Circulation Research

BACKGROUND: Pulmonary venous remodeling is a key pathological feature of pulmonary hypertension associated with left heart disease (PH-LHD). This study aims to investigate the role of regulatory T (Treg) cells in this process. METHODS: We used mouse models with transverse aortic constriction and cell depletion of Foxp3-DTR/tdTomato mice to examine Treg cells’ function around pulmonary veins in PH-LHD in vivo. To confirm the effect of Nlrc3−/− Treg cells on PH-LHD, we utilized 3 mouse models: Nlrc3 knockout mice, athymic mice, and endothelial cell lineage tracing Cdh5CreERT2+/−-mT/mG+/− mice. The interaction proteins and signaling pathways of Treg cells during endothelial-to-mesenchymal transition were elucidated by protein docking prediction, coimmunoprecipitation and cocultivation of Treg cells with venous endothelial cells. RESULTS: Treg cells were abundant around pulmonary veins of transverse aortic constriction–induced PH-LHD and were essential for promoting inflammation resolution and inhibiting pulmonary venous remodeling. Nlrc3 expression was reduced in mice and patients with PH-LHD. NLRC3 (nucleotide-oligomerization domain-like receptor family CARD domain containing 3) deficiency inhibited Treg cell proliferation and impaired their immunosuppressive and endothelial-to-mesenchymal transition–protective effects. Mechanistically, NLRC3 interacted with TRAM (TRIF-related adaptor molecule) and regulated interferon regulatory factor 3 (IRF3)/NF-κB (nuclear factor-κB) p65 signaling in cluster differentiation 4+ (CD4+) T cells. NLRC3-deficient Treg cells promoted interleukin (IL)-18 expression through IRF3/NF-κB p65 signaling, and thus IL-18 secretion activated endothelial receptor tyrosine kinase (RTK) signaling, favoring endothelial-to-mesenchymal transition progression in pulmonary veins and PH-LHD progress. This process was reversible with IL-18 binding protein in vivo. CONCLUSIONS: NLRC3 is crucial for Treg cells to prevent pulmonary venous remodeling in PH-LHD, primarily by modulating IL-18 secretion, which inhibits endothelial-to-mesenchymal transition and thereby improves disease progression and prognosis.

doi: 10.1161/circresaha.124.325310pmid: 40211947

BACKGROUND: Cardiometabolic heart failure with preserved ejection fraction (cHFpEF) is a highly prevalent and deadly condition. Histone 3 trimethylation at lysine 36 (H3k36me3)—a chromatin signature induced by the histone methyltransferase SETD2 (SET domain containing 2)—correlates with changes in gene expression in human failing hearts; however, its role remains poorly understood. This study investigates the role of SETD2 in cHFpEF. METHODS: Chromatin immunoprecipitation sequencing and RNA sequencing were used to investigate H3k36me3-related transcriptional regulation. Mice with cardiomyocyte-specific deletion of SETD2 (c-SETD2−/−) were generated and subjected to high-fat diet feeding and L-NAME treatment for 15 weeks to induce cHFpEF. Cardiac function and exercise tolerance were assessed by echocardiography and treadmill exhaustion test. A selective pharmacological inhibitor of SETD2, EZM0414, was also tested in cHFpEF mice. Mechanistic experiments were performed in cultured cardiomyocytes exposed to palmitic acid. SETD2 signaling and the effects of EZM0414 were also investigated in cardiomyocytes from patients with cHFpEF and control donors. RESULTS: SETD2 was upregulated in cHFpEF mouse hearts, and its chromatin mark H3k36me3 was involved in lipid metabolism and highly enriched on the promoter of the Srebf1 gene, encoding for SREBP1 (sterol regulatory binding protein 1). SETD2 activation in cHFpEF led to SREBP1 upregulation, triglyceride accumulation, and lipotoxic damage. Of note, cardiomyocyte-specific deletion of SETD2 in mice prevented heart failure with preserved ejection fraction–related hypertrophy, diastolic dysfunction, and lung congestion while improving exercise tolerance. SETD2 deletion blunted H3K36me3 enrichment on Srebf1 promoter, thus leading to a marked rewiring of the cardiac lipidome and restoration of autophagic flux. In vivo treatment with the SETD2 inhibitor EZM0414 recapitulated the effects of SETD2 deletion. Silencing of SETD2 in palmitic acid–treated cardiomyocytes prevented SREBP1 upregulation, whereas SETD2 overexpression mirrored lipotoxic damage. Finally, SETD2 was upregulated in left ventricle specimens from patients with cHFpEF while EZM0414 attenuated cardiomyocyte stiffness. CONCLUSIONS: Targeting SETD2 might prevent lipotoxic injury in cHFpEF.

doi: 10.1161/circresaha.125.326156pmid: 40207377

BACKGROUND: Mouse models of myocardial ischemia with subsequent heart failure are common approaches to examine heart failure pathology and possible treatment strategies. We sought to establish a high-throughput approach for echocardiography-guided induction of myocardial ischemia/reperfusion (IR) in mice. METHODS: After visualization of the left coronary artery with high-resolution ultrasound imaging and echocardiographic definition of the level of coronary occlusion, the left anterior descending artery was temporarily occluded with 2 micromanipulator-controlled needles. Functional and molecular changes were assessed and compared with commonly performed surgical techniques. RESULTS: Echocardiography-guided induction of myocardial IR enabled standardized induction of myocardial IR injury with subsequent left ventricular remodeling. Incorporation of various quality control measures throughout the procedure ensured a high success rate and the absence of relevant postinterventional mortality in experienced hands. Compared with surgical approaches, echocardiography-guided induction of myocardial IR showed a quicker recovery time and induced a less pronounced inflammatory response characterized by decreased local and systemic neutrophil counts. Notably, infarct size and subsequent post–myocardial infarction cardiac dysfunction were comparable between methods. The novel procedure was successfully implemented at different academic institutions with imaging expertise and demonstrated high interinstitutional reproducibility. CONCLUSIONS: Echocardiography-guided induction of myocardial IR enables high-throughput induction of myocardial IR injury with precise echocardiographic definition of the occlusion level and immediate evaluation of cardiac function during ischemia. The method provides a more clinically relevant assessment of IR sequelae and offers notable animal welfare advantages by eliminating the need for ventilation and thoracotomy, thereby mitigating potential surgery-related confounders.

BACKGROUND: Heart failure with preserved ejection fraction (HFpEF) has overtaken heart failure with reduced ejection fraction as the leading type of heart failure globally and is marked by high morbidity and mortality rates, yet with only a single approved pharmacotherapy: SGLT2i (sodium-glucose co-transporter 2 inhibitor). A prevailing theory for the mechanism underlying SGLT2i is nutrient deprivation signaling, of which ketogenesis is a hallmark. However, it is unclear whether the canonical ketogenic enzyme, HMGCS2 (3-hydroxy-3-methylglutaryl-coenzyme A synthase 2), plays any cardiac role in HFpEF pathogenesis or therapeutic response. METHODS: We used human myocardium, human HFpEF and heart failure with reduced ejection fraction transcardiac blood sampling, an established murine model of HFpEF, ex vivo Langendorff perfusion, stable isotope tracing in isolated cardiomyocytes, targeted metabolomics, proteomics, lipidomics, and a novel cardiomyocyte-specific conditional HMGCS2-deficient model that we generated. RESULTS: We demonstrate, for the first time, the intrinsic capacity of the human heart to produce ketones via HMGCS2. We found that increased acetylation of HMGCS2 led to a decrease in the enzyme’s specific activity. However, this was overcome by an increase in the steady-state levels of protein. Oxidized form of nicotinamide adenine dinucleotide repletion restored HMGCS2 function via deacetylation, increased fatty acid oxidation, and rescued cardiac function in HFpEF. Critically, using a conditional, cardiomyocyte-specific HMGCS2 knockdown murine model, we revealed that the oxidized form of nicotinamide adenine dinucleotide is unable to rescue HFpEF in the absence of cardiomyocyte HMGCS2. CONCLUSIONS: The canonical ketogenic enzyme, HMGCS2, mediates the therapeutic effects of the oxidized form of nicotinamide adenine dinucleotide repletion in HFpEF by restoring normal lipid metabolism and mitochondrial function.