Ex vivo modelling of cardiac injury identifies ferroptosis-related pathways as a potential therapeutic avenue for translational medicine.

BACKGROUND: Heart failure (HF) is a burgeoning health problem worldwide. Often arising as a result of cardiac injury, HF has become a major cause of mortality with limited availability of effective treatments. Ferroptotic pathways, triggering an iron-dependent form of cell death, are known to be potential key players in heart disease. This form of cell death does not exhibit typical characteristics of programmed cell death, and is mediated by impaired iron metabolism and lipid peroxidation signalling. OBJECTIVES: The aim of this study is to establish an ex-vivo model of myocardial injury in living myocardial slices (LMS) and to identify novel underlying mechanisms and potential therapeutic druggable target(s). METHODS AND RESULTS: In this study, we employed LMS as an ex vivo model of cardiac injury to investigate underlying mechanisms and potential therapeutic targets. Cryoinjury was induced in adult rat LMS, resulting in 30 % tissue damage. Cryoinjured LMS demonstrated impaired contractile function, cardiomyocyte hypertrophy, inflammation, and cardiac fibrosis, closely resembling in vivo cardiac injury characteristics. Proteomic analysis revealed an enrichment of factors associated with ferroptosis in the injured LMS, suggesting a potential causative role. To test this hypothesis, we pharmacologically inhibited ferroptotic pathways using ferrostatin (Fer-1) in the cryoinjured rat LMS, resulting in attenuation of structural changes and repression of pro-fibrotic processes. Furthermore, LMS generated from failing human hearts were used as a model of chronic heart failure. In this model, Fer-1 treatment was observed to reduce the expression of ferroptotic genes, enhances contractile function and improves tissue viability. Blocking ferroptosis-associated pathways in human cardiac fibroblasts (HCFs) resulted in a downregulation of fibroblast activation genes, a decrease in fibroblast migration capacity, and a reduction in reactive oxygen species production. RNA sequencing analysis of Fer-1-treated human LMS implicated metallothioneins as a potential underlying mechanism for the inhibition of these pathways. This effect is possibly mediated through the replenishment of glutathione reserves. CONCLUSIONS: Our findings highlight the potential of targeting ferroptosis-related pathways and metallothioneins as a promising strategy for the treatment of heart disease.

Amyloid precursor protein as a fibrosis marker in infants with biliary atresia.

BACKGROUND: Biliary atresia (BA) is a rare condition of unknown origin in newborns with jaundice. In BA bile ducts are non-functional, causing neonatal cholestasis and following liver fibrosis and failure. METHODS: This retrospective study included liver biopsies of 14 infants with BA aged [mean ± SD] 63 ± 23 days. Patients were grouped according to the clinical course (jaundice-free vs recurrent jaundice vs required liver transplantation or liver fibrosis (Ishak fibrosis score)) and followed for 1.61-5.64 years (mean 4.03). Transcriptome profiles were assessed using a panel of 768 fibrosis-specific genes, reanalyzed via qRT-PCR, and confirmed via immunostaining. Plasma from an additional 30 BA infants and 10 age-matched controls were used for amyloid precursor protein (APP) quantification by ELISA. RESULTS: Different clinical outcome groups showed a homogeneous mRNA expression. Altered amyloid-metabolism-related gene expression was found between cases with Ishak fibrosis score greater than 4. Immunostaining confirmed a distinct presence of APP in the livers of all BA subjects. APP plasma levels were higher in BA than in age-matched controls and correlated with the histological fibrosis grade. CONCLUSIONS: These results suggest that amyloidosis may contribute to BA and liver fibrosis, indicating that APP could serve as a potential liquid biomarker for these conditions. IMPACT: Biliary atresia patients with higher fibrosis scores according to Ishak have higher hepatic expression of amyloid-related genes while amyloid precursor protein accumulates in the liver and increases in the circulation. After a recent study revealed beta-amyloid deposition as a mechanism potentially involved in biliary atresia, we were able to correlate amyloid-metabolism-related transcript levels as well as amyloid precursor protein tissue and plasma levels with the degree of hepatic fibrosis. These findings suggest that amyloid precursor protein is a fibrosis marker in infants with biliary atresia, reinforcing the role of amyloid metabolism in the pathogenesis of this serious disease.

Deep Learning for 3D Vascular Segmentation in Phase Contrast Tomography.

Automated blood vessel segmentation is critical for biomedical image analysis, as vessel morphology changes are associated with numerous pathologies. Still, precise segmentation is difficult due to the complexity of vascular structures, anatomical variations across patients, the scarcity of annotated public datasets, and the quality of images. Our goal is to provide a foundation on the topic and identify a robust baseline model for application to vascular segmentation using a new imaging modality, Hierarchical Phase-Contrast Tomography (HiP-CT). We begin with an extensive review of current machine learning approaches for vascular segmentation across various organs. Our work introduces a meticulously curated training dataset, verified by double annotators, consisting of vascular data from three kidneys imaged using Hierarchical Phase-Contrast Tomography (HiP-CT) as part of the Human Organ Atlas Project. HiP-CT, pioneered at the European Synchrotron Radiation Facility in 2020, revolutionizes 3D organ imaging by offering resolution around 20µm/voxel, and enabling highly detailed localized zooms up to 1µm/voxel without physical sectioning. We leverage the nnU-Net framework to evaluate model performance on this high-resolution dataset, using both known and novel samples, and implementing metrics tailored for vascular structures. Our comprehensive review and empirical analysis on HiP-CT data sets a new standard for evaluating machine learning models in high-resolution organ imaging. Our three experiments yielded Dice scores of 0.9523 and 0.9410, and 0.8585, respectively. Nevertheless, DSC primarily assesses voxel-to-voxel concordance, overlooking several crucial characteristics of the vessels and should not be the sole metric for deciding the performance of vascular segmentation. Our results show that while segmentations yielded reasonably high scores-such as centerline Dice values ranging from 0.82 to 0.88, certain errors persisted. Specifically, large vessels that collapsed due to the lack of hydro-static pressure (HiP-CT is an ex vivo technique) were segmented poorly. Moreover, decreased connectivity in finer vessels and higher segmentation errors at vessel boundaries were observed. Such errors, particularly in significant vessels, obstruct the understanding of the structures by interrupting vascular tree connectivity. Through our review and outputs, we aim to set a benchmark for subsequent model evaluations using various modalities, especially with the HiP-CT imaging database.

Multidimensional Analysis of the Adult Human Heart in Health and Disease Using Hierarchical Phase-Contrast Tomography.

Background Current clinical imaging modalities such as CT and MRI provide resolution adequate to diagnose cardiovascular diseases but cannot depict detailed structural features in the heart across length scales. Hierarchical phase-contrast tomography (HiP-CT) uses fourth-generation synchrotron sources with improved x-ray brilliance and high energies to provide micron-resolution imaging of intact adult organs with unprecedented detail. Purpose To evaluate the capability of HiP-CT to depict the macro- to microanatomy of structurally normal and abnormal adult human hearts ex vivo. Materials and Methods Between February 2021 and September 2023, two adult human donor hearts were obtained, fixed in formalin, and prepared using a mixture of crushed agar in a 70% ethanol solution. One heart was from a 63-year-old White male without known cardiac disease, and the other was from an 87-year-old White female with a history of multiple known cardiovascular pathologies including ischemic heart disease, hypertension, and atrial fibrillation. Nondestructive ex vivo imaging of these hearts without exogenous contrast agent was performed using HiP-CT at the European Synchrotron Radiation Facility. Results HiP-CT demonstrated the capacity for high-spatial-resolution, multiscale cardiac imaging ex vivo, revealing histologic-level detail of the myocardium, valves, coronary arteries, and cardiac conduction system across length scales. Virtual sectioning of the cardiac conduction system provided information on fatty infiltration, vascular supply, and pathways between the cardiac nodes and adjacent structures. HiP-CT achieved resolutions ranging from gross (isotropic voxels of approximately 20 µm) to microscopic (approximately 6.4-µm voxel size) to cellular (approximately 2.3-µm voxel size) in scale. The potential for quantitative assessment of features in health and disease was demonstrated. Conclusion HiP-CT provided high-spatial-resolution, three-dimensional images of structurally normal and diseased ex vivo adult human hearts. Whole-heart image volumes were obtained with isotropic voxels of approximately 20 µm, and local regions of interest were obtained with resolution down to 2.3-6.4 µm without the need for sectioning, destructive techniques, or exogenous contrast agents. Published under a CC BY 4.0 license Supplemental material is available for this article. See also the editorial by Bluemke and Pourmorteza in this issue.

Immunohistochemical marker profiles for the differentiation of collagenous spherulosis from adenoid cystic carcinoma of the breast.

Collagenous spherulosis (CS) is a rare breast lesion of unknown histogenesis. Adenoid cystic carcinoma (ACC) is a rare basal-like breast carcinoma with low histological grade. CS is a benign lesion but resembles ACC. Both lesions show a similar histomorphology and feature bilineage differentiation. This study compared immunohistochemical markers in CS and ACC. We compiled n = 13 CS cases and n = 18 mammary ACCs. Fourteen marker proteins (ER, PR, HER2, GATA3, CK7, E-cadherin, CD117, CK5/14, p40, p63, SMA, CD10, calponin, P-cadherin) were evaluated by immunohistochemistry (IHC). MYB rearrangement, a common alteration in ACC, was assessed by fluorescence in situ hybridization. Patient age ranged between 40-60 years for CS lesions and 30-90 years for ACCs. 7/13 (54%) CS cases harbored a lobular carcinoma in situ (LCIS) in the luminal component. One CS/LCIS lesion occurred in a carrier of a pathogenic germline variant in CDH1/E-cadherin. MYB rearrangement was detected in 0/11 (0%) CS and 6/16 (37%) ACC cases (P = 0.054). CS was associated with expression of ER in the luminal component (P < 0.001), E-cadherin loss in the luminal component (P = 0.045), and expression of CD10 and calponin in the basal component (P < 0.001). Furthermore, CS was associated with GATA3 expression in the luminal component (12/13 [92%] versus 5/18 [27%], P < 0.001). In summary, IHC for GATA3 and E-cadherin may contribute to the differential diagnosis between CS and ACC, although these markers are not exclusively expressed in either lesion. Histologic evaluation has to take into account that CS is frequently colonized by LCIS, requiring thorough correlation of histomorphology and immunohistochemical features.

The role of vasculature and angiogenesis in respiratory diseases.

In European countries, nearly 10% of all hospital admissions are related to respiratory diseases, mainly chronic life-threatening diseases such as COPD, pulmonary hypertension, IPF or lung cancer. The contribution of blood vessels and angiogenesis to lung regeneration, remodeling and disease progression has been increasingly appreciated. The vascular supply of the lung shows the peculiarity of dual perfusion of the pulmonary circulation (vasa publica), which maintains a functional blood-gas barrier, and the bronchial circulation (vasa privata), which reveals a profiled capacity for angiogenesis (namely intussusceptive and sprouting angiogenesis) and alveolar-vascular remodeling by the recruitment of endothelial precursor cells. The aim of this review is to outline the importance of vascular remodeling and angiogenesis in a variety of non-neoplastic and neoplastic acute and chronic respiratory diseases such as lung infection, COPD, lung fibrosis, pulmonary hypertension and lung cancer.

Hypoxia Attenuates Pressure Overload-Induced Heart Failure.

BACKGROUND: Alveolar hypoxia is protective in the context of cardiovascular and ischemic heart disease; however, the underlying mechanisms are incompletely understood. The present study sought to test the hypothesis that hypoxia is cardioprotective in left ventricular pressure overload (LVPO)-induced heart failure. We furthermore aimed to test that overlapping mechanisms promote cardiac recovery in heart failure patients following left ventricular assist device-mediated mechanical unloading and circulatory support. METHODS AND RESULTS: We established a novel murine model of combined chronic alveolar hypoxia and LVPO following transverse aortic constriction (HxTAC). The HxTAC model is resistant to cardiac hypertrophy and the development of heart failure. The cardioprotective mechanisms identified in our HxTAC model include increased activation of HIF (hypoxia-inducible factor)-1α-mediated angiogenesis, attenuated induction of genes associated with pathological remodeling, and preserved metabolic gene expression as identified by RNA sequencing. Furthermore, LVPO decreased Tbx5 and increased Hsd11b1 mRNA expression under normoxic conditions, which was attenuated under hypoxic conditions and may induce additional hypoxia-mediated cardioprotective effects. Analysis of samples from patients with advanced heart failure that demonstrated left ventricular assist device-mediated myocardial recovery revealed a similar expression pattern for TBX5 and HSD11B1 as observed in HxTAC hearts. CONCLUSIONS: Hypoxia attenuates LVPO-induced heart failure. Cardioprotective pathways identified in the HxTAC model might also contribute to cardiac recovery following left ventricular assist device support. These data highlight the potential of our novel HxTAC model to identify hypoxia-mediated cardioprotective mechanisms and therapeutic targets that attenuate LVPO-induced heart failure and mediate cardiac recovery following mechanical circulatory support.

Adhesion of vessel wall to stentriever during combined technique for mechanical thrombectomy in acute ischemic stroke: A histomorphological study.

PURPOSE: Detection of vessel wall tissue in thrombus material in patients with ischemic stroke is judged as vascular injury. So far, several studies investigated components of the free clots after mechanical thrombectomy. The aim of this retrospective study was to investigate the involvement and role of the stentriever in vessel wall injury by analysis of the composition of adherent tissue to the stentriever during combined aspiration thrombectomy with stentriever. METHODS: Stentriever with adherent tissue and free clots in aspiration samples from patients undergoing mechanical thrombectomy (aspiration plus stentriever) were separately assessed for the occurrence of parts of vascular tissue together with clinical and interventional data as well as clinical outcome data. Specimens were analyzed histomorphologically and immunohistochemically. Findings, focused on parts of vessel wall were reported together with clinical data. RESULTS: Specimens from 21 identified patients were available. Parts of the vessel wall were detected in 7 out 21 (33%) samples. All specimens revealed fresh thrombus material without signs of organization or atheromatous tissue. In 90% of patients mTICI was greater than 2b without signs of secondary vessel injury. No vascular tissue was found in free clots of the aspiration samples. CONCLUSION: The examination of adherent tissue to the stentriever instead of the examination of free clots may affect the number of detected parts of vessel wall. Further studies in combination with vessel wall imaging may elucidate the origin of remnants of vessel wall.

Organ Mapping Antibody Panels: a community resource for standardized multiplexed tissue imaging.

Multiplexed antibody-based imaging enables the detailed characterization of molecular and cellular organization in tissues. Advances in the field now allow high-parameter data collection (>60 targets); however, considerable expertise and capital are needed to construct the antibody panels employed by these methods. Organ mapping antibody panels are community-validated resources that save time and money, increase reproducibility, accelerate discovery and support the construction of a Human Reference Atlas.

Pre-emptive iron supplementation prevents myocardial iron deficiency and attenuates adverse remodelling after myocardial infarction.

AIMS: Heart failure (HF) after myocardial infarction (MI) is a major cause of morbidity and mortality. We sought to investigate the functional importance of cardiac iron status after MI and the potential of pre-emptive iron supplementation in preventing cardiac iron deficiency (ID) and attenuating left ventricular (LV) remodelling. METHODS AND RESULTS: MI was induced in C57BL/6J male mice by left anterior descending coronary artery ligation. Cardiac iron status in the non-infarcted LV myocardium was dynamically regulated after MI: non-haem iron and ferritin increased at 4 weeks but decreased at 24 weeks after MI. Cardiac ID at 24 weeks was associated with reduced expression of iron-dependent electron transport chain (ETC) Complex I compared with sham-operated mice. Hepcidin expression in the non-infarcted LV myocardium was elevated at 4 weeks and suppressed at 24 weeks. Hepcidin suppression at 24 weeks was accompanied by more abundant expression of membrane-localized ferroportin, the iron exporter, in the non-infarcted LV myocardium. Notably, similarly dysregulated iron homeostasis was observed in LV myocardium from failing human hearts, which displayed lower iron content, reduced hepcidin expression, and increased membrane-bound ferroportin. Injecting ferric carboxymaltose (15 µg/g body weight) intravenously at 12, 16, and 20 weeks after MI preserved cardiac iron content and attenuated LV remodelling and dysfunction at 24 weeks compared with saline-injected mice. CONCLUSION: We demonstrate, for the first time, that dynamic changes in cardiac iron status after MI are associated with local hepcidin suppression, leading to cardiac ID long term after MI. Pre-emptive iron supplementation maintained cardiac iron content and attenuated adverse remodelling after MI. Our results identify the spontaneous development of cardiac ID as a novel disease mechanism and therapeutic target in post-infarction LV remodelling and HF.

Human lung virtual histology by multi-scale x-ray phase-contrast computed tomography.

Objectives.As the central organ of the respiratory system, the human lung is responsible for supplying oxygen to the blood, which reaches the erythrocytes by diffusion through the alveolar walls and is then distributed throughout the body. By exploiting the difference in electron density detected by a phase shift in soft tissue, high-resolution x-ray phase-contrast computed tomography (XPCT) can resolve biological structures in a sub-µm range, shedding new light on the three-dimensional structure of the lungs, physiological functions and pathological mechanisms.Approach.This work presents both synchrotron and laboratory XPCT results of postmortem tissue from autopsies and biopsies embedded with various preparation protocols such as precision-cut lung slices, cryogenically fixed lung tissue, as well as paraffin and alcohol fixed tissue. The selection of pathological abnormalities includes channel of Lambert, bronchus-associated lymphoid tissue and alveolar capillary dysplasia with misalignment of pulmonary veins. Subsequently, quantification and visualization approaches are presented.Main results.The overall high image quality even of in-house XPCT scans for the case of FFPE biopsies can be exploited for a wide range of pulmonary pathologies and translated to dedicated and optimized instrumentation which could be operated in clinical setting. By using synchrotron radiation, contrast can be further increased to resolve sub-µm sized features down to the sub-cellular level. The results demonstrate that a wide range of preparation protocols including sample mounting in liquids can be used.Significance.With XPCT, poorly understood 3D structures can be identified in larger volume overview and subsequently studied in more detail at higher resolution. With the full 3D structure, the respective physiological functions of airways or vascular networks, and the different pathophysiologic mechanisms can be elucidated or at least underpinned with structural data. Moreover, synchrotron data can be used to validate laboratory protocols and provide ground truth for standardizing the method.

FDA-approved Abl/EGFR/PDGFR kinase inhibitors show potent efficacy against pandemic and seasonal influenza A virus infections of human lung explants.

Influenza viruses (IVs) cause substantial global morbidity and mortality. Given the limited range of licensed antiviral drugs and their reduced efficacy due to resistance mutations, repurposing FDA-approved kinase inhibitors as fast-tracked host-targeted antivirals is an attractive strategy. We identified six FDA-approved non-receptor tyrosine kinase-inhibitors (NRTKIs) as potent inhibitors of viral replication of pandemic and seasonal IVs in vitro. We validated their efficacy in a biologically and clinically relevant ex vivo model of human precision-cut lung slices. We identified steps of the virus infection cycle affected by these inhibitors and assessed their effect(s) on host responses. Their overlapping targets suggest crosstalk between Abl, EGFR, and PDGFR pathways during IAV infection. Our data and established safety profiles of these NRTKIs provide compelling evidence for further clinical investigations and repurposing as host-targeted influenza antivirals. Moreover, these NRTKIs have broad-spectrum antiviral potential given that their kinase/pathway targets are critical for the replication of many viruses.

Vitamin A regulates tissue-specific organ remodeling in diet-induced obesity independent of mitochondrial function.

Background: Perturbed mitochondrial energetics and vitamin A (VitA) metabolism are associated with the pathogenesis of diet-induced obesity (DIO) and type 2 diabetes (T2D). Methods: To test the hypothesis that VitA regulates tissue-specific mitochondrial energetics and adverse organ remodeling in DIO, we utilized a murine model of impaired VitA availability and high fat diet (HFD) feeding. Mitochondrial respiratory capacity and organ remodeling were assessed in liver, skeletal muscle, and kidney tissue, which are organs affected by T2D-associated complications and are critical for the pathogenesis of T2D. Results: In liver, VitA had no impact on maximal ADP-stimulated mitochondrial respiratory capacity (VADP) following HFD feeding with palmitoyl-carnitine and pyruvate each combined with malate as substrates. Interestingly, histopathological and gene expression analyses revealed that VitA mediates steatosis and adverse remodeling in DIO. In skeletal muscle, VitA did not affect VADP following HFD feeding. No morphological differences were detected between groups. In kidney, VADP was not different between groups with both combinations of substrates and VitA transduced the pro-fibrotic transcriptional response following HFD feeding. Conclusion: The present study identifies an unexpected and tissue-specific role for VitA in DIO that regulates the pro-fibrotic transcriptional response and that results in organ damage independent of changes in mitochondrial energetics.

Inflammation and vascular remodeling in COVID-19 hearts.

A wide range of cardiac symptoms have been observed in COVID-19 patients, often significantly influencing the clinical outcome. While the pathophysiology of pulmonary COVID-19 manifestation has been substantially unraveled, the underlying pathomechanisms of cardiac involvement in COVID-19 are largely unknown. In this multicentre study, we performed a comprehensive analysis of heart samples from 24 autopsies with confirmed SARS-CoV-2 infection and compared them to samples of age-matched Influenza H1N1 A (n = 16), lymphocytic non-influenza myocarditis cases (n = 8), and non-inflamed heart tissue (n = 9). We employed conventional histopathology, multiplexed immunohistochemistry (MPX), microvascular corrosion casting, scanning electron microscopy, X-ray phase-contrast tomography using synchrotron radiation, and direct multiplexed measurements of gene expression, to assess morphological and molecular changes holistically. Based on histopathology, none of the COVID-19 samples fulfilled the established diagnostic criteria of viral myocarditis. However, quantification via MPX showed a significant increase in perivascular CD11b/TIE2 + -macrophages in COVID-19 over time, which was not observed in influenza or non-SARS-CoV-2 viral myocarditis patients. Ultrastructurally, a significant increase in intussusceptive angiogenesis as well as multifocal thrombi, inapparent in conventional morphological analysis, could be demonstrated. In line with this, on a molecular level, COVID-19 hearts displayed a distinct expression pattern of genes primarily coding for factors involved in angiogenesis and epithelial-mesenchymal transition (EMT), changes not seen in any of the other patient groups. We conclude that cardiac involvement in COVID-19 is an angiocentric macrophage-driven inflammatory process, distinct from classical anti-viral inflammatory responses, and substantially underappreciated by conventional histopathologic analysis. For the first time, we have observed intussusceptive angiogenesis in cardiac tissue, which we previously identified as the linchpin of vascular remodeling in COVID-19 pneumonia, as a pathognomic sign in affected hearts. Moreover, we identified CD11b + /TIE2 + macrophages as the drivers of intussusceptive angiogenesis and set forward a putative model for the molecular regulation of vascular alterations.

Correction of heat-induced susceptibility changes in respiratory-triggered 2D-PRF-based thermometry for monitoring of magnetic resonance-guided hepatic microwave ablation in a human-like in vivo porcine model.

PURPOSE: To develop and evaluate susceptibility corrected 2D proton resonance frequency (PRF)-based magnetic resonance (MR)-thermometry for the accurate assessment of the ablation zone of hepatic microwave ablation (MWA). METHODS AND MATERIALS: Twelve hepatic MWA were performed in five LEWE minipigs with human-like fissure-free liver. Temperature maps during ablation of PRF-based MR-thermometry were corrected by modeling heat induced susceptibility changes. Ablation zones were determined using cumulative equivalent minutes at 43 °C (CEM43) as tissue damage model. T1 weighted (w) post-ablation contrast-enhanced (CE) MR-imaging and manually segmented postmortem histology were used for validation. The agreement of uncorrected (raw) and susceptibility corrected (corr) MR-thermometry with T1w post-ablation CE MR-imaging and histology was evaluated. The Wilcoxon-signed rank test and Bland-Altman analysis were applied. RESULTS: With the susceptibility corrected MR-thermometry a significantly increased dice coefficient (raw: 77% vs. corr: 83%, p < 0.01) and sensitivity (raw: 72% vs. corr: 82%, p < 0.01) was found for the comparison to T1w-CE imaging as well as histopathology (dice coefficients: raw: 76% vs. corr: 79%, p < 0.001; sensitivity: raw: 72% vs. corr: 74%, p < 0.001). While major axis length was significantly increased (7.1 mm, p < 0.001) and minor axis length significantly decreased (2.2 mm, p < 0.001) in uncorrected MR-thermometry compared to T1w-CE MR-imaging, no significant bias was found after susceptibility correction. CONCLUSION: Using susceptibility corrected 2D PRF-based MR-thermometry to predict the ablation zones of hepatic MWA provided a good agreement in comparison to T1w post-ablation CE MR-imaging and histopathology.

Vitamin A preserves cardiac energetic gene expression in a murine model of diet-induced obesity.

Perturbed vitamin-A metabolism is associated with type 2 diabetes and mitochondrial dysfunction that are pathophysiologically linked to the development of diabetic cardiomyopathy (DCM). However, the mechanism, by which vitamin A might regulate mitochondrial energetics in DCM has previously not been explored. To test the hypothesis that vitamin-A deficiency accelerates the onset of cardiomyopathy in diet-induced obesity (DIO), we subjected mice with lecithin retinol acyltransferase (Lrat) germline deletion, which exhibit impaired vitamin-A stores, to vitamin A-deficient high-fat diet (HFD) feeding. Wild-type mice fed with a vitamin A-sufficient HFD served as controls. Cardiac structure, contractile function, and mitochondrial respiratory capacity were preserved despite vitamin-A deficiency following 20 wk of HFD feeding. Gene profiling by RNA sequencing revealed that vitamin A is required for the expression of genes involved in cardiac fatty acid oxidation, glycolysis, tricarboxylic acid cycle, and mitochondrial oxidative phosphorylation in DIO as expression of these genes was relatively preserved under vitamin A-sufficient HFD conditions. Together, these data identify a transcriptional program, by which vitamin A preserves cardiac energetic gene expression in DIO that might attenuate subsequent onset of mitochondrial and contractile dysfunction.NEW & NOTEWORTHY The relationship between vitamin-A status and the pathogenesis of diabetic cardiomyopathy has not been studied in detail. We assessed cardiac mitochondrial respiratory capacity, contractile function, and gene expression by RNA sequencing in a murine model of combined vitamin-A deficiency and diet-induced obesity. Our study identifies a role for vitamin A in preserving cardiac energetic gene expression that might attenuate subsequent development of mitochondrial and contractile dysfunction in diet-induced obesity.