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High-mobility group box protein 1 (HMGB1) is released during tissue damage and activates the innate immune system through toll-like receptor 4. Because mortality in dilated cardiomyopathy (DCM) is associated with activation of the innate immune system, we hypothesized that HMGB1 possesses a prognostic value in estimating mortality in patients with DCM. We determined HMGB1 and N-terminal B-type natriuretic peptide (NT-proBNP) levels in 67 patients with DCM (12 women, mean age 53.6 ± 1.5 years). Kaplan-Meier analyzes revealed that higher levels of HMGB1 and NT-proBNP are related to increased all-cause mortality. Multivariable Cox regression confirmed HMGB1 as a risk factor for mortality in patients with DCM, independent of NT-proBNP, age, and gender (hazard ratio per 1 SD 1.920, 95% confidence interval 1.401 to 2.631, plt;0.001). HMGB1 is a promising candidate to estimate the prognosis of patients with DCM.

Post-COVID-19 syndrome (PCS) has been described as ‘the pandemic after the pandemic’ with more than 65 million people worldwide being affected. The enormous range of symptoms makes both diagnosis complex and treatment difficult. In a post-COVID rehabilitation outpatient clinic, 184 patients, mostly non-hospitalized, received a comprehensive, interdisciplinary diagnostic assessment with fixed follow-up appointments. At baseline, three in four patients reported more than 10 symptoms, the most frequent symptoms were fatigue (84.9%), decreased physical capacity (83.0%), tiredness (81.1%), poor concentration (73.6%), sleeping problems (66.7%) and shortness of breath (67.3%). Abnormalities were found in the mean values of scores for fatigue (FAS = 34.3), cognition (MoCA = 25.5), psychological alterations (anxiety, depression, post-traumatic stress disorder), limitation of lung function (CAT) and severity scores for PCS (PCFS, MCRS). Clinical abnormalities were found in elevated values of heart rate, breathing rate at rest, blood pressure and NT-proBNP levels. As the frequency of the described symptoms decreases only slowly but most often significantly over the course, it is important to monitor the patients over a longer period of time. Many of them suffer from an immense symptom burden, often without pre-existing clinical correlates. Our results show a clear association with objectifiable assessments and tests as well as pronounced symptoms.

Development and progression of heart failure involve endothelial and myocardial dysfunction as well as a dysregulation of the NO-sGC-cGMP signalling pathway. Recently, we reported that the sGC stimulator riociguat has beneficial effects on cardiac remodelling and progression of heart failure in response to chronic pressure overload. Here, we examined if these beneficial effects of riociguat were also reflected in alterations of the myocardial proteome and microRNA profiles.Male C57BL/6N mice underwent transverse aortic constriction (TAC) and sham-operated mice served as controls. TAC and sham animals were randomised and treated with either riociguat or vehicle for 5 weeks, starting 3 weeks after surgery, when cardiac hypertrophy was established. Afterwards, we performed mass spectrometric proteome analyses and microRNA sequencing of proteins and RNAs, respectively, isolated from left ventricles (LVs).TAC-induced changes of the LV proteome were significantly reduced by treatment with riociguat. Bioinformatics analyses revealed that riociguat improved TAC-induced cardiovascular disease-related pathways, metabolism and energy production, for example, reversed alterations in the levels of myosin heavy chain 7, cardiac phospholamban and ankyrin repeat domain-containing protein 1. Riociguat also attenuated TAC-induced changes of microRNA levels in the LV.The sGC stimulator riociguat exerted beneficial effects on cardiac structure and function during pressure overload, which was accompanied by a reversal of TAC-induced changes of the cardiac proteome and microRNA profile. Our data support the potential of riociguat as a novel therapeutic agent for heart failure.

Plasma levels of myeloid differentiation factor-2 (MD-2), a co-receptor of toll-like-receptor 4 (TLR4), independently predict mortality in patients with dilated cardiomyopathy (DCM). We tested whether monocyte activation by MD-2 contributes to immune activation and inflammatory status in DCM patients. We found increased MD-2 plasma levels in 25 patients with recent-onset DCM (1250 ± 80.7 ng/ml) compared to 25 age- and gender-matched healthy controls (793.4 ± 52.0 ng/ml; p 0.001). Monocytes isolated from DCM patients showed a higher expression (141.7 ± 12.4%; p = 0.006 vs. controls) of the MD-2 encoding gene, LY96 and an increased NF-κB-activation. Further, the TLR4-activator lipopolysaccharide (LPS) caused a higher increase in interleukin (IL)-6 in monocytes from DCM patients compared to controls (mean fluorescence intensity: 938.7 ± 151.0 vs. 466.9 ± 51.1; p = 0.005). MD-2 increased IL-6 secretion in a TLR4/NF-κB-dependent manner in monocyte-like THP-1-cells as demonstrated by TLR4-siRNA and NF-κB-inhibition. Since endothelial cells (ECs) are responsible for recruiting monocytes to the site of inflammation, ECs were treated with MD-2 leading to an activation of Akt and increased secretion of monocyte-chemoattractant-protein-1 (MCP-1). Activation of ECs by MD-2 was accompanied by an increased expression of the adhesion molecules CD54, CD106 and CD62E, resulting in an increased monocyte recruitment, which was attenuated by CD54 inhibition. In addition, in murine WT but not LY96-KO bone marrow-derived macrophages LPS increased the amount of CD54 and CD49d/CD29. MD-2 facilitates a pro-inflammatory status of monocytes and EC-mediated monocyte recruitment via TLR4/NF-κB. Elevated MD-2 plasma levels are possibly involved in monocyte-related inflammation-promoting disease progression in DCM. Our results suggest that MD-2 contributes to increasing monocytic inflammatory activity and triggers the recruitment of monocytes to ECs in DCM.

(1) Background: COVID-19 is often associated with significant long-term symptoms and disability, i.e., the long/post-COVID syndrome (PCS). Even after presumably mild COVID-19 infections, an increasing number of patients seek medical help for these long-term sequelae, which can affect various organ systems. The pathogenesis of PCS is not yet understood. Therapy has so far been limited to symptomatic treatment. The Greifswald Post COVID Rehabilitation Study (PoCoRe) aims to follow and deeply phenotype outpatients with PCS in the long term, taking a holistic and comprehensive approach to the analysis of their symptoms, signs and biomarkers. (2) Methods: Post-COVID outpatients are screened for symptoms in different organ systems with a standardized medical history, clinical examination, various questionnaires as well as physical and cardiopulmonary function tests. In addition, biomaterials are collected for the analysis of immunomodulators, cytokines, chemokines, proteome patterns as well as specific (auto)antibodies. Patients are treated according to their individual needs, adhering to the current standard of care. PoCoRe’s overall aim is to optimize diagnostics and therapy in PCS patients.

Background Septic cardiomyopathy worsens the prognosis of critically ill patients. Clinical data suggest that interleukin‐1β (IL‐1β), activated by the NLRP3 inflammasome, compromises cardiac function. Whether or not deleting Nlrp3 would prevent cardiac atrophy and improve diastolic cardiac function in sepsis was unclear. Here, we investigated the role of NLRP3/IL‐1β in sepsis‐induced cardiomyopathy and cardiac atrophy. Methods Male Nlrp3 knockout (KO) and wild‐type (WT) mice were exposed to polymicrobial sepsis by caecal ligation and puncture (CLP) surgery (KO, n = 27; WT, n = 33) to induce septic cardiomyopathy. Sham‐treated mice served as controls (KO, n = 11; WT, n = 16). Heart weights and morphology, echocardiography and analyses of gene and protein expression were used to evaluate septic cardiomyopathy and cardiac atrophy. IL‐1β effects on primary and immortalized cardiomyocytes were investigated by morphological and molecular analyses. IonOptix and real‐time deformability cytometry (RT‐DC) analysis were used to investigate functional and mechanical effects of IL‐1β on cardiomyocytes. Results Heart morphology and echocardiography revealed preserved systolic (stroke volume: WT sham vs. WT CLP: 33.1 ± 7.2 μL vs. 24.6 ± 8.7 μL, P

Introduction Our clinical research unveiled chronic heart failure with preserved ejection fraction (HFpEF) as a long-term sequel in survivors of severe pediatric burn injury due to a yet unknown molecular pathomechanism. Applying a standardized rat model, we systematically determined the pathophysiological impact of burn injury on long-term cardiac performance to uncover systemic and molecular pathomechanisms that may cause post-burn HFpEF development. Methods Male adolescent SD-rats were subjected to a 60 % total body surface area (TBSA) full-thickness burn- or sham-trauma and subsequently characterized after burn-injury by serial transthoracic echocardiography, bulk myocardial next-generation sequencing and proteomics as well as RT-PCR, immuno-blotting (IB), histology and plasma proteomics for cardiac performance and molecular alterations, respectively, at 3, 7, 30 and 90days. Results In comparison to the sham-group (SG), animals from the burn-group (BG) recapitulated typical post-burn clinical traits, such as significant loss in body weight (BG 27 % less than SG at 30d, p< 0.05) or skeletal muscle wasting (27 % less at 30d, p< 0.05) in accord with elevated molecular atrophy markers. We show post-burn cardiac muscle wasting (BG 22 % less at 30d, p< 0.05) and persistent markers of cardiac dysfunction in accord with significant histological cardiomyocyte hypotrophy (BG -8 % at 30d, p< 0.05) and significantly diminished left ventricular (LV) global longitudinal strain and isovolumic relaxation time in BGs, while LV-EF remained unchanged. Weighted gene network correlation analysis from bulk myocardial NGS and clinical traits related activation of immunological and pro-fibrotic pathways in post-burn injury hearts to cardiac dysfunction in BGs. Subsequent RT-PCR and histology confirmed significant myocardial accumulation of cardio-depressive damage associated molecular patterns (i.e., S100A8 and A9) and infiltration by granulocytes and monocytes as well as significant LV fibrosis. Serial plasma proteomic analysis indicated elevated plasma levels of S100A8 and A9 and other heart failure markers that mirrored similar changes in human post-burn plasma samples. Conclusions Here we report the development of HFpEF as a novel systemic consequence of severe burn injury in a rodent model, which warrants further mechanistic and translational studies. Cardiac inflammation and fibrosis are known to negatively impact cardiac performance and may be mechanistic key findings that will guide further therapeutic studies and subsequent validation of post-burn heart failure biomarkers Applicability of Research to Practice This model is part of a translational and interdisciplinary experimental and clinical effort to inform pathophysiology and mechanistics of long-term heart failure in burn patients. Echocardiographic data and parameters from this model are currently being used to evaluate adult survivors of severe burn injury for signs of HFpEF.

Background: We described chronic heart failure with preserved ejection fraction (HFpEF) as a long-term sequel in survivors of severe pediatric burn injury (BI) (Hundeshagen et al., Lancet Child & Adolescent Health, 2017). Applying a widely used standardized scald injury rat model in burn research, we sought to uncover systemic and molecular pathomechanisms that may cause post-BI HFpEF development. Methods and Results: Male adolescent SD-rats were subjected to a 60% total body surface area full-thickness BI (B; 100% survival) or sham (S) procedure (each n=10) and characterized them up to 90 days (3, 7, 30 and 90d) by serial echocardiography (E), bulk myocardial NGS and -proteomics, RT-PCR, IB, histology (H) and plasma proteomics for cardiac performance and molecular alterations, respectively. B rats mirrored typical post-burn clinical traits as significant loss in body (-27%*) or skeletal muscle weight (-30%*) e.g., with elevated atrophy markers as Murf1 (5-fold*) throughout the observation period vs S (30d, *P3-fold*) infiltration as well LV fibrosis (2.2-fold*). Cardiac proteomics yielded e.g., neutrophil degranulation as lead GO-term. Serial blood and plasma proteomic and ELISA analysis indicated elevated WBC (+26%*) and levels e.g., of IL6, S100A8/A9, CH3L1 and other HF markers alike changes in human post-BI plasma samples. WGCNA for bulk myocardial NGS and clinical traits related activated immunological and pro-fibrotic pathways in post-BI hearts to cardiac dysfunction in B. Conclusion: The first ever report of the development of HFpEF as a novel systemic consequence of severe burn injury in a rodent model prepares the ground for further mechanistic and translational studies. Cardiac inflammation and fibrosis that negatively impact cardiac performance may be mechanistic key findings guiding further therapeutic studies and validation of post-BI HF biomarkers.

Heart failure is associated with an impaired NO-soluble guanylyl cyclase (sGC)-cGMP pathway and its augmentation is thought to be beneficial for its therapy. We hypothesized that stimulation of sGC by the sGC stimulator riociguat prevents pathological cardiac remodelling and heart failure in response to chronic pressure overload.Transverse aortic constriction or sham surgery was performed in C57BL/6N mice. After 3 weeks of transverse aortic constriction when heart failure was established, animals receive either riociguat or its vehicle for 5 additional weeks. Cardiac function was evaluated weekly by echocardiography. Eight weeks after surgery, histological analyses were performed to evaluate remodelling and the transcriptome of the left ventricles (LVs) was analysed by RNA sequencing. Cell culture experiments were used for mechanistically studies.Transverse aortic constriction resulted in a continuous decrease of LV ejection fraction and an increase in LV mass until week 3. Five weeks of riociguat treatment resulted in an improved LV ejection fraction and a decrease in the ratio of left ventricular mass to total body weight (LVM/BW), myocardial fibrosis and myocyte cross-sectional area. RNA sequencing revealed that riociguat reduced the expression of myocardial stress and remodelling genes (e.g. Nppa, Nppb, Myh7 and collagen) and attenuated the activation of biological pathways associated with cardiac hypertrophy and heart failure. Riociguat reversed pathological stress response in cultivated myocytes and fibroblasts.Stimulation of the sGC reverses transverse aortic constriction-induced heart failure and remodelling, which is associated with improved myocardial gene expression.This article is part of a themed issue on cGMP Signalling in Cell Growth and Survival. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v179.11/issuetoc.

The transcription factor EB (TFEB) promotes protein degradation by the autophagy and lysosomal pathway (ALP) and overexpression of TFEB was suggested for the treatment of ALP-related diseases that often affect the heart. However, TFEB-mediated ALP induction may perturb cardiac stress response. We used adeno-associated viral vectors type 9 (AAV9) to overexpress TFEB (AAV9-Tfeb) or Luciferase-control (AAV9-Luc) in cardiomyocytes of 12-week-old male mice. Mice were subjected to transverse aortic constriction (TAC, 27G; AAV9-Luc: n = 9; AAV9-Tfeb: n = 14) or sham (AAV9-Luc: n = 9; AAV9-Tfeb: n = 9) surgery for 28 days. Heart morphology, echocardiography, gene expression, and protein levels were monitored. AAV9-Tfeb had no effect on cardiac structure and function in sham animals. TAC resulted in compensated left ventricular hypertrophy in AAV9-Luc mice. AAV9-Tfeb TAC mice showed a reduced LV ejection fraction and increased left ventricular diameters. Morphological, histological, and real-time PCR analyses showed increased heart weights, exaggerated fibrosis, and higher expression of stress markers and remodeling genes in AAV9-Tfeb TAC compared to AAV9-Luc TAC. RNA-sequencing, real-time PCR and Western Blot revealed a stronger ALP activation in the hearts of AAV9-Tfeb TAC mice. Cardiomyocyte-specific TFEB-overexpression promoted ALP gene expression during TAC, which was associated with heart failure. Treatment of ALP-related diseases by overexpression of TFEB warrants careful consideration.

Background: Men and women with valvular heart disease have different risk profiles for clinical endpoints. Non-esterified fatty acids (NEFA) are possibly involved in cardio-metabolic disease. However, it is unclear whether NEFA concentrations are associated with physical performance in patients undergoing transcatheter aortic valve implantation (TAVI) and whether there are sex-specific effects. Methods: To test the hypothesis that NEFA concentration is associated with sex-specific physical performance, we prospectively analysed data from one hundred adult patients undergoing TAVI. NEFA concentrations, physical performance and anthropometric parameters were measured before and 6 and 12 months after TAVI. Physical performance was determined by a six-minute walking test (6-MWT) and self-reported weekly bicycle riding time. Results: Before TAVI, NEFA concentrations were higher in patients (44 women, 56 men) compared to the normal population. Median NEFA concentrations at 6 and 12 months after TAVI were within the reference range reported in the normal population in men but not women. Men but not women presented with an increased performance in the 6-MWT over time (p = 0.026, p = 0.142, respectively). Additionally, men showed an increased ability to ride a bicycle after TAVI compared to before TAVI (p = 0.034). NEFA concentrations before TAVI correlated with the 6-MWT before TAVI in women (Spearman’s rho −0.552; p = 0.001) but not in men (Spearman’s rho −0.007; p = 0.964). No association was found between NEFA concentrations and physical performance 6 and 12 months after TAVI. Conclusions: NEFA concentrations improved into the reference range in men but not women after TAVI. Men but not women have an increased physical performance after TAVI. No association between NEFA and physical performance was observed in men and women after TAVI.

Supplemental Digital Content is available in the text., Background: Arterial hypertension and its organ sequelae show characteristics of T cell–mediated inflammatory diseases. Experimental anti-inflammatory therapies have been shown to ameliorate hypertensive end-organ damage. Recently, the CANTOS study (Canakinumab Antiinflammatory Thrombosis Outcome Study) targeting interleukin-1β demonstrated that anti-inflammatory therapy reduces cardiovascular risk. The gut microbiome plays a pivotal role in immune homeostasis and cardiovascular health. Short-chain fatty acids (SCFAs) are produced from dietary fiber by gut bacteria and affect host immune homeostasis. Here, we investigated effects of the SCFA propionate in 2 different mouse models of hypertensive cardiovascular damage. Methods: To investigate the effect of SCFAs on hypertensive cardiac damage and atherosclerosis, wild-type NMRI or apolipoprotein E knockout–deficient mice received propionate (200 mmol/L) or control in the drinking water. To induce hypertension, wild-type NMRI mice were infused with angiotensin II (1.44 mg·kg–1·d–1 subcutaneous) for 14 days. To accelerate the development of atherosclerosis, apolipoprotein E knockout mice were infused with angiotensin II (0.72 mg·kg–1·d–1 subcutaneous) for 28 days. Cardiac damage and atherosclerosis were assessed using histology, echocardiography, in vivo electrophysiology, immunofluorescence, and flow cytometry. Blood pressure was measured by radiotelemetry. Regulatory T cell depletion using PC61 antibody was used to examine the mode of action of propionate. Results: Propionate significantly attenuated cardiac hypertrophy, fibrosis, vascular dysfunction, and hypertension in both models. Susceptibility to cardiac ventricular arrhythmias was significantly reduced in propionate-treated angiotensin II–infused wild-type NMRI mice. Aortic atherosclerotic lesion area was significantly decreased in propionate-treated apolipoprotein E knockout–deficient mice. Systemic inflammation was mitigated by propionate treatment, quantified as a reduction in splenic effector memory T cell frequencies and splenic T helper 17 cells in both models, and a decrease in local cardiac immune cell infiltration in wild-type NMRI mice. Cardioprotective effects of propionate were abrogated in regulatory T cell–depleted angiotensin II–infused mice, suggesting the effect is regulatory T cell–dependent. Conclusions: Our data emphasize an immune-modulatory role of SCFAs and their importance for cardiovascular health. The data suggest that lifestyle modifications leading to augmented SCFA production could be a beneficial nonpharmacological preventive strategy for patients with hypertensive cardiovascular disease.

With more than 25 million people affected, heart failure (HF) is a global threat. As energy production pathways are known to play a pivotal role in HF, we sought here to identify key metabolic changes in ischemic- and non-ischemic HF by using a multi-OMICS approach. Serum metabolites and mRNAseq and epigenetic DNA methylation profiles were analyzed from blood and left ventricular heart biopsy specimens of the same individuals. In total we collected serum from n = 82 patients with Dilated Cardiomyopathy (DCM) and n = 51 controls in the screening stage. We identified several metabolites involved in glycolysis and citric acid cycle to be elevated up to 5.7-fold in DCM (p = 1.7 × 10−6). Interestingly, cardiac mRNA and epigenetic changes of genes encoding rate-limiting enzymes of these pathways could also be found and validated in our second stage of metabolite assessment in n = 52 DCM, n = 39 ischemic HF and n = 57 controls. In conclusion, we identified a new set of metabolomic biomarkers for HF. We were able to identify underlying biological cascades that potentially represent suitable intervention targets.

Background: Inflammatory processes are a cause of accelerated loss of muscle mass. Metabolic syndrome (MetS) is a highly prevalent age-related condition, which may promote and be promoted by inflammation. However, whether inflammation in MetS (metaflammation) is associated with lower muscle mass is still unclear. Methods: Complete cross-sectional data on body composition, MetS, and the inflammatory markers interleukin (IL)-1β, IL-6, IL-10, tumor necrosis factor (TNF), and C-reactive protein (CRP) were available for 1,377 BASE-II participants (51.1% women; 68 ± 4 years old). Appendicular lean mass (ALM) was assessed by dual-energy X-ray absorptiometry. Low muscle mass (low ALM-to-BMI ratio [ALMBMI]) was defined according to the Foundation for the National Institutes of Health (FNIH) Sarcopenia Project. Regression models, adjusted for an increasing number of confounders (sex, age, physical activity, morbidities, diabetes mellitus type II, TSH, albumin, HbA1c, smoking habits, alcohol intake, education, and energy intake/day), were used to calculate the association between low ALMBMI and high inflammation (tertile 3) according to MetS. Results: MetS was present in 36.2% of the study population, and 9% had low ALMBMI. In the whole study population, high CRP (odds ratio [OR]: 2.7 [95% CI: 1.6–4.7; p = 0.001]) and high IL-6 (OR: 2.1 [95% CI: 1.2–1.9; p = 0.005]) were associated with low ALMBMI. In contrast, no significant association was found between TNF, IL-10, or IL-1β with low ALMBMI. When participants were stratified by MetS, results for IL-6 remained significant only in participants with MetS. Conclusions: Among BASE-II participants, low ALMBMI was associated with inflammation. Low-grade inflammation triggered by disease state, especially in the context of MetS, might favor loss of muscle mass, so a better control of MetS might help to prevent sarcopenia. Intervention studies to test whether strategies to prevent MetS might also prevent loss of muscle mass seem to be promising.

Cachexia is associated with poor prognosis in chronic heart failure patients, but the underlying mechanisms of cachexia triggered disease progression remain poorly understood. Here, we investigate whether the dysregulation of myokine expression from wasting skeletal muscle exaggerates heart failure. RNA sequencing from wasting skeletal muscles of mice with heart failure reveals a reduced expression of Ostn, which encodes the secreted myokine Musclin, previously implicated in the enhancement of natriuretic peptide signaling. By generating skeletal muscle specific Ostn knock-out and overexpressing mice, we demonstrate that reduced skeletal muscle Musclin levels exaggerate, while its overexpression in muscle attenuates cardiac dysfunction and myocardial fibrosis during pressure overload. Mechanistically, Musclin enhances the abundance of C-type natriuretic peptide (CNP), thereby promoting cardiomyocyte contractility through protein kinase A and inhibiting fibroblast activation through protein kinase G signaling. Because we also find reduced OSTN expression in skeletal muscle of heart failure patients, augmentation of Musclin might serve as therapeutic strategy., Cachexia is associated with poor prognosis in heart failure. Here the authors show that mice and patients with cardiac cachexia display reduced skeletal muscle expression and circulating levels of Musclin. Musclin ablation in skeletal muscle worsens, while its muscle-specific overexpression ameliorates heart failure in mice.

Background Studying patients with rare Mendelian diabetes has uncovered molecular mechanisms regulating β‐cell pathophysiology. Previous studies have shown that Class IIa histone deacetylases (HDAC4, 5, 7, and 9) modulate mammalian pancreatic endocrine cell function and glucose homeostasis. Methods We performed exome sequencing in one adolescent nonautoimmune diabetic patient and detected one de novo predicted disease‐causing HDAC4 variant (p.His227Arg). We screened our pediatric diabetes cohort with unknown etiology using Sanger sequencing. In mouse pancreatic β‐cell lines (Min6 and SJ cells), we performed insulin secretion assay and quantitative RT‐PCR to measure the β‐cell function transfected with the detected HDAC4 variants and wild type. We carried out immunostaining and Western blot to investigate if the detected HDAC4 variants affect the cellular translocation and acetylation status of Forkhead box protein O1 (FoxO1) in the pancreatic β‐cells. Results We discovered three HDAC4 mutations (p.His227Arg, p.Asp234Asn, and p.Glu374Lys) in unrelated individuals who had nonautoimmune diabetes with various degrees of β‐cell loss. In mouse pancreatic β‐cell lines, we found that these three HDAC4 mutations decrease insulin secretion, down‐regulate β‐cell‐specific transcriptional factors, and cause nuclear exclusion of acetylated FoxO1. Conclusion Mutations in HDAC4 disrupt the deacetylation of FoxO1, subsequently decrease the β‐cell function including insulin secretion, resulting in diabetes.

OBJECTIVE: In sepsis, the disease course of critically ill patients is often complicated by muscle failure leading to ICU-acquired weakness. The myokine transforming growth factor-{beta}1 increases during inflammation and mediates muscle atrophy in vivo. We observed that the transforming growth factor-{beta}1 inhibitor, secreted frizzled-related protein 2, was down-regulated in skeletal muscle of ICU-acquired weakness patients. We hypothesized that secreted frizzled-related protein 2 reduction enhances transforming growth factor-{beta}1-mediated effects and investigated the interrelationship between transforming growth factor-{beta}1 and secreted frizzled-related protein 2 in inflammation-induced atrophy. DESIGN: Observational study and prospective animal trial. SETTING: Two ICUs and research laboratory. PATIENTS/SUBJECTS: Twenty-six critically ill patients with Sequential Organ Failure Assessment scores greater than or equal to 8 underwent a skeletal muscle biopsy from the vastus lateralis at median day 5 in ICU. Four patients undergoing elective orthopedic surgery served as controls. To search for signaling pathways enriched in muscle of ICU-acquired weakness patients, a gene set enrichment analysis of our recently published gene expression profiles was performed. Quantitative reverse transcriptase-polymerase chain reaction, Western blot, and immunohistochemistry were used to analyze secreted frizzled-related protein 2 expression and protein content. A mouse model of inflammation-induced skeletal muscle atrophy due to polymicrobial sepsis and cultured myocytes were used for mechanistic analyses. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Gene set enrichment analysis uncovered transforming growth factor-{beta}1 signaling activation in vastus lateralis from ICU-acquired weakness patients. Muscular secreted frizzled-related protein 2 expression was reduced after 5 days in ICU. Likewise, muscular secreted frizzled-related protein 2 expression was decreased early and continuously in mice with inflammation-induced atrophy. In muscle, secreted frizzled-related protein 2 was predominantly contained in fast twitch/type II myofibers. Secreted frizzled-related protein 2 physically interacted and colocalized with transforming growth factor-{beta}1 through its cysteine-rich domain. Finally, secreted frizzled-related protein 2 prevented transforming growth factor-{beta}1-induced atrophy in C2C12 myotubes. CONCLUSIONS: Muscular secreted frizzled-related protein 2 is down-regulated in ICU-acquired weakness patients and mice with inflammation-induced muscle atrophy. Decreased secreted frizzled-related protein 2 possibly establishes a positive feedback loop enhancing transforming growth factor-{beta}1-mediated atrophic effects in inflammation-induced atrophy.

Cachexia is a life threatening syndrome associated with several diseases, such as end‐stage heart failure, end‐stage renal disease, chronic obstructive pulmonary disease, chronic inflammation (i.e. rheumatoid arthritis), acquired immune deficiency syndrome, and cancer.1, 2 Cachexia is found in 31–87% of cancer patients especially in advanced disease stages.3 It is characterized by progressive weight loss, metabolic alterations, fatigue, and persistent reduction of body cell mass in response to a malignant tumour.2, 4, 5, 6 The incidence of cachexia in cancer patients is dependent on the type and site of the tumour. While patients with non‐Hodgkin's lymphoma, breast cancer, and sarcomas show low incidences, rates up to 83% in pancreatic cancer patients, and over 85% in patients with gastric cancer have been found. Additionally, around 60% of small‐cell and non‐small‐cell lung cancer patients develop cachexia.7, 8, 9 Cancer cachexia affects the function of several organs such as muscle, adipose tissue, liver, brain, immune system, and heart, collectively decreasing patients' quality of life and worsening their prognosis. Therefore, cachexia must be considered as a true multi‐organ syndrome.10 Because cancer cachexia leads to a decrease in physical performance and quality of life,11 and is associated with poor survival (accounting for more than 20% of cancer deaths,7, 12, 13, 14, 15) it is of major clinical relevance. Even more so since cachectic patients show lower response rates to chemotherapy7 and a reduced tolerance to anticancer treatment.16 Despite its importance, weight loss in cancer patients is rarely recognized, assessed,17 or treated actively.18, 19 Thus, cancer cachexia represents an important underappreciated clinical syndrome.

BACKGROUND Cardiodepressant antibodies contribute to cardiac dysfunction in dilated cardiomyopathy (DCM). Changes in immunoglobulin G (IgG) glycosylation modulate the activity of various autoimmune diseases and influence disease activity as well as severity of various autoimmune diseases. We hypothesized that alterations in IgG glycosylation are involved in the disease course of DCM. METHODS AND RESULTS IgG glycosylation was analyzed in plasma samples of 50 DCM patients using a lectin-based ELISA. Negative inotropic (cardiodepressant) activity (NIA) of antibodies was assessed by measuring the effect of purified DCM-IgG on the shortening of isolated rat cardiomyocytes by means of a video-edge detection system. IgG obtained from plasma of healthy blood donors served as control. DCM-IgG contained significantly less sialic acid (-25%) and galactose (-16%; both P

The muscle-specific RING-finger protein MuRF1 (also known as TRIM63) constitutes a bona fide ubiquitin ligase that routes proteins like several different myosin heavy chain proteins (MyHC) to proteasomal degradation during muscle atrophy. In two unbiased screens, we identified DCAF8 as a new MuRF1-binding partner. MuRF1 physically interacts with DCAF8 and both proteins localize to overlapping structures in muscle cells. Importantly, similar to what is seen for MuRF1, DCAF8 levels increase during atrophy, and the downregulation of either protein substantially impedes muscle wasting and MyHC degradation in C2C12 myotubes, a model system for muscle differentiation and atrophy. DCAF proteins typically serve as substrate receptors for cullin 4-type (Cul4) ubiquitin ligases (CRL), and we demonstrate that DCAF8 and MuRF1 associate with the subunits of such a protein complex. Because genetic downregulation of DCAF8 and inhibition of cullin activity also impair myotube atrophy in C2C12 cells, our data imply that the DCAF8 promotes muscle wasting by targeting proteins like MyHC as an integral substrate receptor of a Cul4A-containing ring ubiquitin ligase complex (CRL4A).This article has an associated First Person interview with the first author of the paper.

Spinal cord injury (SCI) above mid-thoracic levels leads to autonomic dysfunction affecting both the cardiovascular system and thermoregulation. The renin-angiotensin system (RAS) which is a potent regulator of blood pressure, including its novel beneficial arm with the receptor Mas could be an interesting target in post-SCI hemodynamics. To test the hypothesis that hemodynamics, activity and diurnal patterns of those are more affected in the Mas deficient mice post-SCI we used a mouse model of SCI with complete transection of spinal cord at thoracic level 4 (T4-Tx) and performed telemetric monitoring of blood pressure (BP) and heart rate (HR). Our data revealed that hypothermia deteriorated physiological BP and HR control. Preserving normothermia by keeping mice at 30°C prevented severe hypotension and bradycardia post-SCI. Moreover, it facilitated rapid return of diurnal regulation of BP, HR and activity in wild type (WT) mice. In contrast, although Mas deficient mice had comparable reacquisition of diurnal HR rhythm, they showed delayed recovery of diurnal rhythmicity in BP and significantly lower nocturnal activity. Exposing mice with T4-Tx (kept in temperature-controlled cages) to 23°C room temperature for one hour at different time-points post-SCI, demonstrated their inability to maintain core body temperature, Mas deficient mice being significantly more impaired than WT littermates. We conclude that Mas deficient mice were more resistant to acute hypotension, delayed nocturnal recovery, lower activity and more severely impaired thermoregulation. The ambient temperature had significant effect on hemodynamics and, thus it should be taken into account when assessing cardiovascular parameters post-SCI in mice.

Objective: Inflammation drives cardiovascular disease, anti-inflammatory approaches may be beneficial. Short-chain fatty acids (SCFA) are bacterial metabolites with anti-inflammatory properties affecting host immune homeostasis including regulatory T cells (Treg). We investigated effects of the SCFA propionate (administered in drinking water, NaCl as control) in two mouse models, namely hypertensive heart disease (wild-type NMRI (WT), angiotensin (Ang)II infusion 1.44 mg/kg/d s.c. for 14 days) and atherosclerosis (Apolipoprotein E knockout (ApoE), AngII infusion 0.72 mg/kg/d s.c. for 28 days), respectively. Results: Propionate attenuated cardiac hypertrophy and fibrosis in both models significantly. Susceptibility to cardiac ventricular arrhythmias was significantly reduced in propionate-treated WT mice. Aortic atherosclerotic lesion area was significantly reduced in propionate-treated ApoE (27.6±8 vs. 7.9±2.4%). Treatment reduced splenic effector memory (CD4+ CD44+ CD62L-) T cell frequencies (WT: 30.5±4.6 vs. 19.1±1.6; ApoE: 41.1±3.1 vs. 32.7±1.4%) and splenic Th17 cells (WT: 1.0±0.2 vs. 0.6±0.1; ApoE: 1.3±0.1 vs. 0.9±0.1%) in both models, indicating beneficial effects on systemic inflammation. Similarly, propionate reduced cardiac immune cell infiltration (CD4+, CD8+, F4/80+) in WT mice. Propionate improved vascular dysfunction and moderately reduced blood pressure in both models. Organ-protective actions of propionate (cardiac inflammation and fibrosis) were abrogated in Treg-depleted (antiCD25-treated) AngII-infused WT mice, suggesting a central role for Treg. To verify our findings in a human cohort, we re-analyzed clinical and metagenomic data from a recent randomized controlled trial investigating the effect of a 90-day synbiotic intervention in 84 subjects with metabolic syndrome including healthy controls. Interestingly, in synbiotic-treated subjects an increased capacity for SFCA production was significantly correlated to blood pressure reduction. Conclusion: Data underscore the importance of SCFA for cardiovascular health and suggest that lifestyle modifications leading to augmented SCFA production could be a beneficial non-pharmacological add-on strategy for cardiovascular disease.

Background The Muscle-specific RING-finger (MuRF) protein family of E3 ubiquitin ligases is important for maintenance of muscular structure and function. MuRF proteins mediate adaptation of striated muscles to stress. MuRF2 and MuRF3 bind to microtubules and are implicated in sarcomere formation with noticeable functional redundancy. However, if this redundancy is important for muscle function in vivo is unknown. Our objective was to investigate cooperative function of MuRF2 and MuRF3 in the skeletal muscle and the heart in vivo. Methods MuRF2 and MuRF3 double knockout mice (DKO) were generated and phenotypically characterized. Skeletal muscle and the heart were investigated by morphological measurements, histological analyses, electron microscopy, immunoblotting, and real-time PCR. Isolated muscles were subjected to in vitro force measurements. Cardiac function was determined by echocardiography and working heart preparations. Function of cardiomyocytes was measured in vitro. Cell culture experiments and mass-spectrometry were used for mechanistic analyses. Results DKO mice showed a protein aggregate myopathy in skeletal muscle. Maximal force development was reduced in DKO soleus and extensor digitorum longus. Additionally, a fibre type shift towards slow/type I fibres occurred in DKO soleus and extensor digitorum longus. MuRF2 and MuRF3-deficient hearts showed decreased systolic and diastolic function. Further analyses revealed an increased expression of the myosin heavy chain isoform beta/slow and disturbed calcium handling as potential causes for the phenotype in DKO hearts. Conclusions The redundant function of MuRF2 and MuRF3 is important for maintenance of skeletal muscle and cardiac structure and function in vivo.

Excitotoxicity, a critical process in neurodegeneration, induces oxidative stress and neuronal death through mechanisms largely unknown. Since oxidative stress activates protein kinase D1 (PKD1) in tumor cells, we investigated the effect of excitotoxicity on neuronal PKD1 activity. Unexpectedly, we find that excitotoxicity provokes an early inactivation of PKD1 through a dephosphorylation-dependent mechanism mediated by protein phosphatase-1 (PP1) and dual specificity phosphatase-1 (DUSP1). This step turns off the IKK/NF-κB/SOD2 antioxidant pathway. Neuronal PKD1 inactivation by pharmacological inhibition or lentiviral silencing in vitro, or by genetic inactivation in neurons in vivo, strongly enhances excitotoxic neuronal death. In contrast, expression of an active dephosphorylation-resistant PKD1 mutant potentiates the IKK/NF-κB/SOD2 oxidative stress detoxification pathway and confers neuroprotection from in vitro and in vivo excitotoxicity. Our results indicate that PKD1 inactivation underlies excitotoxicity-induced neuronal death and suggest that PKD1 inactivation may be critical for the accumulation of oxidation-induced neuronal damage during aging and in neurodegenerative disorders.

Progression of idiopathic dilated cardiomyopathy (IDCM) is marked with extensive left ventricular remodeling whose clinical manifestations and molecular basis are poorly understood. We aimed to evaluate the clinical potential of titin ligands in monitoring progression of cardiac remodeling associated with end-stage IDCM. Expression patterns of 8 mechanoptotic machinery-associated titin ligands (ANKRD1,ANKRD2,TRIM63,TRIM55,NBR1,MLP,FHL2, andTCAP) were quantitated in endomyocardial biopsies from 25 patients with advanced IDCM. When comparing NYHA disease stages, elevatedANKRD1expression levels marked transition from NYHA < IV to NYHA IV.ANKRD1expression levels closely correlated with systolic strain depression and short E wave deceleration time, as determined by echocardiography. On molecular level, myocardialANKRD1and serum adiponectin correlated with lowBAX/BCL-2ratios, indicative of antiapoptotic tissue propensity observed during the worsening of heart failure. ANKRD1 is a potential marker for cardiac remodeling and disease progression in IDCM.ANKRD1expression correlated with reduced cardiac contractility and compliance. The association ofANKRD1with antiapoptotic response suggests its role as myocyte survival factor during late stage heart disease, warranting further studies on ANKRD1 during end-stage heart failure.

Excitotoxicity, a critical process in neurodegeneration, induces oxidative stress and neuronal death through mechanisms largely unknown. Since oxidative stress activates protein kinase D1 (PKD1) in tumor cells, we investigated the effect of excitotoxicity on neuronal PKD1 activity. Unexpectedly, we find that excitotoxicity provokes an early inactivation of PKD1 through a dephosphorylation-dependent mechanism mediated by protein phosphatase-1 (PP1) and dual specificity phosphatase-1 (DUSP1). This step turns off the IKK/NF-¿B/SOD2 antioxidant pathway. Neuronal PKD1 inactivation by pharmacological inhibition or lentiviral silencing in vitro, or by genetic inactivation in neurons in vivo, strongly enhances excitotoxic neuronal death. In contrast, expression of an active dephosphorylation-resistant PKD1 mutant potentiates the IKK/NF-¿B/SOD2 oxidative stress detoxification pathway and confers neuroprotection from in vitro and in vivo excitotoxicity. Our results indicate that PKD1 inactivation underlies excitotoxicity-induced neuronal death and suggest that PKD1 inactivation may be critical for the accumulation of oxidation-induced neuronal damage during aging and in neurodegenerative disorders., This work was supported by grants SAF2014-52737-P to T.I., SAF2013-45258-P to M.R. C., BFU2016-77885-P to F.H., SAF2014-54070-JIN to A.M. from Ministerio de Economía, Industria y Competitividad (Spain). It was also funded by P2010/BMD-2332 (Neurodegmodels) from Comunidad de Madrid to T.I., F.H. and J.A.) and Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED, Instituto de Salud Carlos III, Spain) to T.I., F.H., J.A., and I.F.). J.P.-U. is a recipient of a predoctoral contract from SAF2014-52737-P; L.G.-G. was funded by a contract from CIBERNED; A.G.-M. and A.S.S. were funded by contracts from CIBERNED cooperative projects 2013/07 and CIBERNED 2015-2/06, respectively. A.M., A.D.P. is a recipient of a Juan de la Cierva formación fellowship (Ministerio de Economía, Industria y Competitividad, Spain) associated to CIBERNED. J.J.-A. was funded by a predoctoral contract from CSIC (JAEPredoc program) and by CIBERNED. The cost of this publication has been paid in part by FEDER (European Funds for Regional Development) funds.

Ββ-Cell adaptation to insulin resistance is necessary to maintain glucose homeostasis in obesity. Failure of this mechanism is a hallmark of type 2 diabetes (T2D). Hence, factors controlling functional β-cell compensation are potentially important targets for the treatment of T2D. Protein kinase D1 (PKD1) integrates diverse signals in the β-cell and plays a critical role in the control of insulin secretion. However, the role of β-cell PKD1 in glucose homeostasis in vivo is essentially unknown. Using β-cell–specific, inducible PKD1 knockout mice (βPKD1KO), we examined the role of β-cell PKD1 under basal conditions and during high-fat feeding. βPKD1KO mice under a chow diet presented no significant difference in glucose tolerance or insulin secretion compared with mice expressing the Cre transgene alone; however, when compared with wild-type mice, both groups developed glucose intolerance. Under a high-fat diet, deletion of PKD1 in β-cells worsened hyperglycemia, hyperinsulinemia, and glucose intolerance. This was accompanied by impaired glucose-induced insulin secretion both in vivo in hyperglycemic clamps and ex vivo in isolated islets from high-fat diet–fed βPKD1KO mice without changes in islet mass. This study demonstrates an essential role for PKD1 in the β-cell adaptive secretory response to high-fat feeding in mice.

Intensive care unit (ICU)-acquired muscle wasting is a devastating complication leading to persistent weakness and functional disability. The mechanisms of this myopathy are unclear, but a disturbed balance of myosin heavy chain (MyHC) is implicated.To investigate pathways of myosin turnover in severe critically ill patients at high risk of ICU-acquired weakness.Prospective, mechanistic, observational study.Interdisciplinary ICUs of a university hospital.Twenty-nine patients with Sequential Organ Failure Assessment (SOFA) scores of at least 8 on three consecutive days within the first 5 days in ICU underwent two consecutive open skeletal muscle biopsies from the vastus lateralis at median days 5 and 15. Control biopsy specimens were from healthy subjects undergoing hip-replacement surgery.None.Time-dependent changes in myofiber architecture, MyHC synthesis, and degradation were determined and correlated with clinical data.ICU-acquired muscle wasting was characterized by early, disrupted myofiber ultrastructure followed by atrophy of slow- and fast-twitch myofibers at later time points. A rapid decrease in MyHC mRNA and protein expression occurred by day 5 and persisted at day 15 (P0.05). Expression of the atrophy genes MuRF-1 and Atrogin1 was increased at day 5 (P0.05). Early MuRF-1 protein content was closely associated with late myofiber atrophy and the severity of weakness.Decreased synthesis and increased degradation of MyHCs contribute to ICU-acquired muscle wasting. The rates and time frames suggest that pathogenesis of muscle failure is initiated very early during critical illness. The persisting reduction of MyHC suggests that sustained treatment is required.

Brain activity requires a flux of glucose to active regions to sustain increased metabolic demands. Insulin, the main regulator of glucose handling in the body, has been traditionally considered not to intervene in this process. However, we now report that insulin modulates brain glucose metabolism by acting on astrocytes in concert with IGF-I. The cooperation of insulin and IGF-I is needed to recover neuronal activity after hypoglycemia. Analysis of underlying mechanisms show that the combined action of IGF-I and insulin synergistically stimulates a mitogen-activated protein kinase/protein kinase D pathway resulting in translocation of GLUT1 to the cell membrane through multiple protein-protein interactions involving the scaffolding protein GAIP-interacting protein C terminus and the GTPase RAC1. Our observations identify insulin-like peptides as physiological modulators of brain glucose handling, providing further support to consider the brain as a target organ in diabetes., This work was funded by MINECO (Spain) grants SAF2010-60051 and SAF2013-40710-R, and by CIBERNED. E.H.-G. was partially funded by a fellowship from ColFuturo and CIBERNED. T.I. is funded by MINECO (SAF2014-52737-P). T.I. and I.T.A. are funded by CIBERNED and Comunidad de Madrid, Spain (P2010/BMD-2331-Neurodegmodels).