Abstract
The endocardium is a major source of coronary angiogenesis and arterialization, through coordinated cell fate transition and migration. However, the transcriptional regulatory network synchronizing cell fate determination and movement remains unclear. Here, we identified transcription factor HAND2 as a key candidate for coronary vascular formation. Endocardial deletion of Hand2 in mice disrupted arterial-venous networks and stunted coronary arteries, paralleling a ventricular noncompaction phenotype. Moreover, deletion of Hand2 produced excessive tip cells with defective movement. RNA-seq analysis revealed enhanced hypoxic metabolic activation but declined TGF-β/p38MAPK-dependent endothelial-to-mesenchymal transition (Endo-MT). In consistence, genetic inhibition of the core hypoxic regulators or pharmaceutical administration of TGFβ2 partially recovered the coronary arterial defects in Hand2 mutants. Furthermore, HAND2 was found directly bound to promoters of the target genes, harmonizing cell migration and cell fate transition. These findings pinpoint HAND2 as an essential regulator of the endocardial transcriptional regulatory network for coronary arterialization and provide potential therapeutic targets for coronary artery diseases.
Authors
Huijuan Wang, Haosheng Zhang, Leiyin Zheng, Peihan Zhang, Yuqian Wang, Sijia Ding, Wenping Liu, Yuanming Cheng, Zhongzhou Yang, Wen Luo
Abstract
Myofibroblasts are the cells responsible for collagen production, leading to tissue fibrosis. Because 20.5% of the total amino acids in collagen are proline, myofibroblasts must acquire a well-developed proline-producing mechanism during their differentiation. However, the detailed mechanism for myofibroblasts to acquire and keep the developed proline biosynthesis machinery remains obscure. Here, we show branched-chain amino acid transaminase 1 (Bcat1) is up-regulated in a substantial subset of Postn-expressing proto-myofibroblast-like fibroblasts, transitional cells en route to fully differentiated myofibroblasts, as well as in myofibroblasts in the fibrotic heart and liver of mice and humans and promotes the proline production. The branched-chain amino acid (BCAA) production by BCAT1 promotes SMAD3 phosphorylation via HDAC5 phosphorylation at Ser488, thereby enhancing SMAD3-dependent transcription of proline biosynthesis-related genes, Aldh18a1, Pycr1, and Eprs, in proto-myofibroblast-like fibroblasts and myofibroblasts. In BCAT1-deficient mice, expression of proline biosynthesis-related genes is significantly attenuated in their hearts after myocardial infarction, resulting in decreased cardiac fibrosis. Moreover, BCAT1 inhibitor treatment of mice with myocardial infarction reduces cardiac fibrosis. Our results identified a BCAT1-mediated pathway that promotes collagen production via proline biosynthesis regulation in proto-myofibroblast-like fibroblasts and myofibroblasts, which may provide a therapeutic target for cardiac fibrosis.
Authors
Noburo Takizawa, Takanori Hironaka, Hayato Watanabe, Haruna Suetsugu, Keisuke Yoshioka, Yuma Horii, Yuri Nagata, Hiroaki Matoba, Hidetaka Kosako, Kenji Hamase, Go Hirai, Michio Nakaya
Abstract
Gut microbiota-derived trimethylamine N-oxide (TMAO) plays a role in the pathogenesis of cardiovascular disease. The role of TMAO in the pathogenesis of atrial fibrillation (AF) remains uncertain. TMAO levels were quantified in plasma from serial subjects undergoing elective cardiac catheterizations (N=5090) and shown to independently associate with prevalent AF following adjustment for risk factors (TMAO adjusted odds ratio 1.7 [95% confidence interval 1.3-2.1]; P<0.01). Human cAMP response element modulator isoform IbΔC-X transgenic mice (CREM-IbΔC-X), a spontaneous mouse model of AF, supplemented with a TMAO diet developed AF sooner. C57BL/6J mice on and off a TMAO had more inducible AF via a transesophageal pacing study compared to chow controls. Dietary choline supplementation increased circulating TMAO levels and significantly accelerated AF onset in CREM-IbΔC-X mice (P<0.01). Iodomethylcholine (IMC), the gut microbial CutC/D inhibitor that suppresses choline→TMA(O) metabolic transformation, reduced circulating TMAO levels (P<0.0001) and choline induced AF onset (P<0.01). Cecal metagenomic analyses showed that choline supplementation induced changes in microbial communities associated with AF, while many of these changes were attenuated by IMC. Choline supplementation promoted overall adverse atrial remodeling with left atrial dilation. Optical mapping studies showed that mice supplemented with choline exhibited reduced conduction velocity, shortened action potential duration at 80% repolarization, and decreased wavelength. TMAO inhibits muscarinic receptor 2 resulting in autonomic dysfunction that promotes AF. In summary, the gut microbial metabolite TMAO, independently associated with AF risk in subjects, enhances AF in multiple AF mouse models via autonomic dysfunction, and is a therapeutic target for prevention of AF.
Authors
Selvam Arjunan, Isaiah Pemberton, Xinmin S. Li, Naseer Sangwan, Lydia Akino, Emmanuel Opoku, Dmitriy Verbovetskiy, Ina Nemet, Hyun Su Kim, Haruko Masumiya, Zeneng Wang, Joseph A. Lupica, Melissa Y. Tian, Karis Mao, Deepthi P. Mallela, Maradumane Mohan, Sarah Schumacher, Julie H. Rennison, Sathyamangla Prasad, Kenneth R. Laurita, Vamsi Chodisetty, Mina K. Chung, David R. Van Wagoner, John Barnard, Jonathan D. Smith, Oussama Wazni, Stanley L. Hazen, Robert A. Koeth
Abstract
Inflammation contributes to the pathogenesis of myocardial infarction and heart failure and represents a viable therapeutic target. Monocytes and their progeny are highly abundant and display striking functional diversity, serving as key determinants of myocardial inflammation and tissue repair. Much remains to be learned regarding mechanisms and signaling events that instruct monocyte fate decisions. We devised a genetic lineage tracing strategy using Ccr2crERT2Rosa26LSL-tdTomato mice in combination with single cell RNA-sequencing to map the differentiation trajectories of monocytes that infiltrate the heart after reperfused myocardial infarction. Monocytes were recruited to the heart early after injury and gave rise to transcriptionally distinct and spatially restricted macrophage and dendritic cell-like subsets that were specified prior to extravasation and chronically persisted within the myocardium. Pseudotime analysis predicted two differentiation trajectories of monocyte-derived macrophages that are partitioned into the border and infarct zones, respectively. Among these trajectories, we demonstrated that macrophages expressing a type I interferon responsive signature were an intermediate population that gave rise to MHC-IIhi macrophages, were localized within the border zone, induce regulatory T cells, and promote myocardial protection. Collectively, these data uncover complexities of monocyte differentiation in the infarcted heart and suggest that modulating monocyte fate decisions may have clinical implications.
Authors
Andrew L. Koenig, Farid F. Kadyrov, Junedh M. Amrute, Steven Yang, Carla J. Weinheimer, Jessica M. Nigro, Attila Kovacs, Wenjun Li, Gabriella B. Smith, Lance Yeh, Daniel Kreisel, Kory J. Lavine
Abstract
Gene therapy-based biological pacemakers have been proposed as an alternative to their hardware-based counterparts. In this context, short-term ectopic expression of the T-box transcription factor 18 (TBX18) in the ventricle has been reported to generate potent short-term pacemaker function in various animal models. Here, we investigated the impact of adeno-associated virus (AAV)-mediated long-term expression of TBX18, and compared the outcomes to those of the pacemaker ion channel Hcn2. Our findings revealed that CMV-driven ectopic TBX18 expression in mouse hearts led to severe cardiac fibrosis. At lower, non-fibrogenic levels, TBX18 maintained its transcriptional function but failed to induce pacemaker phenotypes. TBX18-expressing cells showed suppressed expression of key working myocardial genes, but the pacemaker gene program was not induced. Electrophysiological studies showed abnormal automaticity in TBX18-expressing cells, combined with prolonged repolarization and various current changes. However, no hyperpolarization-activated funny current was detected. In a complete AV-block rat model, AAV-mediated Hcn2 expression induced robust ectopic pacemaker activity in the presence of isoproterenol, whereas TBX18 expression neither generated such activity nor augmented Hcn2-mediated pacing. In conclusion, at functionally non-fibrogenic levels, TBX18 is neither sufficient nor necessary to induce pacemaker activity. In contrast, Hcn2 generates reliable pacing, making it a more viable candidate for biological pacemaker development.
Authors
Jianan Wang, Mathilde R. Rivaud, Mischa Klerk, Arie R. Boender, Ruud N. Visser, Rinske Sparrius, Hee Young Lee, Karel van Duijvenboden, Huiling Zhou, Yuting Yang, Emiel J.M. Kramer, Kyung Ho Park, Larry C. Park, Silke Schrödel, Christian Thirion, Eric Ehrke-Schulz, Anja Ehrhardt, Osne F. Kirzner, Klaus Neef, Hanno L. Tan, Arie O. Verkerk, Vincent M. Christoffels, Gerard J.J. Boink
Abstract
Enhanced TGFβ signaling caused by mutations in Fibrillin-1 (FBN1) in patients with Marfan syndrome (MFS) leads to myxomatous degeneration of the mitral valve (MDMV). MDMV can result in mitral valve prolapse, severe regurgitation, and sudden cardiac death. However, it remains unknown whether lymphatic vessel (LV) dysfunction contributes to MDMV development in MFS. Here, we show that lymphangiogenesis in murine mitral valves (MVs) begins postnatally. However, this process is inhibited in a mouse MFS model, Fbn1 mutant (Fbn1C1039G/+) mice, accompanied by disrupted lymphatic cell-cell junctions, impaired lymphatic drainage, and an abnormally widespread distribution of MHCII+ infiltrating macrophages. Treatment of Fbn1 mutant mice with VEGF-C156S, a selective VEGFR3 agonist, stimulates the ERK and Akt pathways, increases LV density in MVs, and ameliorates MDMV. Fbn1 mutant MVs display disorganized valvular endothelial cells (VECs) and decreased expression of the anti-inflammatory modulator Zfp36 (zinc finger protein 36) in VECs and immune cells. Treatment with FTY720 (Fingolimod), a ZFP36 activator and S1P antagonist, rescues MDMV phenotypes in Fbn1 mutant mice by reducing immune cell infiltration and restoring lymphatic cell junctions and drainage. These findings suggest that the Fbn1 mutation causes LV hypoplasia and defective lymphatic drainage in MVs, driven in part by pro-inflammatory VECs, leading to MFS-related MDMV.
Authors
Can Tan, Ziyou Ren, Shreya Kurup, Xianpeng Liu, Zhi-Dong Ge, Shodai Suzuki, Pritika Jakka, Cheryl Tang, M. Luisa Iruela-Arispe, Tsutomu Kume
Abstract
Cardiac macrophages are broadly studied as two subtypes, tissue resident C-X3-C motif chemokine receptor 1 positive (CX3CR1+) that are also C-C motif chemokine receptor 2 negative (CCR2–), and monocyte derived CCR2+. Previous systemic loss of function approaches suggested unique roles for each subtype in the heart with CCR2+ being inflammatory and CX3CR1+ being pro-healing. Here we employed a cardiac-specific gain of function approach to selectively enhance either macrophage subtype. A robust increase in basal CCR2+ macrophages in the heart by targeted C-C motif chemokine ligand 2 (Ccl2) expression did not induce inflammation, cause fibroblast activation, or impair cardiac function. However, increased CCR2+ macrophages reciprocally diminished self-renewing tissue resident macrophages and worsened cardiac fibrosis due to pressure overload stimulation. Conversely, augmented expression of colony-stimulating factor-1 (Csf1) in the heart promoted selective expansion of resident CX3CR1+ macrophages, which exerted no pathophysiological consequences at steady-state. However, pressure overload in these mice with expanded CX3CR1+ macrophages showed a CCR2+ macrophage-dependent inflammation leading to exacerbated cardiac dysfunction, simultaneously still protecting from adverse remodeling and cardiac fibrosis. In conclusion, cardiac-specific selective enrichment of macrophage subtypes shows their intricate interplay and unique functional roles in regulating myocardial inflammation and fibrosis during hypertrophy and at homeostasis.
Authors
Rajesh K. Kasam, Ronald J. Vagnozzi, Yasuhide Kuwabara, Anne Katrine Z. Johansen, N. Scott Blair, Vikram Prasad, Suh-Chin J. Lin, Akanksha Rajput, Michelle Nieman, Jeffery D. Molkentin
Abstract
Dilated cardiomyopathy (DCM) is a genetically heterogeneous disorder, characterized by ventricular dilatation and impaired systolic function, leading to heart failure and sudden cardiac death. Despite advances in genomic technologies, the genetic cause of DCM remains unidentified in more than half of the cases. Here, we performed an integrative analysis of genomic and transcriptomic data from patient-derived cardiac tissue to identify causative variants in genetically undiagnosed DCM. This approach enabled us to identify a homozygous splice-site variant (c.243+6T>A) in the sarcoglycan gene SGCB, which results in exon 2 skipping. This variant was significantly enriched in patients with DCM compared with the general population, with consistent genotype–phenotype correlations observed across multiple families. Protein-level analysis of cardiac tissue from homozygous individuals revealed loss of β-sarcoglycan, the protein product of SGCB, and destabilization of the sarcoglycan complex. Although SGCB has been previously associated with limb-girdle muscular dystrophy, these homozygous individuals showed no biochemical or clinical signs of skeletal muscle involvement, indicating an absence of myopathy. Compared with variant-negative patients with DCM, homozygous individuals also had a higher risk of early-onset adverse cardiac events. Together, these findings identify c.243+6T>A in SGCB as a cause of isolated DCM associated with unfavorable clinical outcomes.
Authors
Fangfang Li, Haruki Shinomiya, Yuki Kuramoto, Koshiro Kanaoka, Yuji Sakahashi, Yasuki Ishihara, Hidetaka Kioka, Seiko Ide, Yumi Yamaguchi-Kabata, Shu Tadaka, Ikuko N. Motoike, Kengo Kinoshita, Kinuko Ohneda, Hidetoshi Sakurai, Takahiro Okumura, Yohei Miyashita, Kota Jojima, Hisakazu Kato, Ken Matsuoka, Kazuya Tanabe, Shunsuke Nishimura, Seiji Takashima, Yoshihiro Asano, Yasushi Sakata
Abstract
BACKGROUND. Right ventricular failure (RVF) is a major determinant of mortality in pulmonary arterial hypertension (PAH), and hepatic dysfunction predicts adverse outcomes. However, the cell-specific effects of PAH/RVF on the human liver remain poorly defined. METHODS. We performed single-nucleus RNA sequencing of autopsy-derived liver tissue from 5 PAH patients and 4 non-PAH controls and compared these findings with publicly available single-nucleus RNA sequencing datasets from non-alcoholic steatohepatitis (NASH) and Fontan-associated liver disease (FALD). Transcriptomic analyses were integrated with histologic assessment, mitochondrial-enriched proteomics, and correlated with clinical markers of PAH/RVF severity. RESULTS. PAH livers showed cell-specific metabolic, inflammatory, and fibrotic remodeling distinct from NASH and FALD. PAH hepatocytes exhibited a hypoxia-adapted, Warburg-like metabolic phenotype with reduced fatty acid metabolism, gluconeogenesis, cytochrome P450 activity, and ketone metabolism. PAH endothelial cells demonstrated increased glycolytic pathway activity and disrupted adhesion/barrier signaling. PAH hepatic stellate cells displayed HIF-1 and PI3K-Akt pathway activation, and increased IL6 expression, which resulted in central vein fibrotic remodeling. PAH macrophages showed complement activation with reduced JAK-STAT signaling. Finally, HSC HIF-1 activity correlated with clinical markers of PAH/RVF severity. CONCLUSION. PAH induces a distinct metabolic and inflammatory hepatopathy characterized by hepatocyte metabolic reprogramming, HSC activation, and macrophage complement signaling. These findings support PAH-associated hepatopathy as a disease-specific end-organ phenotype linked to RVF severity.
Authors
Madelyn J. Blake, Sally E. Prins, Jeffrey C. Blake, Lynn M. Hartweck, Jenna B. Mendelson, Steeve Provencher, Sandra Breuils-Bonnet, Sebastien Bonnet, Kurt W. Prins
Abstract
Authors
Ryan A. Moon, Rafael Sobrano Fais, Minwoo Kim, Neal T. Vogel, Jenna B. Mendelson, Lynn M. Hartweck, John P. Carney, Melissa K. Gardner, Sally E. Prins, Sasha Z. Prisco, Tim Lahm, Kurt W. Prins
Copyright © 2026 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)