinase cGMP-dependent 1 (PRKG1) to restore VSMCs contractility significantly mitigated CCB-induced aortic stiffness and the incidence of AAD. These findings suggest that CCBs may increase AAD risk and post-stent surgery prognosis, highlighting the need for caution when prescribing CCBs to hypertensive patients at risk for AAD.

2.BackgroundGenetic cardiomyopathies display variable penetrance and phenotypic expression, highlighting the influence of environmental modulators. Myocarditis, commonly triggered by cardiotropic viruses, overlaps clinically with genetic cardiomyopathies. Consequently, these infections are implicated as secondary factors that accelerate disease onset and progression, yet their precise impact in specific genetic settings remains unexplored. MethodsTo interrogate this, genetic mouse models heterozygous for a mutant allele of desmosomal plakophilin-2 (Pkp2+/-) or sarcomeric titin (Ttn+/-), genes frequently linked to acute myocarditis, were challenged with murine cytomegalovirus (MCMV) to determine how latent infection influences myocardial inflammation, tissue remodeling, and cardiac performance. Integrated experimental approaches, including echocardiography, histology, flow cytometry, single-cell RNA sequencing, as well as cytokine and kinome analyses, defined immune and signaling responses in infected versus non-infected hearts. ResultsAcute, MCMV-induced viral myocarditis and subsequent latent MCMV infection unmasked early disease onset in Pkp2+/- animals, leading to progressive systolic impairment, whereas in Ttn+/- mice cardiac structure and function remained preserved throughout infection. Cardiac immune profiling uncovered infection- and genotype-specific divergence: both genetic models showed a stable myocardial effector-memory CD8+ T-cell response to MCMV, but only Pkp2+/- hearts recruited additional Ly6C+ CCR2+ monocytes and macrophages with distinct inflammatory signatures. In the absence of infection, Pkp2 insufficiency initiated subclinical CCL2 secretion and subsequent recruitment of CCR2+ cells, reflecting early immune activation preceding age-associated functional and structural decline. At this stage, cytokine and kinase evaluations indicated a balance between proinflammatory and compensatory signals. However, with aging or following MCMV challenge, this balance shifted towards persistent inflammation, evidenced by chronic upregulation of cytokines and activation of signaling pathways, which ultimately led to adverse effects and myocardial dysfunction. ConclusionsManifestation of genetic cardiomyopathies depends on interactions between inherited susceptibility and environmental stressors. Here, we show that cytomegalovirus infection intensifies inflammation in PKP2-related cardiomyopathy. In contrast, TTN-linked cardiomyopathy does not exhibit increased inflammation under the same conditions. For individuals carrying desmosomal variants, infection control and tailored anti-inflammatory strategies may attenuate or delay disease manifestation and progression.

Atherogenesis involves an interplay of inflammation, tissue remodeling and cellular transdifferentiation (CTD), making it especially difficult to precisely delineate its pathophysiology. Here we examine the single-cell transcriptome of entire atherosclerotic core (AC) plaques and patient-matched proximal adjacent (PA) portions of carotid artery tissue from patients undergoing carotid endarterectomy. We use a novel tissue dissociation strategy, single-cell RNA sequencing, and systems-biology approaches to analyze the transcriptional profiles of six main cell populations and identify key gene drivers of pathogenic biological processes in vascular smooth muscle cells (VSMCs) and endothelial cells (ECs). Our results reveal an anatomic continuum whereby PA cells promote and respond to inflammatory processes and eventually transition through CTD into matrix-secreting cells in the AC. Inflammatory signaling in PA ECs is driven by IL6, while TNFa signaling defines inflammation in both PA ECs and VSMCs. Furthermore, we identify POSTN, SPP1 and IBSP in AC VSMCs, and ITLN1, SCX and S100A4 in AC ECs as key drivers of CTD in the atherosclerotic core. These results establish an anatomic framework for atherogenesis and suggest a site-specific strategy for disruption of disease progression.

Controlling hyperlipidemia has reduced but not eliminated atherosclerotic cardiovascular disease as a predominant cause of human mortality. Here, we report that loss of the immunoregulatory gene GTPase of immunity-associated protein 6 (GIMAP6), causes an inflammatory vasculopathy and accelerated atherosclerosis in the absence of hyperlipidemia. These pathologic changes in turn result in progressive cardiac ischemia, myocardial infarction, and heart failure, culminating in early death. In humans, rare deleterious GIMAP6 variants are associated with premature severe cardiovascular disease. These findings reveal GIMAP6 to play an important protective role against atherosclerotic cardiovascular disease whose identification offers opportunities for improved risk management and a target for new therapies.

Hypertrophic cardiomyopathy (HCM) is associated with phenotypic variability. To gain insights into transcriptional regulation of cardiac phenotype, single-nucleus linked RNA-/ATAC-seq was performed in 5-week-old control mouse-hearts (WT) and two HCM-models (R92W-TnT, R403Q-MyHC) that exhibit differences in heart size/function and fibrosis; mutant data was compared to WT. Analysis of 23,304 nuclei from mutant hearts, and 17,669 nuclei from WT, revealed similar dysregulation of gene expression, activation of AP-1 TFs (FOS, JUN) and the SWI/SNF complex in both mutant ventricular-myocytes. In contrast, marked differences were observed between mutants, for gene expression/TF enrichment, in fibroblasts, macrophages, endothelial cells. Cellchat predicted activation of pro-hypertrophic IGF-signaling in both mutant ventricular-myocytes, and profibrotic TGF{beta}-signaling only in mutant-TnT fibroblasts. In summary, our bioinformatics analyses suggest that activation of IGF-signaling, AP-1 TFs and the SWI/SNF chromatin remodeler complex promotes myocyte hypertrophy in early-stage HCM. Selective activation of TGF{beta}-signaling in mutant-TnT fibroblasts contributes to genotype-specific differences in cardiac fibrosis.

BackgroundAscending thoracic aortic dissection (ATAD) is characterized by extensive macrophage (M{Phi}) accumulation and profound inflammation; however, the mechanisms sustaining pro-inflammatory M{Phi} activation remain incompletely defined. Emerging evidence indicates that epigenetically generated immune memory drives innate immune cells toward persistent inflammatory states. In this study, we investigated whether epigenetic reprogramming governs M{Phi} phenotypic fate and contributes to ATAD pathogenesis. MethodsWe performed single-cell RNA sequencing of human ascending aortic tissues from controls, patients with ascending thoracic aortic aneurysm (ATAA), and patients with acute ascending thoracic aortic dissection (ATAD). We also performed integrated single-cell RNA sequencing, single-cell ATAC sequencing, and spatial transcriptomics in an angiotensin II (Ang II)-infused mouse model. The role of the STING-IRF3 signaling axis in M{Phi} epigenetic programming was examined using M{Phi}-Sting -/- and M{Phi}-Irf3-/- mice. ResultsIn human and mouse aortic tissues, we identified multiple functional M{Phi} populations including pro-inflammatory, phagocytic/anti-inflammatory, proliferative, and reparative/healing M{Phi}s. Aortic M{Phi}s in both sporadic ATAD patients and Ang II-induced ATAD mice exhibited a pronounced pro-inflammatory bias with enhanced differentiation toward pro-inflammatory M{Phi}s and impaired differentiation toward phagocytic/anti-inflammatory states. Pro-inflammatory M{Phi}s were particularly abundant in dissection sites, whereas phagocytic M{Phi}s were enriched in discrete adventitial niches. Origin analyses revealed a substantial increase in CCR2 recruited M{Phi}s within the aortic wall, which preferentially differentiated into pro-inflammatory M{Phi}s. In contrast, LYVE1 resident M{Phi}s-- predominantly biased toward phagocytic phenotypes--were markedly depleted in ATAD. Single-cell ATAC sequencing identified coordinated chromatin remodeling with increased accessibility at pro-inflammatory gene loci and decreased accessibility at phagocytic gene loci. Among candidate transcriptional regulators identified, IRF family TFs, including IRF3 emerged as unique factors capable of simultaneously promoting pro-inflammatory gene programs while suppressing phagocytic gene expression. Mechanistically, STING-IRF3 signaling orchestrates this biased transcriptional state, likely through coordinated BRG1-dependent chromatin opening at pro-inflammatory gene loci and chromatin closing at phagocytic/anti-inflammatory gene loci. M{Phi} specific Sting -/- and Irf3-/- mice exhibited attenuated inflammatory reprogramming and reduced aortic destruction and dissection. ConclusionsThese findings identify STING-IRF3-mediated epigenetic programming of M{Phi}s as a fundamental mechanism driving aortic inflammation and ATAD development. Targeting M{Phi} epigenetic programming may represent a promising therapeutic strategy to prevent aortic dissection. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=189 HEIGHT=200 SRC="FIGDIR/small/701198v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@c97bcdorg.highwire.dtl.DTLVardef@1df0ca8org.highwire.dtl.DTLVardef@b7fd04org.highwire.dtl.DTLVardef@1443e16_HPS_FORMAT_FIGEXP M_FIG C_FIG

Acquisition and maintenance of vascular smooth muscle cell (VSMC) fate are important for vascular development and homeostasis; however, little is known about the key determinant for VSMC fate and vascular homeostasis. We found that VSMC-specific Lkb1 ablation in Lkb1flox/flox;Tagln-Cre mice caused severe vascular abnormalities and embryonic lethality. VSMC-specific deletion of Lkb1 in tamoxifen-inducible Lkb1flox/flox;Myh11-Cre/ERT2 mice progressively induced aortic/arterial dilation, aneurysm, rupture, and premature death. Single-cell RNA sequencing and imaging-based lineage tracing showed that Lkb1-deficient VSMCs underwent dynamic transcriptional reprogramming and transformed gradually from early modulated VSMCs to fibroblast-like, chondrocyte-like, and even osteocyte-like cells. VSMC transformation followed by extracellular matrix remodeling and inflammatory cell infiltration contributed to the arterial aneurysm formation in tamoxifen-induced Lkb1flox/flox;Myh11-Cre/ERT2 mice. Finally, we found that VSMC-specific Lkb1 ablation resulted in decreased vascular contractility, hypotension, and impaired responses to angiotensin II and vessel injury in vivo. Lkb1 is therefore a key determinant of mouse VSMC fate that prevents VSMC reprogramming and sustains vascular homeostasis. Our findings have important implications for understanding the pathogenesis of aortic aneurysm.

BackgroundLeft ventricular noncompaction (LVNC) is a prevalent cardiomyopathy associated with excessive trabeculation and thin compact myocardium. Patients with LVNC are vulnerable to cardiac dysfunction and at high risk of sudden death. Although sporadic and inherited mutations in cardiac genes are implicated in LVNC, understanding of the mechanisms responsible for human LVNC is limited. MethodsWe screened the complete exome sequence database of the Pediatrics Cardiac Genomics Consortium and identified a cohort with a de novo chromodomain helicase DNA-binding protein 4 (CHD4) proband, CHD4M202I, with congenital heart defects. We engineered a patient-specific model of CHD4M202I (mouse CHD4M195I). Histological analysis, immunohistochemistry, flow cytometry, transmission electron microscopy, and echocardiography were used to analyze cardiac anatomy and function. Ex vivo culture, immunopurification coupled with mass spectrometry, transcriptional profiling, and chromatin immunoprecipitation were performed to deduce the mechanism of CHD4M195I-mediated ventricular wall defects. ResultsCHD4M195I/M195I mice developed biventricular hypertrabeculaion and noncompaction and died at birth. Proliferation of cardiomyocytes was significantly increased in CHD4M195I hearts, and the excessive trabeculation was associated with accumulation of extracellular matrix (ECM) proteins and a reduction of ADAMTS1, an ECM protease. We rescued the hyperproliferation and hypertrabeculation defects in CHD4M195I hearts by administration of ADAMTS1. Mechanistically, the CHD4M195I protein showed augmented affinity to endocardial BRG1. This enhanced affinity resulted in failure of derepression of Adamts1 transcription such that ADAMTS1-mediated trabeculation termination was impaired. ConclusionsOur study reveals how a single mutation in the chromatin remodeler CHD4, in mice or humans, modulates ventricular chamber maturation and that cardiac defects associated with the missense mutation CHD4M195I can be attenuated by the administration of ADAMTS1. Clinical PerspectiveO_ST_ABSWhat Is New?C_ST_ABSO_LIA patient-specific mouse model of CHD4M202I develops ventricular hypertrabeculation and dies at birth. C_LIO_LIProliferation of cardiomyocytes is significantly enhanced in CHD4M195I mice. C_LIO_LIADAMTS1 is significantly downregulated in CHD4M195I mice. C_LIO_LIClose interaction between CHD4M195I and BRG1 robustly and continuously represses Adamts1 transcription, which impairs ADAMTS1-mediated termination of trabeculation in the developing mutant heart. C_LI What Are the Clinical Implications?O_LIThis study provides a unique mouse model of ventricular noncompaction cardiomyopathy that faithfully reflects human patients genetic condition without disturbing the target genes expression and localization. C_LIO_LITranscriptional repression of ECM protease ADAMTS1 by CHD4-BRG1 interaction is detrimental to ventricular wall maturation; maintaining appropriate ADAMTS1 levels in the heart could be a promising therapeutic approach for treating ventricular noncompaction cardiomyopathy. C_LI

Background: Non-monogenic arteriopathies are often classified as distinct entities according to the arterial territory involved, yet they share clinical features and may co-occur in the same individual. This pattern suggests shared susceptibility across anatomically distinct arteriopathies, potentially driven by common biological and genetic mechanisms. Methods: We investigated the shared genetic architecture of five arteriopathies (cervical artery dissection (CeAD), intracranial aneurysm (IA), spontaneous coronary artery dissection (SCAD), aortic aneurysm and dissection (AAD), and fibromuscular dysplasia (FMD)) using LD score regression, Association analysis based on SubSETs (ASSET), pairwise Multi-Trait Analysis of Genome-wide association summary statistics (MTAG), pleiotropy mapping and Mendelian randomization (MR) to identify shared loci and prioritise candidate causal genes. Results: LD score regression identified significant positive genetic correlations between CeAD-SCAD (rg = 0.64), IA-AAD (rg = 0.33), IA-SCAD (rg = 0.37), CeAD-AAD (rg = 0.56) and SCAD-AAD (rg = 0.20). ASSET identified 37 shared independent loci, and in MTAG analyses, one novel locus was identified for CeAD and SCAD (SLC39A8) and one for IA (FGF5). 13 loci showed strong cross-trait colocalization, including PHACTR1, LRP1, and CDKN2B-AS1. Using the Genotype-Phenotype Map, we found that arteriopathy-associated variants colocalized with blood pressure- and migraine-related traits, while many showed effect directions opposite to those observed for coronary artery disease. Proteome-wide MR identified 67 circulating proteins associated with at least one trait, including ECM1 and SHISA5 for CeAD and FGF5 for IA, with 17 supported by colocalization. Transcriptome-wide MR identified 204 colocalized tissue?specific signals, of which, 14 were shared across multiple traits. Enrichment analyses implicated pathways related to vascular development, smooth muscle cell function, extracellular matrix organization, and TGF-? signaling. Conclusions: These findings support shared genetic architecture across anatomically distinct arteriopathies, implicating pathways involved in vascular structure and prioritising therapeutic targets for future mechanistic investigation.

Arterial restenosis following balloon angioplasty - a procedure performed to re-establish vessel patency in atherosclerotic cardiovascular disease - remains a major clinical challenge and a key barrier to durable revascularization. Endothelial denudation induced by angioplasty triggers an inflammatory cascade that drives vascular smooth muscle cell (VSMC) proliferation, migration, and phenotypic switching, culminating in neointimal hyperplasia and restenosis1. Human genetics-guided target discovery has proven more effective than non-guided approaches in revealing causal pathways of complex cardiovascular traits2. Genetic variants at Histone Deacetylase 9 (HDAC9) are a major risk factor for cardiovascular disease3,4 and is associated with increased carotid intima-media thickness and modulation of VSMC phenotype4. Here, using an experimental model of arterial injury that faithfully mirrors the vascular response to balloon angioplasty in humans, we show that HDAC9 drives maladaptive remodeling of the arterial wall following vascular injury.

ObjectiveVascular smooth muscle cells (VSMCs) are the primary contractile component of blood vessels and can undergo phenotypic switching from a contractile to a synthetic phenotype in vascular diseases such as coronary artery disease (CAD). This process leads to decreased expression of SMC lineage genes and increased proliferative, migratory and secretory abilities that drive disease progression. Super-enhancers (SE) and occupied transcription factors are believed to drive expression of genes that maintain cell identify and homeostasis. The goal of this study is to identify novel regulator of VSMC homeostasis by screening for SE-regulated transcription factors in arterial tissues. Approach and ResultsWe characterized human artery SEs by analyzing the enhancer histone mark H3K27ac ChIP-seq data of multiple arterial tissues. We unexpectedly discovered the transcription factor PRDM16, a GWAS identified CAD risk gene with previously well-documented roles in brown adipocytes but with an unknown function in vascular disease progression, is enriched with artery-specific SEs. Further analysis of public bulk RNA-seq and scRNA-seq datasets, as well as qRT-PCR and Western blotting analysis, demonstrated that PRDM16 is preferentially expressed in arterial tissues and in contractile VSMCs but not in visceral SMCs, and down-regulated in phenotypically modulated VSMCs. To explore the function of Prdm16 in vivo, we generated Prdm16 SMC-specific knockout mice and performed histological and bulk RNA-Seq analysis of aortic tissues. SMC-deficiency of Prdm16 does not affect the aortic morphology but significantly alters expression of many CAD risk genes and genes involved in VSMC phenotypic modulation. Specifically, Prdm16 negatively regulates the expression of Tgfb2 that encodes for an upstream ligand of TGF-{beta} signaling pathway, potentially through binding to the promoter region of Tgfb2. These transcriptomic changes likely disrupt VSMC homeostasis and predispose VSMCs to a disease state. ConclusionsOur results suggest that the CAD risk gene PRDM16 is preferentially expressed in VSMCs and is a novel regulator of VSMC homeostasis. Future studies are warranted to investigate its role in VSMCs under pathological conditions such as atherosclerosis.

Key findings1. PRC2 regulates EC shear stress responses. 2. PRC2 governs Klf2/4 suppression downstream of Pcdhg. 3. High PRC2 in ASCVD-prone arterial regions suppresses Klf2/4 to promote ASCVD. 4. Athero-protective Klf2/4 induction upon PRC2 inhibition requires Notch signaling. 5. Tazemetostat, an FDA approved PRC2 inhibitor, slows ASCVD progression and improves markers of plaque stability. Atherosclerotic cardiovascular disease (ASCVD), the leading cause of mortality worldwide, is driven by endothelial cell inflammatory activation and counter-balanced by anti-inflammatory transcription factors Klf2 and Klf4 (Klf2/4). Understanding vascular endothelial inflammation to develop effective treatments is thus essential. Here, we identify, Polycomb Repressive Complex (PRC) 2, which blocks gene transcription by trimethylating histone3 Lysine27 in gene promoter/enhancers, as a potent, therapeutically targetable determinant of vascular inflammation and ASCVD progression. Bioinformatics identified PRC2 as a direct suppressor of Klf2/4 transcription. Klf2/4 transcription requires Notch signaling, which reverses PRC2 modification of Klf2/4 promoter/enhancers. PRC2 activity is elevated in human ASCVD endothelium. Treating mice with established ASCVD with tazemetostat, an FDA approved pharmacological inhibitor of PRC2, slowed plaque progression by 50% and drastically improved markers of plaque stability. This study elucidates a fundamental mechanism of vascular inflammation, thus identifying a potential method for treating ASCVD and possibly other vascular inflammatory diseases.

Proliferating cells rewire glucose metabolism away from oxidative ATP production towards glycolysis and anabolic branch pathways (the Warburg effect). In contrast, the contractile function of terminally differentiated cardiomyocytes in the heart critically depends upon mitochondrial ATP generation. Adult cardiomyocytes are largely non-proliferative but undergo hypertrophy in response to increased heart workload. A fundamental question in the field is how cardiac metabolism is modified to balance competing energetic and anabolic demands. We previously reported that the redox-signalling H2O2-generating enzyme, NADPH oxidase 4 (NOX4), facilitates compensated cardiac function in hearts undergoing hypertrophic remodelling. Here, we show that NOX4 induces Warburg-type reprogramming of glucose metabolism in the healthy heart, with increased flux into the pentose phosphate, serine and nucleotide biosynthetic pathways. Using an integrated multiomics approach, we uncover a NOX4-regulated network involving interplay between direct transcriptional activation of metabolic genes via NRF2 [aka NFE2L2] and ATF4 and a broader epigenetic regulation. This reprogramming of glucose intermediary metabolism along with previously reported NOX4-mediated enhancement of fatty acid oxidation may serve to optimally support the dual requirements of increased energy demand and remodelling in the heart. Our findings identify a novel paradigm for redox-regulated Warburg-type metabolic reprogramming in the terminally differentiated heart.

Aortic dissection (AD) is a cardiovascular disease with rapid onset and extremely high short-term mortality, currently lacking specific peripheral blood-based biomarkers and effective treatments. Here, our analysis of AD samples from animal models and human patients revealed elevated blood levels of CTHRC1, a protein secreted by vascular fibroblasts. Furthermore, CTHRC1 regulates the phenotypic switch of vascular smooth muscle cells (VSMCs) during arterial remodeling by binding to ADAM9 on the VSMC membrane, activating the ERK1/2 signaling pathway, and promoting a contractile-to-synthetic transition. In Ang-II/BAPN induced mouse models, genetic ablation or antibody-mediated blockade of CTHRC1 effectively prevented AD development. These findings unveil CTHRC1 as a critical regulator of VSMC phenotype and aortic structural integrity via the ERK1/2 pathway, suggesting its potential as a novel serum diagnostic biomarker for AD diagnostic and a promising therapeutic target. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/649636v1_ufig1.gif" ALT="Figure 1"> View larger version (75K): org.highwire.dtl.DTLVardef@6a3f14org.highwire.dtl.DTLVardef@15353e7org.highwire.dtl.DTLVardef@1a9928forg.highwire.dtl.DTLVardef@1d4ef3d_HPS_FORMAT_FIGEXP M_FIG C_FIG In briefCTHRC1 is aberrantly expressed in aortic adventitial fibroblasts from dissected aortas and function as an exocrine mediator that induces phenotypic switching of vascular smooth muscle cells. A monoclonal antibody targeting CTHRC1 demonstrates therapeutic potential in mouse models of aortic dissection. HighlightsO_LICTHRC1 is highly expressed in aortic adventitial fibroblasts from dissected aortas C_LIO_LICTHRC1 interacts with ADAM9 to induce phenotypic switching of VSMCs C_LIO_LIAn anti-CTHRC1 antibody inhibits aortic dissection formationinvivo C_LIO_LIElevated levels of serum CTHRC1 can be used to identify AD in patients presenting with chest pain C_LI

Cardiomyocytes die and do not regenerate after an injury such as a myocardial infarction (MI), a leading cause of mortality worldwide. Following MI, cardiac fibroblasts (CFs) proliferate and differentiate into myofibroblasts, which then produce increased collagen and extracellular matrix (ECM) leading to fibrosis. Fibrosis can weaken cardiac output via excessive stiffening and interference with electric signal transmission, but can also prevent wall rupture under load (reviewed in (1)). Thus, dampening fibrosis has been investigated as a potential therapeutic intervention. Most mammalian cells possess a single primary cilium involved in intercellular communication. We investigated the role of CF primary cilia in sensing injury signals and initiating fibrotic remodeling. We found that deleting CF cilia reduced fibrosis and improved cardiac output after MI, demonstrating that cilia act as a signaling hub that amplifies the fibrotic response in the injured heart.

To elucidate the pathogenesis of vein of Galen malformations (VOGMs), the most common and severe congenital brain arteriovenous malformation, we performed an integrated analysis of 310 VOGM proband-family exomes and 336,326 human cerebrovasculature single-cell transcriptomes. We found the Ras suppressor p120 RasGAP (RASA1) harbored a genome-wide significant burden of loss-of-function de novo variants (p=4.79x10-7). Rare, damaging transmitted variants were enriched in Ephrin receptor-B4 (EPHB4) (p=1.22x10-5), which cooperates with p120 RasGAP to limit Ras activation. Other probands had pathogenic variants in ACVRL1, NOTCH1, ITGB1, and PTPN11. ACVRL1 variants were also identified in a multi-generational VOGM pedigree. Integrative genomics defined developing endothelial cells as a key spatio-temporal locus of VOGM pathophysiology. Mice expressing a VOGM-specific EPHB4 kinase-domain missense variant exhibited constitutive endothelial Ras/ERK/MAPK activation and impaired hierarchical development of angiogenesis-regulated arterial-capillary-venous networks, but only when carrying a "second-hit" allele. These results illuminate human arterio-venous development and VOGM pathobiology and have clinical implications.

BACKGROUNDMono-ADP ribosylation is a post-translational modification that regulates various cellular physiological processes, including cell cycle progression, genomic stability, transcription, and cellular protein turnover. PARP16 is an endoplasmic reticulum (ER)-localized mono-ADP-ribosyltransferase that has been shown to regulate the unfolded protein response and maintain ER homeostasis under stress conditions. Despite its established role in ER stress signaling, the functional significance of PARP16 in cardiac pathophysiology, particularly in cardiac hypertrophy and heart failure, remains poorly understood. In this study, we aim to investigate the role of PARP16 in cardiac hypertrophy and heart failure using in vitro and mouse model systems. METHODSWe analysed PARP16 expression in human heart failure samples as well as in heart failure-based mouse models. We evaluated gene expression by RT-PCR, immunoblotting, and confocal microscopy to understand the role of PARP16 in heart failure under phenylephrine- or isoproterenol-treated conditions. We also investigated the role of PARP16 in regulating cardiac function in genetically engineered mouse models, including whole-body PARP16 knockout, cardiac-specific PARP16 knockout, inducible cardiac-specific PARP16 knockout, and cardiac-specific PARP16 Transgenic mice. We performed echocardiography to assess cardiac function. We also used an in vitro primary cardiomyocyte system to knock down and overexpress PARP16. We performed RNA sequencing and mass spectrometry, followed by molecular docking, molecular dynamics simulation, immunoprecipitation, and luciferase assay to characterise the molecular mechanism by which PARP16 regulates cardiac function. RESULTSHuman heart failure samples showed reduced PARP16 expression. PARP16 expression was also significantly reduced in models of heart failure, including the hearts of isoproterenol-treated C57B/L6 mice and phenylephrine-treated primary cardiomyocytes. PARP16-deficient NRCMs showed signs of pathological remodelling. Whole-body, cardiac-specific, and inducible cardiac-specific PARP16 KO mice exhibited cardiac remodelling and dysfunction. In contrast, cardiac-specific PARP16-overexpressing mice were protected from iso-induced cardiac hypertrophy. Mechanistically, several hypertrophic signalling pathway genes are dysregulated in PARP16 knockout mouse hearts concomitant with upregulated NFAT1 transcriptional activity and nuclear translocation. PARP16 binds to and catalytically downregulates NFAT activity, thereby maintaining cardiac function. Mass spectrometry analysis showed that PARP16 is involved in ADP-ribosylation of NFAT1 at E398 and T533. Pharmacological inhibition of NFAT activation attenuates structural and functional abnormalities associated with PARP16 deficiency. CONCLUSIONSPARP16 binds to and inhibits NFAT1 activity to regulate cardiac function in mice, and its downregulation may activate NFAT1 signalling, leading to hypertrophy. In this manner, PARP16 plays a critical role in cardiac hypertrophy and failure and may serve as a potential therapeutic target for the treatment of heart failure.

The risk of early recurrent events after stroke remains high despite currently established secondary prevention strategies. Risk is particularly high in patients with atherosclerosis, with more than 10% of patients experiencing early recurrent events. However, despite the enormous medical burden of this clinical phenomenon, the underlying mechanisms leading to increased vascular risk and recurrent stroke are largely unknown. Here, using a novel mouse model of stroke-induced recurrent ischemia, we show that stroke leads to activation of the AIM2 inflammasome in vulnerable atherosclerotic plaques via an increase of circulating cell-free DNA from the ischemic tissue. Enhanced plaque inflammation post-stroke results in plaque destabilization and atherothrombosis, finally leading to arterio-arterial embolism and recurrent stroke within days after the index stroke. We confirm key steps of plaque destabilization also after experimental myocardial infarction and in carotid artery plaque samples from patients with acute stroke. Neutralization of cell-free DNA by DNase treatment or inhibition of inflammasome activation reduced the rate of stroke recurrence after experimental stroke. Our findings present an explanation for the high recurrence rate after incident ischemic events in atherosclerotic patients. The detailed mechanisms uncovered here provide so far clinically uncharted therapeutic targets for which we show high efficacy to prevent recurrent events. Targeting DNA-mediated inflammasome activation after remote tissue injury represents a promising avenue for further clinical development in the prevention of early recurrent events.

Cardiovascular disease is the leading cause of death worldwide. Advanced insights into disease mechanisms and strategies to improve therapeutic opportunities require deeper understanding of the molecular processes of the normal heart. Knowledge of the full repertoire of cardiac cells and their gene expression profiles is a fundamental first step in this endeavor. Here, using large-scale single cell and nuclei transcriptomic profiling together with state-of-the-art analytical techniques, we characterise the adult human heart cellular landscape covering six anatomical cardiac regions (left and right atria and ventricles, apex and interventricular septum). Our results highlight the cellular heterogeneity of cardiomyocytes, pericytes and fibroblasts, revealing distinct subsets in the atria and ventricles indicative of diverse developmental origins and specialized properties. Further we define the complexity of the cardiac vascular network which includes clusters of arterial, capillary, venous, lymphatic endothelial cells and an atrial-enriched population. By comparing cardiac cells to skeletal muscle and kidney, we identify cardiac tissue resident macrophage subsets with transcriptional signatures indicative of both inflammatory and reparative phenotypes. Further, inference of cell-cell interactions highlight a macrophage-fibroblast-cardiomyocyte network that differs between atria and ventricles, and compared to skeletal muscle. We expect this reference human cardiac cell atlas to advance mechanistic studies of heart homeostasis and disease.