Icosapent ethyl, a high-purity ethyl ester of eicosapentaenoic acid (EPA), has demonstrated cardiovascular benefit in clinical trials; however, its effects on post-myocardial infarction (MI) cardiac remodeling and the underlying cellular mechanisms remain incompletely understood. Here, we investigated the role of icosapent ethyl in regulating post-MI remodeling using a murine MI model with myeloid-specific deletion of the omega-3 fatty acid receptor GPR120. Icosapent ethyl treatment significantly improved survival, preserved left ventricular systolic function, attenuated adverse ventricular remodeling and interstitial fibrosis, reduced cardiomyocyte apoptosis, and preserved myocardial microvascular density after MI. The protective effects of icosapent ethyl were markedly attenuated in myeloid GPR120-deficient mice. Mechanistically, icosapent ethyl activated GPR120 signaling in myeloid cells, suppressed CXCL10 expression, and reduced recruitment of proinflammatory CXCR3[+] leukocytes into the infarcted myocardium. Pharmacological inhibition of CXCR3 rescued adverse cardiac remodeling in myeloid GPR120-deficient mice, supporting a functional link between myeloid GPR120 activation and CXCR3-dependent inflammatory responses. These findings identify the myeloid GPR120-CXCL10/CXCR3 signaling axis as a critical immunomodulatory pathway by which icosapent ethyl improves post-MI cardiac remodeling, highlighting therapeutic potential of icosapent ethyl to limit inflammation-driven cardiac dysfunction after ischemic injury.

Canonical Wnt/β-catenin pathway activation can promote cardiomyocyte (CM) cell cycle activity, but translation is limited by off-target and functional liabilities associated with global pathway activation. We identified two small molecules, LRN9 and DF6, which potentiate Wnt/β-catenin signaling through a non-GSK3β mechanism and induce CM cell-cycle re-entry in human iPSC-CMs. In cardiac organoids, the effects was largely restricted to CMs and were accompanied by minimal changes in contractile and relaxation parameters, in contrast to reported impairment with GSK3β inhibition. RNA-sequencing showed enrichment of DNA replication and cell-cycle pathways. In embryonic zebrafish, both compounds increased ventricular growth consistent with CM hyperplasia.

Myocardial ischemia/reperfusion (I/R) injury is a major cause of morbidity and mortality. Although both nuclear factor erythroid 2-related factor 2 (Nrf2) and O-GlcNAcylation play critical protective roles in myocardial I/R injury, it remains unclear the role of Nrf2 O-GlcNAcylation in cardiac injury following I/R. An in vivo I/R model in mice and an in vitro oxygen-glucose deprivation/reperfusion (OGD/R) model in AC16 cardiomyocytes were employed. Histology, serum markers, and oxidative stress assays were used to assess injury. Site-directed mutagenesis, co-immunoprecipitation, western blot, and qPCR were utilized to identify O-GlcNAcylation sites and investigate the functional consequences of Nrf2 mutations (S215A and S33A) on its localization, transcriptional activity, and cell injury. Nrf2 underwent time-dependent O-GlcNAcylation following myocardial I/R injury in vivo and in vitro. During the early phase (6 h) of OGD/R, O-GlcNAcylation at Ser215 promoted Nrf2 nuclear translocation, activated the expression of antioxidant genes (HO-1, NQO1, and SOD2), and conferred cyto-protection in AC16 cardiomyocytes. In contrast, during the later phase (24 h), O-GlcNAcylation at Ser33 inhibited Nrf2 nuclear accumulation, suppressed its transcriptional activity, and exacerbated oxidative injury. The S215A mutation abolished the early protective effects, whereas the S33A mutation significantly attenuated the late-phase damage. This study identified a novel O-GlcNAcylation switch at Nrf2 Ser215 and Ser33 that governs the shift from protection to injury in the myocardium following I/R, highlighting the therapeutic potential of site-specific O-GlcNAcylation for cardioprotection.

Limited proliferative capacity of cardiomyocytes (CMs) underlies persistent CMs loss and cardiac dysfunction after myocardial infarction (MI). Although neonatal mammalian hearts retain transient regenerative potential, the molecular mechanisms governing this process remain incompletely understood. Protein phosphatase 2A (PP2A) has been implicated in cardiac protection; however, its role in CMs proliferation and heart regeneration is unclear. Here, using neonatal mouse MI models, cardiomyocyte-specific PP2Acα knockdown mice, pharmacological inhibition with LB-100, adenoviral mediated gain and loss of function approaches, and transcriptomic analysis, we investigated the functional significance and molecular mechanism of PP2Acα in cardiac regeneration. We found that PP2Acα was highly expressed in neonatal hearts, declined with postnatal maturation, and was reinduced following neonatal MI. Cardiomyocyte-specific PP2Acα knockdown markedly reduced CMs proliferation, impaired neonatal heart regeneration, exacerbated post MI cardiac dysfunction, and increased fibrosis, while pharmacological inhibition of PP2A similarly suppressed CMs proliferation both in vivo and in vitro. In contrast, PP2Acα overexpression significantly enhanced mitotic activity in neonatal mouse cardiomyocytes. Mechanistically, co-immunoprecipitation and molecular docking analyses revealed that PP2Acα directly interacted with Yes-associated protein (YAP) and reduced its phosphorylation, thereby promoting YAP nuclear translocation and activation of cell cycle-related gene transcription. YAP knockdown abolishes PP2Acα-induced cardiomyocyte proliferation. Conversely, LB-100 increased YAP phosphorylation without altering its mRNA expression, indicating regulation dependent on phosphatase activity. Collectively, these findings identify PP2Acα as a critical phosphatase that promotes neonatal cardiac regeneration by directly dephosphorylating YAP and activating the Hippo-YAP signaling axis, uncovering a previously unrecognized regulatory mechanism controlling cardiomyocyte proliferation and highlighting a potential therapeutic target for cardiac repair.

Transmembrane protein 175 (TMEM175) is a lysosomal proton-activated and proton-selective channel critical for regulating lysosomal membrane potential and acidity. However, its role in cardiomyocyte physiological and stress response remains unclear. Here, we investigated the function of TMEM175 in mitochondrial homeostasis using H9c2 cardiomyocytes and neonatal rat ventricular myocytes (NRVMs) under physiological conditions and during hydrogen peroxide (H2O2)-induced oxidative stress. Under physiological conditions, genetic knockout of TMEM175 impaired mitochondrial respiration, reduced mitochondrial superoxide, and attenuated autophagic clearance. In contrast, under H2O2-induced stress, TMEM175 deletion significantly alleviated mitochondrial dysfunction and cell death, despite autophagic flux being primarily stalled at the degradation stage. Mechanistically, TMEM175 deficiency activated AMP-activated protein kinase (AMPK), and silencing AMPK reversed the cytoprotective effects of TMEM175 deletion against H2O2 injury. Pharmacological inhibition of TMEM175 with 2-phenylpyridin-4-ylamine (2-PPA) in H9c2 and NRVMs recapitulated key phenotypes observed in genetic knockout models. Furthermore, 2-PPA improved cardiac function and attenuated histopathological injury in myocardial infarction mice. Together, these findings reveal a dual role for TMEM175: it maintains lysosomal-mitochondrial communication under basal conditions, yet its inhibition protects against oxidative stress primarily through AMPK activation. This study identifies TMEM175 as a novel lysosomal regulator of cardiac mitochondrial resilience and highlights its potential role in the cellular response to oxidative injury in cardiomyocytes.

Fibrosis is a maladaptive process common to many diseases throughout the human body. During fibroblast activation, the cell undergoes metabolic reprogramming to increase glycolysis and support increased cellular growth. Alterations in metabolism have been shown to play a significant role in determining cell identity and function. However, the contribution of many ancillary metabolic pathways in cardiac fibroblasts remains poorly defined. Here we investigate the role of Gfpt2, the rate-limiting enzyme of the hexosamine biosynthesis pathway, and metabolism in fibroblast activation. We demonstrate loss of Gfpt2 in cardiac fibroblasts results in a shift towards an activated fibroblast phenotype suggesting Gfpt2 acts as a regulator of fibroblast activation. Furthermore, untargeted metabolomics identifies glutathione metabolism as a downstream pathway and shows decreased glutathione following Gfpt2 loss. Consistent with this result, treatment with glutathione following either Gfpt2 knockdown or TGFβ stimulation is sufficient to prevent fibroblast activation. Finally, screening fibroblasts from other tissues illustrates that the Gfpt2-glutathione regulatory axis is shared across some, but not all, tested fibroblast populations. Overall, our study highlights the nuanced role of metabolism in cardiac fibroblasts and identifies a novel regulatory axis that could be a shared pathway in fibroblast activation.

Cardiac mitochondrial remodelling is a hallmark of type 2 diabetes-linked heart failure (T2DM-HF). We previously reported that mitochondrial morphological changes occur in early-stage disease and identified down-regulation of the mitochondrial protein Miro1 (Rhot1). Neuronal Miro1 regulates mitochondrial movement but the role of cardiac Miro1 remains poorly understood. Therefore, we generated a cardiac-specific Miro1 knockout (Miro1[cko]) mouse model to investigate how cardiomyocyte-Miro1 deficiency affects cardiac and mitochondrial structure-function. Miro1[cko] mice compared to controls develop mild diastolic and systolic dysfunction and electrical abnormalities, cellular hypertrophy and fibrosis. Miro1[cko] leads to aberrant mitochondrial respiration and elevated H2O2 production, consistent with electron microscopy showing disrupted cristae morphology, with putative links to Myosin19 down-regulation. Three-dimensional electron microscopy identified mitochondrial remodelling with interfibrillar mitochondria (IFM) ∼50% smaller with an increased surface complexity. Since fusion-fission protein expression was unchanged these data identify Miro1 as a regulator of mitochondrial morphology. Mitochondrial density increases (34% Miro1[cko]; 30% control), with abnormal IFM clustering, which we suggest is associated with impaired mitophagy since PINK1 and Parkin are down-regulated (∼80% and ∼ 60% respectively) and imaging flow cytometry of isolated primary cardiomyocytes identified an ∼2-fold reduction to mitochondrial clearance indicative of blunted mitophagy. Heterozygous knockout mice, which display a milder cardiac phenotype, rapidly developed HF symptoms when given a high fat diet with L-NAME. In conclusion, loss of Miro1 drives multiple aberrant mitochondrial remodelling events culminating in cardiac dysfunction and predisposes towards accelerated metabolic-HF development. Loss of Miro1 may represent a critical mechanistic link in T2DM-HF pathogenesis and therefore a potential therapeutic target.

Heart failure with preserved ejection fraction (HFpEF) predominantly affects older women. The widely used two-hit model of HFpEF has mostly been applied to young animals and fails to induce HFpEF in female mice, limiting clinical and epidemiologic relevance. We challenged 19-month-old female mice with the two-hit protocol. Unlike young female mice, aged mice developed HFpEF, coinciding with impaired cardiac mitochondrial function and disrupted mitochondrial proteostasis. Our findings connect aging with increased female susceptibility to cardiometabolic stress and demonstrate the contribution of mitochondrial dysfunction in HFpEF. Incorporating aging into the two-hit model enables future investigation of sex-specific mechanisms of HFpEF.

Hypertrophic cardiomyopathy (HCM) is a condition characterized by variable patterns of myocardial hypertrophy. Progressive functional remodeling of material properties contributes to symptoms, disease progression, and variability in treatment response. However, it is not clear how these properties interact to determine cardiac performance and response to treatment. We quantified how cardiac anatomy variability, functional remodeling, and pharmacological treatment influence the relative importance of biophysical mechanisms that govern cardiac function. We built five four-chamber electromechanical heart models representing anatomical clusters. We performed global sensitivity analyses on 46 parameters across 32 outputs. We repeated this process 18 times to model different types of functional remodeling and two times to model mavacamten and aficamten. Across all representative HCM phenotypes, sensitivity profiles were preserved. Within the parameter ranges examined, ventricular afterload explained the largest share of variance in hemodynamic outputs across anatomies (up to 69%). Functional remodeling led to shifts in parameter importance, mostly when ventricular stiffness was increased. The modeling of pharmacological treatments led to modest but targeted changes, with mavacamten and aficamten producing comparable effects, particularly for outputs related to arterial pressures. Across major anatomical phenotypes of hypertrophic cardiomyopathy, functional remodeling, rather than anatomy alone, reshaped the relative importance of cellular, myocardial, and loading-related parameters, with the largest shifts under increased ventricular stiffness and under aficamten in obstructive physiology. These hypothesis-generating results suggest that the integration of biomarkers reflecting ventricular-arterial coupling, myocardial stiffness, and contractile state may help anticipate symptoms and treatment response in HCM.

Increased cardiac risk in diabetes has been linked to disturbances in myocardial metabolism. Circulating and cardiac fructose levels are elevated in diabetes but the relationship between fructose and cardiac pathology is unclear. The goal of this study was to assess myocardial capacity for fructose metabolism and evaluate the time-course of cardiac fructose accumulation relative to the emergence of cardiac functional impairment in diabetic rats. Cardiac capability for fructose metabolism to support function was demonstrated in ex vivo working mouse hearts perfused with 11 mM fructose. Using isotope-labeled fructose [U[13]C], we observed [13]C enrichment into downstream metabolites glyceraldehyde, glycerate, pyruvate, lactate, and mitochondrial acetyl-CoA in perfused working mouse hearts. Metabolite profiling demonstrated that relative to glucose, myocardial fructose metabolism favored glycerate production. In diabetic rats (streptozotocin, 55 mg/kg), cardiac fructose elevation was evident prior to the onset of cardiac dysfunction. This study provides proof-of-principle evidence that fructose metabolism is operational in the working heart and identifies key fructose-derived metabolites. The finding that cardiac fructose elevation precedes functional impairment supports the contention that fructose may be an early instigator of diabetic cardiomyopathy and further investigation is now warranted. NEW AND NOTEWORTHY (<75 WORDS): Circulating and cardiac fructose levels are elevated in diabetes but the relationship between fructose and cardiac pathology is unclear. This study provides proof-of-principle evidence that fructose metabolism is operational in the working heart and identifies that cardiac fructose metabolism favors production of glycerate. In diabetes, cardiac fructose elevation precedes functional impairment supporting the contention that fructose may be an early instigator of diabetic cardiomyopathy.

Biallelic titin truncation variants (TTNtvs) are linked to severe cardiac and skeletal muscle diseases, due to unclear mechanisms. Using induced pluripotent stem cell-derived cardiomyocytes from a biallelic TTNtv patient with dilated cardiomyopathy, we investigated sarcomere structure/function. Only the longest of the TTNtvs was detected as protein, and this nearly full-length titin was incorporated into the sarcomere. Subtle structural alterations occurred, with shortened A-bands observed in a subset of sarcomeres. Resulting reduction and imbalance of force development was linked to lowered contractility, and many sarcomeres being stretched by their neighbors. Thus, inefficient sarcomere assembly and interaction promotes cardiomyopathy in biallelic TTNtv.

Extracellular signal-regulated kinases 1 and 2 (ERK1/2) are central regulators of cardiovascular development and stress responses and have been implicated in the cardiovascular toxicity of tyrosine kinase inhibitors. While ERK signaling in cardiomyocytes has been extensively investigated, the isoform- and cell-specific contribution of ERK2 in vascular smooth muscle cells (VSMCs) to cardiac dysfunction remains incompletely understood. To address this issue, we generated mice with systemic deletion of Erk1 (Erk1[-][/][-]), SM22α-Cre-mediated deletion of Erk2 (SM22α-E2KO; targeting VSMCs and cardiomyocytes), and α-myosin heavy chain-Cre-mediated deletion of Erk2 in cardiomyocytes alone (MHCα-E2KO). SM22α-E2KO mice developed progressive eccentric left ventricular hypertrophy, systolic dysfunction, impaired exercise capacity, and markedly reduced survival, whereas MHCα-E2KO and Erk1[-][/][-] mice exhibited preserved cardiac structure and function under basal conditions. Vascular functional studies revealed enhanced phenylephrine-induced vasoconstriction and impaired acetylcholine-mediated relaxation in SM22α-E2KO mice, despite preserved ERK2 expression in aortic endothelial cells. Aortic RhoA/Rho-kinase activity was significantly increased in SM22α-E2KO mice, and pharmacological inhibition of Rho-kinase normalized vascular hypercontractility. Transcriptomic analyses of the heart demonstrated downregulation of excitation-contraction coupling and metabolic pathways, accompanied by activation of inflammatory and fibrotic signaling. Notably, partial loss of Erk1 further exacerbated mortality and cardiac dysfunction in SM22α-E2KO mice, indicating a compensatory role of ERK1 in the setting of ERK2 deficiency. Collectively, these findings identify ERK2 signaling in VSMCs as a critical determinant of vascular tone and cardiac systolic function and demonstrate that combined vascular and myocardial ERK2 deficiency promotes heart failure through Rho-kinase-dependent vascular dysfunction.

Cardiac myosin binding protein-C (cMyBP-C) is a key regulator of sarcomere structure and contractile function that modulates actomyosin interactions to control the speed and strength of cardiac muscle contraction and relaxation. As an integral part of the sarcomeric machinery, this modular protein has been extensively studied; however, the regulatory contributions of its middle and C-terminal regions remain incompletely understood. This review highlights the structural organization and emerging roles of key cMyBP-C domains, including multiple phosphorylation sites in the N-terminal M-domain, the flexible C4-C5 linker, the cardiac-specific C5 loop, and the extended linker between domains C9 and C10. We discuss how N-terminal domains interact with actin and myosin subfragments (S1/S2) to influence contractile dynamics, particularly through phosphorylation-dependent modulation of actomyosin interactions. We further examine the middle domains (C4-C6), which contain the C4-C5 linker and the C5 loop, originally proposed to function primarily as structural spacers but are now increasingly implicated in regulating cMyBP-C function. The C-terminal domains, particularly C7-C10, are essential for anchoring cMyBP-C to the thick filament's C-zone, though additional functional roles remain to be uncovered. Post-translational modifications, including phosphorylation, S-glutathionylation, nitrosylation, and acetylation, are discussed in the context of cardiac stress and disease, while recognizing that their physiological significance and potential interplay remain incompletely understood. Finally, we highlight the current limitations in visualizing dynamic cMyBP-C interactions and emphasize the need for integrative structural, biochemical, and in vivo studies to fully elucidate the regulatory network orchestrated by this multi-domain sarcomeric regulator. Improved mechanistic understanding may ultimately reveal therapeutic opportunities for cardiomyopathies associated with mutations in MYBPC3 and dysregulated cMyBP-C signaling.