Abstract
Haploinsufficiency of TBX1, which occurs in 22q11.2 deletion syndrome (22q11.2DS), leads to a heterogeneous spectrum of clinical manifestations, including craniofacial anomalies, immunodeficiency, and congenital heart defects. The variability in syndromic presentation between patients may be partially explained by variants in chromatin regulatory genes that act to further modify TBX1 function. To investigate this relationship, we selected KMT2D as a candidate gene because of its role in the etiology of Kabuki syndrome, which shares overlapping features with 22q11.2DS. We demonstrate that conditional inactivation of Kmt2d in the Tbx1 lineage in Tbx1-heterozygous mice leads to fully penetrant perinatal lethality and increased incidence of craniofacial dysmorphism, thymus and parathyroid gland hypoplasia, and aortic arch anomalies. At early stages, mutant embryos were found to have defects of the caudal pharyngeal apparatus, including abnormal patterning of the third pouch endoderm, hypoplastic fourth arches, and defective fourth arch arteries. Finally, analysis of single-cell RNA sequencing revealed dysregulation, and largely downregulation, of genes involved in basic cellular functions, suggesting that Tbx1 and Kmt2d developmentally converge upon essential biological processes. Overall, these results indicate that reduced dosage of Kmt2d perturbs the developmental landscape of the Tbx1 heterozygote, eliciting phenotypes that are shared between 22q11.2DS and Kabuki syndrome.
Authors
Daniella Miller, Kevyn Jackson, Timothy C. Cox, Bernice E. Morrow
Abstract
Atrial fibrillation (AF) contributes to cardiovascular morbidity and mortality. Ubiquitin-specific peptidase 10 (USP10) plays a crucial role in numerous cellular processes; however, its particular role in AF remains largely unexplored. In the present study, USP10 expression was assessed in human atrial samples and angiotensin II–treated (Ang II–treated) mouse atrial tissues. An Ang II–induced AF mouse model was employed to investigate the effects of USP10 on atrial remodeling and AF susceptibility. Calcium imaging and patch clamp techniques were used to evaluate USP10’s influence on calcium handling and triggered activity. Additionally, RNA sequencing, coimmunoprecipitation, and ubiquitination assays were performed to explore the regulatory interactions between USP10 and NADH:ubiquinone oxidoreductase subunit S1 (NDUFS1). Our findings demonstrate that USP10 is downregulated in atrial tissues from mouse models and patients with AF. USP10 overexpression counteracts Ang II–induced atrial remodeling and reduces AF susceptibility. Furthermore, USP10 contributes to the restoration of mitochondrial function in AF. Mechanistically, USP10 deubiquitinates NDUFS1 at lysine 621, stabilizing NDUFS1 protein levels and mitigating Ang II–induced mitochondrial dysfunction. This study uncovers a critical mechanistic link between USP10 and NDUFS1. Our findings suggest that upregulating USP10 or targeting NDUFS1 degradation could provide an alternative therapeutic strategy to mitigate AF progression and associated cardiovascular risk.
Authors
Wanrong Fu, Xiao-Xu Tian, Jianghua Zhou, Yu-Xu Huang, Huan Li, Zhenya Wang, Tong-You Wade Wei, Li Li, Guo-Jun Zhao
Abstract
A single-nucleotide missense polymorphism (rs1800449, R158Q) in the propeptide domain of lysyl oxidase (LOX-PP) is associated with increased risk of coronary artery disease (CAD) independent of changes in plasma lipid levels. Although the enzymatic function of LOX has an essential role for the cross-linking of extracellular matrix proteins in connective tissues, whether and how LOX-PP R158Q contributes to the development of atherosclerosis has not been clearly established. Here, hypercholesterolemia was induced in mice that were WT or homozygous for the LOX-PP R158Q polymorphism by adeno-associated virus-8–mediated overexpression of Pcsk9 followed by high-fat diet feeding for 16 weeks. We found that the R158Q polymorphism promoted atherosclerosis and induced proliferation of macrophages and vascular smooth muscle cells without altering LOX enzymatic activity. Using single-cell RNA sequencing, we found the transcriptional program of atherosclerotic plaques from mice harboring R158Q was strongly enriched for proliferation- and calcification-related genes in a regionally distinct manner. Together, these results establish an enzymatically independent proatherogenic role for the LOX-PP and suggest its potential as a novel therapeutic target.
Authors
In-Hyuk Jung, Junedh M. Amrute, Sofia E. Luna, Ryan E. Wagoner, Arturo Alisio, Paul C. Lee, Kendall H. Burks, Joohee Oh, Hannah C. Plunkett Paletta, Chul Joo Kang, Nathan O. Stitziel
Abstract
The pericardium plays an important homeostatic function for the neighbouring heart providing both lubricating and structural support. In vivo models have further identified a protective role for the pericardium in modulating cardiac remodelling following myocardial infarction possibly through the actions of tissue-resident pericardial macrophages. Using patient derived pericardial samples, we establish that human pericardial immune cells directly inhibit cardiac fibroblast fibrotic activity and this action is dampened following myocardial infarction. Performing single-cell RNA sequencing of patient pericardial fluid cells, we identify two pericardial macrophage subsets that are uniquely altered in response to myocardial infarction, which contributes to a shift in their effector molecule expression profiles. We confirm that fibronectin-expressing human pericardial macrophages are the primary driver of the pericardial anti-fibrotic actions through the release of cystatin C. Finally, we establish cystatin C as a macrophage-derived cardioprotective effector molecule in an in vivo model of myocardial infarction. Collectively, we uncover a new molecular mechanism of the local immune environment that regulates cardiac remodelling post myocardial infarction.
Authors
Ali Fatehi Hassanabad, Sarthak Sinha, Arzina Jaffer, Darrell Belke, Nicole L. Rosin, Elodie Labit, Daniel Young, Friederike I. Schoettler, Keerthana Chockalingam, Benjamin Haeyul Lee, Jameson A. Dundas, Emilie de Chantal, Carmina A. Isidoro, Alexander Tam, Hanjoo B. Shim, Anna N. Zarzycki, Afshin Derakhshani, Elisabeth Gorgiogianni, Jeannine D. Turnbull, Antoine Dufour, Shalina S. Ousman, Jeff A. Biernaskie, Paul W.M. Fedak, Justin F. Deniset
Abstract
Pulmonary Arterial Hypertension (PAH) is a rare vascular disorder characterized by elevated pressure in pulmonary arteries, eventually leading to right ventricular failure. Approximately 50% of pediatric disease and 20% of adult disease can be linked to a genetic mutation, with nearly 70% of these cases involving mutations in the bone morphogenetic protein receptor type 2 (BMPR2) locus. Investigations using rodent models have made substantial advances in our understanding of BMPR2 signaling; however, limited data exist regarding the onset and course of PAH, and etiologies for phenotypic expression in these patients remain unknown. In this work, we describe the development of an ovine model of heritable PAH. Because homozygous disruption of BMPR2 is embryonic lethal, we developed heterozygous BMPR2 sheep by using a PAM-disrupting synonymous single stranded oligodeoxyribonucleotide alongside a single guide RNA and Cas9 mediated gene editing strategy. The resulting BMPR2(+/-) lambs demonstrated cardiac and pulmonary vascular pathology that are consistent with BMPR2 mutation-driven PAH observed in humans. Given the genetic and physiological similarities of BMPR2(+/-) sheep to humans with heritable PAH, this large animal model will serve as a vital platform for mechanistic molecular studies and will provide a much-needed pre-clinical model for extensive treatment evaluations.
Authors
Sanjeev A. Datar, Nicholas Werry, Austin R. Brown, Devon S. Fitzpatrick, Oluwafemi Falade, Josephine F. Trott, Rachel Hutchings, Elena K. Amin, Jessica M. Morgan, Hythem Nawaytou, Gail H. Deutsch, Eric G. Johnson, Omar A. Gonzales Viera, Thomas F. Bishop, Tara Urbano Beach, Bret R. McNabb, Eric D. Austin, Jeffrey R. Fineman, Alison L. Van Eenennaam
Abstract
Extracellular vesicles (EVs)-mediated inter-organ communication represents a promising frontier in transplant immunology; however, its role in cardiac allograft rejection remains poorly characterized. We performed proteomic profiling of plasma-derived EVs in a rat heterotopic heart transplantation model and identified a distinct liver-predominant protein signature during acute rejection, with Antithrombin III (ATIII) emerging as a top candidate. Functional validation revealed that pharmacological EV inhibition intensified systemic and intragraft inflammation, whereas adeno-associated virus (AAV)-mediated silencing of hepatic ATIII directly accelerated allograft rejection. Conversely, AAV-mediated hepatocyte-specific ATIII overexpression attenuated rejection pathology, reduced immune cell recruitment, and markedly prolonged median graft survival. This protective effect was achieved without evidence of coagulopathic complications, indicating an immunomodulatory mechanism beyond ATIII’s canonical anticoagulant function. Mechanistically, ATIII overexpression was associated with upregulation of heme oxygenase-1 (HO-1) in the liver and suppression of proinflammatory cytokine expression in the graft. These findings highlight hepatocyte-derived EVs as important mediators of a liver-heart signaling axis in transplant rejection, and further implicate the protein ATIII as a contributor to this axis. Our study reveals a therapeutically targetable liver-heart signaling axis in transplant rejection, whereby enhancing liver-derived ATIII or its downstream pathways (such as HO-1) could attenuate acute cardiac allograft rejection.
Authors
Shiyu Dai, Wei Zhou, Fangyu Chen, Huanyu Zhang, Zhenchun Ji, Xuejing Zong, Wanruo Zhang, Jie Hu, Shumin Jiang, Fei Wang, Zhenya Shen
Abstract
Adults with type 2 diabetes mellitus (T2DM) are at increased risk for stroke, myocardial infarction, and cardiovascular death, yet individual risk is heterogeneous and incompletely captured by clinical models. In the Exenatide Study of Cardiovascular Event Lowering (EXSCEL), adults with T2DM were randomized to a GLP-1 RA (exenatide) or placebo and followed longitudinally for major adverse cardiovascular events (MACE). High-throughoput discovery proteomics was done in plasma collected at baseline and 12-months. Proteins associated with time-to-MACE were identified using multivariable regression and incorporated into supervised machine learning models. A multi-protein score was developed and externally validated in two independent population-based and trial cohorts, Cardiovascular Health Study and the Prospective Multicentre Imaging Study for Evaluation of Chest Pain (PROMISE). The proteomic score showed incremental improvement in cardiovascular risk discrimination beyond clinical factors alone, and several proteins were consistently prioritized across modeling approaches. The protein score and a top-ranked protein, tetranectin, were modified by GLP-1 RA treatment, and a decrease in the protein score was associated with improved outcomes, supporting modifiability of MACE risk. External validation confirmed generalizability across cohorts with and without diabetes. Together, these findings demonstrate that plasma proteomic signatures can enhance cardiovascular risk stratification and identify treatment-responsive biomarkers in T2DM, supporting their potential role in precision prevention strategies.
Authors
Kristin M. Corey, Maggie Nguyen, Michael Y. Mi, Megan E. Ramaker, Ilya Zhbannikov, Harald Sourij, G. Michael Felker, Naveed Sattar, Jennifer B. Green, Pamela S. Douglas, Robert E. Gerszten, Robert J. Mentz, Adrian F. Hernandez, Rury R. Holman, Bruce M. Psaty, James S. Floyd, Svati H. Shah
Abstract
Pulmonary arterial hypertension (PAH) is a progressive vascular syndrome characterized by aberrant signaling, severe pulmonary artery remodeling, and right ventricular (RV) failure, a major driver of morbidity and mortality. Dysregulation of the apelinergic pathway has been implicated in pulmonary vascular remodeling in PAH. Using a sugen-hypoxia rat model of PAH, we assessed the ability of a novel apelin analog, resistant to native peptidase degradation, to reverse the pathological hallmarks of PAH and RV dysfunction. Apelin analog therapy corrected the vascular lesions in the lungs and nearly normalized pulmonary arterial pressures. Early cardiorenal syndrome, RV dilation and dysfunction as well as RV cardiomyocyte and fibroblast activation induced by pressure overload, were also reversed by apelin analog treatment. Single-nucleus RNA sequencing of the lungs and RV revealed apelin-analog treatment activated several protective pathways, including rebalancing protective BMPR2 (bone morphogenetic protein receptor type 2) signaling to counteract excessive pathogenic TGFBR2 (transforming growth factor β receptor 2) activity in PAH. These findings highlight the therapeutic potential of exogenous apelin in reversing pulmonary vascular and cardiac pathologies in PAH and support further investigation to evaluate the clinical benefits of apelin analog treatment in patients with PAH and RV failure.
Authors
Jennie Vu, Pavel Zhabyeyev, Kemar J. Brown, Joshua M. Gorham, Daniel M. DeLaughter, Huachen Chen, Thilina U. Jayawardena, Ander Vergara, Maria Alexiou, Anjalee Wijewardane, Conrad Fischer, Charlotte Avet, Abby Ewasiuk, Faqi Wang, Mark C. Chappell, Yuri Kim, Michel Bouvier, John C. Vederas, Christine E. Seidman, Jonathan G. Seidman, Gavin Y. Oudit
Abstract
Duchenne muscular dystrophy (DMD) is a lethal pediatric striated muscle disease caused by loss of dystrophin for which there is no cure. Cardiomyopathy is the leading cause of death amongst individuals with DMD, and effective therapeutics to treat DMD cardiomyopathy are a major unmet clinical need. This work investigated adeno-associated viral (AAV) gene therapy approaches to treat DMD cardiomyopathy by overexpression of the calcium binding proteins S100A1 and apoptosis repressor with caspase recruitment domains (ARC). Using the severe D2.mdx mouse model of DMD, we identified that S100A1 gene therapy improves the diastolic dysfunction associated with DMD cardiomyopathy, whereas ARC gene therapy prolongs survival. The combination of both S100A1 and ARC in a single bicistronic vector improves the long-term cardiac outcome and histopathology of D2.mdx mice, development of heart failure caused by micro-dystrophin expression, and exhibits safety via intracoronary delivery in a canine model of DMD. In addition to robust cardiac benefits, S100A1-ARC gene therapy benefits D2.mdx skeletal muscle function and histopathology when driven by a striated muscle promoter. Together, these findings indicate that S100A1-ARC gene therapy represents an effective treatment for DMD cardiomyopathy and may have therapeutic benefits in treating other forms of cardiomyopathy and muscle pathologies.
Authors
David W. Hammers, Cora C. Hart, Eli A. Zerpa, Karen I. Laurent, Young il Lee, Margaret M. Sleeper, H. Lee Sweeney
Abstract
The sodium-dependent multivitamin transporter, encoded by SLC5A6, mediates cellular uptake of biotin and pantothenic acid, essential cofactors for energy metabolism. We identified two families with SLC5A6 mutations presenting with early-onset dilated cardiomyopathy (DCM). To investigate the link between vitamin deficiency and cardiomyopathy, we generated a cardiac-specific SLC5A6 knockout (Slc5a6cKO) mouse model and evaluated the impact of vitamin supplementation. Slc5a6cKO mice developed progressive cardiac dysfunction, culminating in cardiac pathology and premature death at 26 weeks; earlier stages exhibited cardiomyocyte hypertrophy, fibrosis, impaired Coenzyme A synthesis, and metabolic imbalance, indicating progression toward cardiomyopathy. Cardiac magnetic resonance imaging and ECG confirmed progressive functional decline. Proteomic analysis revealed early mitochondrial metabolic disruption and extracellular matrix protein upregulation at 8 weeks, preceding overt cardiac dysfunction. Strikingly, vitamin supplementation from preconception onwards prevented the cardiac phenotype, preserving cardiac structure, function, morphology and survival. This paralleled the clinical outcome in one patient who received early vitamin treatment, compared to another who required a heart transplant without vitamin treatment. This study establishes a direct link between SLC5A6-mediated vitamin transport, mitochondrial function, and cardiac health. It highlights how vitamin deficiency contributes to cardiomyopathy pathogenesis and supports early vitamin supplementation as a potential therapeutic strategy for metabolic cardiomyopathies.
Authors
Millie O. Fullerton, Lauren C. Phillips, Rachael E. Redgrave, Luke Spray, Vincent Haufroid, George Merces, Scott T. Kerridge, Gavin D. Richardson, Nathalie Mercier, Dominique Roland, Rebecca Crossley, Andrew D.H. Morgan, Joseph P. Dewulf, John Burn, Simon D. Bamforth, Helen M. Phillips
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