Skeletal muscle is a major contributor to whole-body glucose homeostasis and is an important endocrine organ. To date, few studies have undertaken the large-scale identification of skeletal muscle-derived secreted proteins (myokines), particularly in response to stimuli that activate pathways governing energy metabolism in health and disease. Whereas the AMP-activated protein kinase (AMPK) and insulin-signaling pathways have received notable attention for their ability to independently regulate skeletal muscle substrate metabolism, little work has examined their ability to re-pattern the secretome. The present study coupled the use of high-resolution MS-based proteomics and bioinformatics analysis of conditioned media derived from 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR—an AMPK activator)- and insulin-treated differentiated C2C12 myotubes. We quantified 858 secreted proteins, including cytokines and growth factors such as fibroblast growth factor-21 (Fgf21). We identified 377 and 118 proteins that were significantly altered by insulin and AICAR treatment, respectively. Notably, the family of insulin growth factor binding-proteins (Igfbp) was differentially regulated by each treatment. Insulin- but not AICAR-induced conditioned media increased the mitochondrial respiratory capacity of myotubes, potentially via secreted factors. These findings may serve as an important resource to elucidate secondary metabolic effects of insulin and AICAR stimulation in skeletal muscle.

Session V of the inaugural biennial Research Symposium on Pulmonary Injury and Repair of the Endothelium showcased cutting-edge research on pulmonary endothelial crosstalk with end organs and its role in vascular disease. Growing evidence suggests that communication between injured organs and distal vascular beds plays a critical role in the pathogenesis of complex conditions such as sepsis, acute respiratory distress syndrome, pulmonary arterial hypertension, and heart failure with preserved ejection fraction. Circulating mediators—including heparan sulfate fragments, proinflammatory cytokines, mitochondrial damage-associated molecular patterns, bone morphogenetic protein 9, bile acids, and nitric oxide—have emerged as key factors linking pulmonary endothelial dysfunction to neural impairment, acute kidney injury, subclinical liver injury, and left-sided heart disease. This review highlights recent advances in the field, identifies critical knowledge gaps, and outlines future research directions aimed at elucidating mechanisms of multiorgan dysfunction and identifying novel therapeutic targets.

As thorough, noninvasive assessment of congestion is not available in small animals, we developed and validated an ultrasonography-based research tool to evaluate pulmonary and central venous congestion in experimental heart failure models.

BACKGROUND: The role of leukocytes in the pathogenesis of right ventricular (RV) dysfunction (RVD) associated with heart failure with preserved ejection fraction (HFpEF) remains poorly defined, partially due to the lack of suitable small animal models. Here, we followed a translational research approach by establishing a murine HFpEF model developing manifest RVD and analyzed human HFpEF cohorts to study the mechanistic link between leukocytes and RVD in HFpEF. METHODS: Young (80-week-old) male and female C57BL/6J mice were divided into 4 experimental groups: chow, HFpEF (L-NAME [Nω-nitro-L-arginine methyl ester] plus a high-fat diet), chronic hypoxia (10% O ₂ ), and HFpEF plus hypoxia (RV-HFpEF). Biventricular function and myeloid cell dynamics were assessed across groups. To test whether myeloid cells are causally involved in the development of RV remodeling in HFpEF, we treated RV-HFpEF mice with a colony-stimulating factor 1 receptor inhibitor to deplete myeloid cells. RESULTS: RV-HFpEF resulted in left ventricular diastolic dysfunction, indicated by increased E/E′ ratio, reduced global longitudinal peak strain, smaller end-diastolic diameters, and increased isovolumetric relaxation time compared with chow. RV-HFpEF animals developed RV hypertrophy and RVD, evident as increased Fulton index as well as elevated RV systolic pressure and reduced tricuspid annular plane systolic excursion, respectively. Total leukocyte, monocyte, and macrophage counts were elevated in RV tissue from RV-HFpEF mice compared with RV tissue from chow mice and left ventricular tissue from young and aged RV-HFpEF animals. These findings were confirmed by unbiased proteomic analyses of RV tissue from RV-HFpEF mice, demonstrating an increased abundance of proteins involved in activation of the innate immune system, macrophage chemotaxis, and leukocyte migration when compared with left ventricular tissue and the other experimental groups. Fate-mapping experiments revealed that recruited monocyte-derived macrophages became the main source of total cardiac macrophages in RV tissue from RV-HFpEF mice. Depletion of myeloid cells was associated with lower RVSP profiles in RV-HFpEF mice compared with controls. In RV biopsies from patients with HFpEF, we found increased expression of adhesion molecules, fibrotic markers, and inflammatory transcripts, as well as an association between RVD and CD68 ⁺ cells. CONCLUSIONS: We demonstrate that dysregulated myeloid cell dynamics are associated with, and directly contribute to, the pathogenesis of RVD associated with HFpEF in humans and mice.

Background: Dyspnea and exercise intolerance are the primary clinical symptoms of heart failure. Heart failure patients experience frequent hypoxemic episodes, yet underlying mechanisms and relevance remain poorly understood. In a cohort of heart failure patients and multiple animal models, we identify pulmonary capillary rarefaction driven by excessive autophagy in endothelial cells as a novel mechanism of hypoxemia and cardiac disease progression. Methods: A cohort of heart failure with preserved ejection fraction (HFpEF) patients was analyzed for parameters of left ventricular (LV) dysfunction and pulmonary gas exchange. Morphological and cellular mechanisms of impaired pulmonary oxygenation were assessed in three animal models of heart failure, namely two HFpEF models, SU5416-treated ZSF1 obese rats and high fat diet/L-NAME treated mice, and in rats subjected to aortic banding. Lung microvascular rarefaction was quantified by micro-computed tomography, stereology, flow cytometry and dye efflux. Cellular mechanisms of capillary loss were analyzed by single-cell transcriptomics, electron microscopy and immunofluorescence, and in mice with endothelial-specific deletion of the autophagy gene Atg7 (Atg7 ^EN-KO ) . Results: In 234 HFpEF patients, advancing NYHA class was associated with progressive worsening of arterial oxygen saturation at rest and during exercise and a reduced lung diffusing capacity. Impaired gas diffusion correlated with indices of LV diastolic dysfunction. Impaired oxygenation and reduced exercise capacity were similarly evident in animal models of left heart disease, which showed a distinct loss of pulmonary microvessels and capillaries. Lung microvascular endothelial cells in HFpEF showed characteristics of increased autophagic flux and apoptosis. Relative to their wild type HFpEF controls, Atg7 ^EN-KO mice had less capillary loss, restored normoxemia, improved exercise tolerance, and mitigated LV diastolic dysfunction. Additional studies in HFpEF mice corroborated the functional relevance of impaired gas exchange for the progression of left heart disease by demonstrating that additional hypoxia aggravated, whereas moderate hyperoxia improved LV function. Conclusion: Our findings identify pulmonary microvascular rarefaction as a novel pathomechanism in heart failure that i) contributes to dyspnea and exercise intolerance, ii) impairs pulmonary gas exchange and iii) accelerates LV disease progression. Strategies targeting this axis such as moderate oxygen therapy may mitigate cardiopulmonary morbidity in heart failure. Clinical Trial Registration: Registered in the DRKS (Deutsches Register für klinische Studien) as trial# DRKS00032974 at https://drks.de/search/en/trial/DRKS00032974 .