Characterization of Biomarkers of Thrombo-Inflammation in Patients with First-Diagnosed Atrial Fibrillation.

Patients with first-diagnosed atrial fibrillation (FDAF) exhibit major adverse cardiovascular events (MACEs) during follow-up. Preclinical models have demonstrated that thrombo-inflammation mediates adverse cardiac remodeling and atherothrombotic events. We have hypothesized that thrombin activity (FIIa) links coagulation with inflammation and cardiac fibrosis/dysfunction. Surrogate markers of the thrombo-inflammatory response in plasma have not been characterized in FDAF. In this prospective longitudinal study, patients presenting with FDAF (n = 80), and 20 matched controls, were included. FIIa generation and activity in plasma were increased in the patients with early AF compared to the patients with chronic cardiovascular disease without AF (controls; p < 0.0001). This increase was accompanied by elevated biomarkers (ELISA) of platelet and endothelial activation in plasma. Pro-inflammatory peripheral immune cells (TNF-α+ or IL-6+) that expressed FIIa-activated protease-activated receptor 1 (PAR1) (flow cytometry) circulated more frequently in patients with FDAF compared to the controls (p < 0.0001). FIIa activity correlated with cardiac fibrosis (collagen turnover) and cardiac dysfunction (NT-pro ANP/NT-pro BNP) surrogate markers. FIIa activity in plasma was higher in patients with FDAF who experienced MACE. Signaling via FIIa might be a presumed link between the coagulation system (tissue factor-FXa/FIIa-PAR1 axis), inflammation, and pro-fibrotic pathways (thrombo-inflammation) in FDAF.

Being Overweight or Obese Is Associated with an Increased Platelet Reactivity Despite Dual Antiplatelet Therapy with Aspirin and Clopidogrel.

PURPOSE: Obese patients exhibit an overall increased platelet reactivity and a reduced sensitivity to antiplatelet therapy. The aim of this study is to evaluate the platelet reactivity measured by impedance aggregometry in overweight and obese patients and chronic coronary syndrome (CCS) that were treated with dual antiplatelet therapy (DAPT). METHODS: Platelet aggregation was assessed by impedance aggregometry in patients with CCS receiving DAPT (aspirin plus clopidogrel). We compared the platelet reactivity in patients with a normal weight versus overweight or obese patients. Furthermore, the correlation between the body mass index (BMI) and adenosine diphosphate- (ADP-) or thrombin receptor-activating peptide- (TRAP-) dependent platelet aggregation was analyzed. RESULTS: 64 patients were included in the study of which 35.9% were patients with normal weight. A higher ADP- and TRAP-dependent platelet reactivity was observed in overweight and obese patients (ADP: median 27 units (U) [IQR 13-39.5] vs. 7 U [6-15], p < 0.001 and TRAP: 97 U [73-118.5] vs. 85 U [36-103], p = 0.035). Significant positive correlations were observed between agonist-induced platelet reactivity and BMI. CONCLUSION: Despite the use of DAPT, a higher platelet reactivity was found in overweight and obese patients with CCS. If these patients will benefit from treatment with more potent platelet inhibitors, it needs to be evaluated in future clinical trials.

Galectin-3 as a Marker for Increased Thrombogenicity in COVID-19.

Galectin-3 is a beta-galactoside-binding lectin involved in inflammation and lung fibrosis and postulated to enhance thrombosis. In COVID-19, it is considered to be a prognostic marker of severity. The aim of this study was to evaluate whether galectin-3 is associated with thrombogenicity in COVID-19. Patients with moderate-to-severe COVID-19 (COVpos; n = 55) and patients with acute respiratory diseases, but without COVID-19 (COVneg; n = 35), were included in the study. We measured the amount of galectin-3, as well as other platelet and coagulation markers, and correlated galectin-3 levels with these markers of thrombogenicity and with the SOFA Score values. We found that galectin-3 levels, as well as von Willebrand Factor (vWF), antithrombin and tissue plasminogen activator levels, were higher in the COVpos than they were in the COVneg cohort. Galectin-3 correlated positively with vWF, antithrombin and D-dimer in the COVpos cohort, but not in the COVneg cohort. Moreover, galactin-3 correlated also with clinical disease severity, as measured by the SOFA Score. In patients with acute respiratory diseases, galectin-3 can be considered as a marker not only for disease severity, but also for increased hypercoagulability. Whether galectin-3 might be a useful therapeutic target in COVID-19 needs to be assessed in future studies.

Uptake of Tropheryma whipplei by Intestinal Epithelia.

BACKGROUND: Tropheryma whipplei (TW) can cause different pathologies, e.g., Whipple's disease and transient gastroenteritis. The mechanism by which the bacteria pass the intestinal epithelial barrier, and the mechanism of TW-induced gastroenteritis are currently unknown. METHODS: Using ex vivo disease models comprising human duodenal mucosa exposed to TW in Ussing chambers, various intestinal epithelial cell (IEC) cultures exposed to TW and a macrophage/IEC coculture model served to characterize endocytic uptake mechanisms and barrier function. RESULTS: TW exposed ex vivo to human small intestinal mucosae is capable of autonomously entering IECs, thereby invading the mucosa. Using dominant-negative mutants, TW uptake was shown to be dynamin- and caveolin-dependent but independent of clathrin-mediated endocytosis. Complementary inhibitor experiments suggested a role for the activation of the Ras/Rac1 pathway and actin polymerization. TW-invaded IECs underwent apoptosis, thereby causing an epithelial barrier defect, and were subsequently subject to phagocytosis by macrophages. CONCLUSIONS: TW enters epithelia via an actin-, dynamin-, caveolin-, and Ras-Rac1-dependent endocytosis mechanism and consecutively causes IEC apoptosis primarily in IECs invaded by multiple TW bacteria. This results in a barrier leak. Moreover, we propose that TW-packed IECs can be subject to phagocytic uptake by macrophages, thereby opening a potential entry point of TW into intestinal macrophages.

Metformin Is Associated with Reduced Tissue Factor Procoagulant Activity in Patients with Poorly Controlled Diabetes.

PURPOSE: Metformin is the first-line antidiabetic drug and shown to reduce cardiovascular risk independent from its glucose lowering action. Particularly in poorly controlled diabetes, tissue factor (TF) is expressed in the vasculature and accounts for thromboembolic complications. Here, we aimed to assess the effect of metformin on TF activity and markers of vascular inflammation in poorly controlled type 2 diabetes. METHODS: In a cohort of patients with uncontrolled type 2 diabetes (glycosylated hemoglobin 8.39 ± 0.24%, 68.1 ± 2.6 mmol/mol, n = 46) of whom half of the individuals were treated with metformin and the other half did not receive metformin as part of an anti-diabetic combination therapy, we assessed TF activity and markers of vascular inflammation. In vitro, human monocytic cells (THP-1) were exposed to metformin and TF expression measured in the presence and absence of the AMP-activated protein kinase (AMPK) activator 5-aminoimidazole-4-carboxamide riboside (AICAR) or the AMPK inhibitor compound C. RESULTS: In the patients, metformin treatment was associated with lower levels of TF protein (241.5 ± 19 vs. 315.4 ± 25 pg/mL, p = 0.03) and reduced TF activity (408.9 ± 49 vs. 643.8 ± 47 U/mL, p = 0.001) compared with controls. Moreover, the patients on metformin showed lower levels of vascular cell adhesion molecule (VCAM)1 (26.6 ± 1.4 vs. 35.03 ± 3.1 ng/mL, p = 0.014) and higher expression of miR-126-3p/U6sno (11.39 ± 2.8 vs. 4.26 ± 0.9, p = 0.006), a known post-transcriptional down regulator of TF and VCAM1. In vitro, metformin dose-dependently reduced lipopolysaccharide (LPS)-induced TF expression in THP-1 cells. The AMPK activator AICAR alone lowered TF expression in THP-1, while the AMPK inhibitor compound C abrogated the metformin-dependent reduction in TF expression. CONCLUSIONS: Our data are the first to report that metformin is associated with reduced plasma TF procoagulant activity possibly explaining-at least in part-the vasculoprotective properties of metformin.

Vascular miR-181b controls tissue factor-dependent thrombogenicity and inflammation in type 2 diabetes.

BACKGROUND: Diabetes mellitus is characterized by chronic vascular inflammation leading to pathological expression of the thrombogenic full length (fl) tissue factor (TF) and its isoform alternatively-spliced (as) TF. Blood-borne TF promotes factor (F) Xa generation resulting in a pro-thrombotic state and cardiovascular complications. MicroRNA (miR)s impact gene expression on the post-transcriptional level and contribute to vascular homeostasis. Their distinct role in the control of the diabetes-related procoagulant state remains poorly understood. METHODS: In a cohort of patients with poorly controlled type 2 diabetes (n = 46) plasma levels of miR-181b were correlated with TF pathway activity and markers for vascular inflammation. In vitro, human microvascular endothelial cells (HMEC)-1 and human monocytes (THP-1) were transfected with miR-181b or anti-miR-181b and exposed to tumor necrosis factor (TNF) α or lipopolysaccharides (LPS). Expression of TF isoforms, vascular adhesion molecule (VCAM) 1 and nuclear factor (NF) κB nuclear translocation was assessed. Moreover, aortas, spleen, plasma, and bone marrow-derived macrophage (BMDM)s of mice carrying a deletion of the first miR-181b locus were analyzed with respect to TF expression and activity. RESULTS: In patients with type 2 diabetes, plasma miR-181b negatively correlated with the procoagulant state as evidenced by TF protein, TF activity, D-dimer levels as well as markers for vascular inflammation. In HMEC-1, miR-181b abrogated TNFα-induced expression of flTF, asTF, and VCAM1. These results were validated using the anti-miR-181b. Mechanistically, we confirmed a miR-181b-mediated inhibition of importin-α3 (KPNA4) leading to reduced nuclear translocation of the TF transcription factor NFκB. In THP-1, miR-181b reduced both TF isoforms and FXa generation in response to LPS due to targeting phosphatase and tensin homolog (PTEN), a principal inducer for TF in monocytes. Moreover, in miR-181-/- animals, we found that reduced levels of miR-181b were accompanied by increased TF, VCAM1, and KPNA4 expression in aortic tissue as well as increased TF and PTEN expression in spleen. Finally, BMDMs of miR-181-/- mice showed increased TF expression and FXa generation upon stimulation with LPS. CONCLUSIONS: miR-181b epigenetically controls the procoagulant state in diabetes. Reduced miR-181b levels contribute to increased thrombogenicity and may help to identify individuals at particular risk for thrombosis.

Protease-activated receptor 2 deficiency mediates cardiac fibrosis and diastolic dysfunction.

AIMS: Heart failure with preserved ejection fraction (HFpEF) and pathological cardiac aging share a complex pathophysiology, including extracellular matrix remodelling (EMR). Protease-activated receptor 2 (PAR2) deficiency is associated with EMR. The roles of PAR1 and PAR2 have not been studied in HFpEF, age-dependent cardiac fibrosis, or diastolic dysfunction (DD). METHODS AND RESULTS: Evaluation of endomyocardial biopsies from patients with HFpEF (n = 14) revealed that a reduced cardiac PAR2 expression was associated with aggravated DD and increased myocardial fibrosis (r = -0.7336, P = 0.0028). In line, 1-year-old PAR2-knockout (PAR2ko) mice suffered from DD with preserved systolic function, associated with an increased age-dependent α-smooth muscle actin expression, collagen deposition (1.7-fold increase, P = 0.0003), lysyl oxidase activity, collagen cross-linking (2.2-fold increase, P = 0.0008), endothelial activation, and inflammation. In the absence of PAR2, the receptor-regulating protein caveolin-1 was down-regulated, contributing to an augmented profibrotic PAR1 and transforming growth factor beta (TGF-ß)-dependent signalling. This enhanced TGF-ß/PAR1 signalling caused N-proteinase (ADAMTS3) and C-proteinase (BMP1)-related increased collagen I production from cardiac fibroblasts (CFs). PAR2 overexpression in PAR2ko CFs reversed these effects. The treatment with the PAR1 antagonist, vorapaxar, reduced cardiac fibrosis by 44% (P = 0.03) and reduced inflammation in a metabolic disease model (apolipoprotein E-ko mice). Patients with HFpEF with upstream PAR inhibition via FXa inhibitors (n = 40) also exhibited reduced circulating markers of fibrosis and DD compared with patients treated with vitamin K antagonists (n = 20). CONCLUSIONS: Protease-activated receptor 2 is an important regulator of profibrotic PAR1 and TGF-ß signalling in the heart. Modulation of the FXa/FIIa-PAR1/PAR2/TGF-ß-axis might be a promising therapeutic approach to reduce HFpEF.

MicroRNA-19a contributes to the epigenetic regulation of tissue factor in diabetes.

BACKGROUND: Diabetes mellitus is characterized by chronic vascular disorder and presents a main risk factor for cardiovascular mortality. In particular, hyperglycaemia and inflammatory cytokines induce vascular circulating tissue factor (TF) that promotes pro-thrombotic conditions in diabetes. It has recently become evident that alterations of the post-transcriptional regulation of TF via specific microRNA(miR)s, such as miR-126, contribute to the pathogenesis of diabetes and its complications. The endothelial miR-19a is involved in vascular homeostasis and atheroprotection. However, its role in diabetes-related thrombogenicity is unknown. Understanding miR-networks regulating procoagulability in diabetes may help to develop new treatment options preventing vascular complications. METHODS AND RESULTS: Plasma of 44 patients with known diabetes was assessed for the expression of miR-19a, TF protein, TF activity, and markers for vascular inflammation. High miR-19a expression was associated with reduced TF protein, TF-mediated procoagulability, and vascular inflammation based on expression of vascular adhesion molecule-1 and leukocyte count. We found plasma expression of miR-19a to strongly correlate with miR-126. miR-19a reduced the TF expression on mRNA and protein level in human microvascular endothelial cells (HMEC) as well as TF activity in human monocytes (THP-1), while anti-miR-19a increased the TF expression. Interestingly, miR-19a induced VCAM expression in HMEC. However, miR-19a and miR-126 co-transfection reduced total endothelial VCAM expression and exhibited additive inhibition of a luciferase reporter construct containing the F3 3'UTR. CONCLUSIONS: While both miRs have differential functions on endothelial VCAM expression, miR-19a and miR-126 cooperate to exhibit anti-thrombotic properties via regulating vascular TF expression. Modulating the post-transcriptional control of TF in diabetes may provide a future anti-thrombotic and anti-inflammatory therapy.

Role of dendritic cells in the pathogenesis of Whipple's disease.

Accumulation of Tropheryma whipplei-stuffed macrophages in the duodenum, impaired T. whipplei-specific Th1 responses, and weak secretion of interleukin-12 (IL-12) are hallmarks of classical Whipple's disease (CWD). This study addresses dendritic cell (DC) functionality during CWD. We documented composition, distribution, and functionality of DC ex vivo or after in vitro maturation by fluorescence-activated cell sorting (FACS) and by immunohistochemistry in situ. A decrease in peripheral DC of untreated CWD patients compared to healthy donors was due to reduced CD11c(high) myeloid DC (M-DC). Decreased maturation markers CD83, CD86, and CCR7, as well as low IL-12 production in response to stimulation, disclosed an immature M-DC phenotype. In vitro-generated monocyte-derived DC from CWD patients showed normal maturation and T cell-stimulatory capacity under proinflammatory conditions but produced less IL-12 and failed to activate T. whipplei-specific Th1 cells. In duodenal and lymphoid tissues, T. whipplei was found within immature DC-SIGN(+) DC. DC and proliferating lymphocytes were reduced in lymph nodes of CWD patients compared to levels in controls. Our results indicate that dysfunctional IL-12 production by DC provides suboptimal conditions for priming of T. whipplei-specific T cells during CWD and that immature DC carrying T. whipplei contribute to the dissemination of the bacterium.

Immunopathology of immune reconstitution inflammatory syndrome in Whipple's disease.

During antimicrobial treatment of classic Whipple's disease (CWD), the chronic systemic infection with Tropheryma whipplei, immune reconstitution inflammatory syndrome (IRIS), is a serious complication. The aim of our study was to characterize the immunological processes underlying IRIS in CWD. Following the definition of IRIS, we describe histological features of IRIS and immunological parameters of 24 CWD IRIS patients, 189 CWD patients without IRIS, and 89 healthy individuals. T cell reconstitution, Th1 reactivity, and the phenotype of T cells were described in the peripheral blood, and infiltration of CD4(+) T cells and regulatory T cells in the duodenal mucosa was determined. During IRIS, tissues were heavily infiltrated by CD3(+), predominantly CD45RO(+)CD4(+) T cells. In the periphery, initial reduction of CD4(+) cell counts and their reconstitution on treatment was more pronounced in CWD patients with IRIS than in those without IRIS. The ratio of activated and regulatory CD4(+) T cells, nonspecific Th1 reactivity, and the proportion of naive among CD4(+) T cells was high, whereas serum IL-10 was low during IRIS. T. whipplei-specific Th1 reactivity remained suppressed before and after emergence of IRIS. The findings that IRIS in CWD mainly are mediated by nonspecific activation of CD4(+) T cells and that it is not sufficiently counterbalanced by regulatory T cells indicate that flare-up of pathogen-specific immunoreactivity is not instrumental in the pathogenesis of IRIS in CWD.

Hand hygiene in emergencies: Multiprofessional perceptions from a mixed methods based online survey in Germany.

Introduction: Despite high vulnerability to infection, hand disinfection compliance in emergencies is low. This is regularly justified as the disinfection procedure delays life support, and instead, wearing disposable gloves is preferred. Simulation studies showed higher achievable compliance than detected in real-life situations. This study aimed to explore healthcare providers' attitudes toward hand disinfection and using gloves in emergencies. Methods: We conducted an anonymous online survey in Germany on the attitude and subjective behavior in the five moments of hand hygiene in a closed environment and an open convenience sampling survey. Statistics included paired student's t-tests corrected for multiple testing. For qualitative analysis, we employed a single-coder approach. Results: In 400 participants, we detected low priority of WHO-1 (before touching a patient) and WHO-2 (before clean/aseptic procedure) hand hygiene moments, despite knowing the risks of omission of hand disinfection. For all moments, self-assessment exceeded the assessment of colleagues (p < 0.001). For WHO-3, we detected a lower disinfection priority for wearing gloves compared to contaminated bare hands. Qualitative analyses revealed five themes: basic conditions, didactic implementations, cognitive load, and uncertainty about feasibility and efficacy. Discussion: Considering bias, the study's subjective nature, the unknown role of emergency-related infections contributing to hospital-acquired infections, and different experiences of healthcare providers, we conclude that hand disinfection before emergencies is de-prioritized and justified by the emergency situation regardless of the objective feasibility. Conclusion: This study reveals subjective and objective barriers to implementation of WHO-1 and WHO-2 moments of hand disinfection to be further evaluated and addressed in educational programs.

Evaluation of arginine metabolism for the analysis of M1/M2 macrophage activation in human clinical specimens.

BACKGROUND: Macrophage heterogeneity reflects their plasticity in response to environmental stimuli. Usually human macrophages are characterized by analysis of surface molecules or cytokine expression while functional assays are established in the mouse system but lacking for various human specimens. METHODS: To evaluate the value of analysis of arginine metabolism for characterization of human macrophage differentiation, we analyzed nitrite production and arginase activity in plasma, duodenal biopsies, and in vitro differentiated macrophages of patients with classical Whipple's disease. RESULTS: We demonstrate that it is feasible to determine the content of urea in supernatants of stimulated duodenal biopsies, arginase activity in fresh duodenal biopsies and plasma samples, and arginase activity and nitrite production in lysates and supernatants of in vitro differentiated macrophages. However, only selected tests are appropriate to define macrophage polarization in human specimens. CONCLUSION: Analysis of arginine metabolism is not suitable for the characterization of in vitro differentiated human macrophages. Besides the measurement of nitrite in duodenal biopsy supernatants, the determination of arginase activity in human plasma seems to be a reasonable functional test to detect enhanced M2 macrophage activation and, thus, is of great value for the analysis of macrophage activity with a minimum of material and costs.

Regulatory T cells in patients with Whipple's disease.

Classical Whipple's disease (CWD) is caused by chronic infection with Tropheryma whipplei that seems to be associated with an underlying immune defect. The pathognomonic hallmark of CWD is a massive infiltration of the duodenal mucosa with T. whipplei-infected macrophages that disperse systemically to many other organ systems. An alleviated inflammatory reaction and the absence of T. whipplei-specific Th1 reactivity support persistence and systemic spread of the pathogen. In this article, we hypothesized that regulatory T cells (T(reg)) are involved in immunomodulation in CWD, and we asked for the distribution, activation, and regulatory capacity of T(reg) in CWD patients. Whereas in the lamina propria of CWD patients before treatment numbers of T(reg) were increased, percentages in the peripheral blood were similar in CWD patients and healthy controls. However, peripheral T(reg) of CWD patients were more activated than those of controls. Elevated secretion of IL-10 and TGF-ß in the duodenal mucosa of CWD patients indicated locally enhanced T(reg) activity. Enhanced CD95 expression on peripheral memory CD4(+) T cells combined with reduced expression of IFN-γ and IL-17A upon polyclonal stimulation by CD4(+) cells from untreated CWD patients further hinted to T(reg) activity-related exhaustion of effector CD4(+) T cells. In conclusion, increased numbers of T(reg) can be detected within the duodenal mucosa in untreated CWD, where huge numbers of T. whipplei-infected macrophages are present. Thus, T(reg) might contribute to the chronic infection and systemic spread of T. whipplei in CWD but in contrast prevent mucosal barrier defect by reducing local inflammation.

The Role of NETosis and Complement Activation in COVID-19-Associated Coagulopathies.

Inflammation-induced coagulopathy is a common complication associated with coronavirus disease 2019 (COVID-19). We aim to evaluate the association of NETosis and complement markers with each other as well as their association with thrombogenicity and disease severity in COVID-19. The study included hospitalized patients with an acute respiratory infection: patients with SARS-CoV2 infection (COVpos, n = 47) or either pneumonia or infection-triggered acute exacerbated COPD (COVneg, n = 36). Our results show that NETosis, coagulation, and platelets, as well as complement markers, were significantly increased in COVpos patients, especially in severely ill COVpos patients. NETosis marker MPO/DNA complexes correlated with coagulation, platelet, and complement markers only in COVpos. Severely ill COVpos patients showed an association between complement C3 and SOFA (R = 0.48; p ≤ 0.028), C5 and SOFA (R = 0.46; p ≤ 0.038), and C5b-9 and SOFA (R = 0.44; p ≤ 0.046). This study provides further evidence that NETosis and the complement system are key players in COVID-19 inflammation and clinical severity. Unlike previous studies that found NETosis and complement markers to be elevated in COVID-19 patients compared to healthy controls, our findings show that this characteristic distinguishes COVID-19 from other pulmonary infectious diseases. Based on our results, we propose that COVID-19 patients at high risk for immunothrombosis could be identified via elevated complement markers such as C5.

Initial therapeutic anticoagulation with rivaroxaban compared to prophylactic therapy with heparins in moderate to severe COVID-19: results of the COVID-PREVENT randomized controlled trial.

BACKGROUND: COVID-19 is associated with a prothrombotic state. Current guidelines recommend prophylactic anticoagulation upon hospitalization. METHODS: COVID-PREVENT, an open-label, multicenter, randomized, clinical trial enrolled patients (≥ 18 years) with moderate to severe COVID-19 and age-adjusted D-dimers > 1.5 upper limit of normal (ULN). The participants were randomly assigned (1:1) to receive either therapeutic anticoagulation with rivaroxaban 20 mg once daily or thromboprophylaxis with a heparin (SOC) for at least 7 days followed by prophylactic anticoagulation with rivaroxaban 10 mg once daily for 28 days or no thromboprophylaxis. The primary efficacy outcome was the D-dimer level and the co-primary efficacy outcome the 7-category ordinal COVID-19 scale by WHO at 7 days post randomization. The secondary outcome was time to the composite event of either venous or arterial thromboembolism, new myocardial infarction, non-hemorrhagic stroke, all-cause death or progression to intubation and invasive ventilation up to 35 days post randomization. RESULTS: The primary efficacy outcome D-dimer at 7 days was not different between patients assigned to therapeutic (n = 55) or prophylactic anticoagulation (n = 56) (1.21 mg/L [0.79, 1.86] vs 1.27 mg/L [0.79, 2.04], p = 0.78). In the whole study population D-dimer was significantly lower at 7 days compared to baseline (1.05 mg/L [0.75, 1.48] vs 1.57 mg/L [1.13, 2.19], p < 0.0001). Therapy with rivaroxaban compared to SOC was not associated an improvement on the WHO 7-category ordinal scale at 7 days (p = 0.085). Rivaroxaban improved the clinical outcome measured by the score in patients with a higher baseline D-dimer > 2.0 ULN (exploratory analysis; 0.632 [0.516, 0.748], p = 0.026). The secondary endpoint occurred in 6 patients (10.9%) in the rivaroxaban group and in 12 (21.4%) in the SOC group (time-to-first occurrence of the components of the secondary outcome: HR 0.5; 95% CI 0.15-1.67; p = 0.264). There was no difference in fatal or non-fatal major or clinically relevant non-major bleeding between the groups. CONCLUSIONS: Therapeutic anticoagulation with rivaroxaban compared to prophylactic anticoagulation with a heparin did not improve surrogates of clinical outcome in patients with moderate to severe COVID-19. Whether initial rivaroxaban at therapeutic doses might be superior to thromboprophylaxis in patients with COVID-19 and a high risk as defined by D-dimer > 2 ULN needs confirmation in further studies.

Intestinal Barrier Dysfunction and Microbial Translocation in Patients with First-Diagnosed Atrial Fibrillation.

BACKGROUND: According to the leaky gut concept, microbial products (e.g., lipopolysaccharide, LPS) enter the circulation and mediate pro-inflammatory immunological responses. Higher plasma LPS levels have been reported in patients with various cardiovascular diseases, but not specifically during early atrial fibrillation (AF). METHODS: We studied data and blood samples from patients presenting with first-diagnosed AF (FDAF) (n = 80) and 20 controls. RESULTS: Circulating biomarkers that are suggestive of mucosal inflammation (zonulin, mucosal adhesion molecule MAdCAM-1) and intestinal epithelium damage (intestinal fatty acid binding protein, IFABP) were increased in the plasma of patients with FDAF when compared to patients with chronic cardiovascular diseases but without AF. Surrogate plasma markers of increased intestinal permeability (LPS, CD14, LPS-binding protein, gut-derived LPS-neutralising IgA antibodies, EndoCAbs) were detected during early AF. A reduced ratio of IgG/IgM EndoCAbs titres indicated chronic endotoxaemia. Collagen turnover biomarkers, which corresponded to the LPS values, suggested an association of gut-derived low-grade endotoxaemia with adverse structural remodelling. The LPS concentrations were higher in FDAF patients who experienced a major adverse cardiovascular event. CONCLUSIONS: Intestinal barrier dysfunction and microbial translocation accompany FDAF. Improving gut permeability and low-grade endotoxaemia might be a potential therapeutic approach to reducing the disease progression and cardiovascular complications in FDAF.

Data processing pipeline for cardiogenic shock prediction using machine learning.

Introduction: Recent advances in machine learning provide new possibilities to process and analyse observational patient data to predict patient outcomes. In this paper, we introduce a data processing pipeline for cardiogenic shock (CS) prediction from the MIMIC III database of intensive cardiac care unit patients with acute coronary syndrome. The ability to identify high-risk patients could possibly allow taking pre-emptive measures and thus prevent the development of CS. Methods: We mainly focus on techniques for the imputation of missing data by generating a pipeline for imputation and comparing the performance of various multivariate imputation algorithms, including k-nearest neighbours, two singular value decomposition (SVD)-based methods, and Multiple Imputation by Chained Equations. After imputation, we select the final subjects and variables from the imputed dataset and showcase the performance of the gradient-boosted framework that uses a tree-based classifier for cardiogenic shock prediction. Results: We achieved good classification performance thanks to data cleaning and imputation (cross-validated mean area under the curve 0.805) without hyperparameter optimization. Conclusion: We believe our pre-processing pipeline would prove helpful also for other classification and regression experiments.

Left-ventricular unloading in extracorporeal cardiopulmonary resuscitation due to acute myocardial infarction - A multicenter study.

BACKGROUND: Guidelines advocate the use of extracorporeal cardio-pulmonary resuscitation with veno-arterial extracorporeal membrane oxygenation (VA-ECMO) in selected patients with cardiac arrest. Effects of concomitant left-ventricular (LV) unloading with Impella® (ECMELLA) remain unclear. This is the first study to investigate whether treatment with ECMELLA was associated with improved outcomes in patients with refractory cardiac arrest caused by acute myocardial infarction (AMI). METHODS: This study was approved by the local ethical committee. Patients treated with ECMELLA at three centers between 2016 and 2021 were propensity score (PS)-matched to patients receiving VA-ECMO based on age, electrocardiogram rhythm, cardiac arrest location and Survival After Veno-Arterial ECMO (SAVE) score. Cox proportional-hazard and Poisson regression models were used to analyse 30-day mortality rate (primary outcome), hospital and intensive care unit (ICU) length of stay (LOS) (secondary outcomes). Sensitivity analyses on patient demographics and cardiac arrest parameters were performed. RESULTS: 95 adult patients were included in this study, out of whom 34 pairs of patients were PS-matched. ECMELLA treatment was associated with decreased 30-day mortality risk (Hazard Ratio [HR] 0.53 [95% Confidence Interval (CI) 0.31-0.91], P = 0.021), prolonged hospital (Incidence Rate Ratio (IRR) 1.71 [95% CI 1.50-1.95], P < 0.001) and ICU LOS (IRR 1.81 [95% CI 1.57-2.08], P < 0.001). LV ejection fraction significantly improved until ICU discharge in the ECMELLA group. Especially patients with prolonged low-flow time and high initial lactate benefited from additional LV unloading. CONCLUSIONS: LV unloading with Impella® concomitant to VA-ECMO therapy in patients with therapy-refractory cardiac arrest due to AMI was associated with improved patient outcomes.

Intestinal barrier dysfunction mediates Whipple's disease immune reconstitution inflammatory syndrome (IRIS).

BACKGROUND & AIMS: Classical Whipple's disease (CWD) affects the gastrointestinal tract and causes chronic diarrhea, malabsorption, and barrier dysfunction with microbial translocation (MT). Immune reconstitution inflammatory syndrome (IRIS) is a serious complication during antimicrobial treatment of CWD. The pathomechanisms of IRIS have not been identified and mucosal barrier integrity has not been studied in patients with IRIS CWD. METHODS: In 96 CWD patients (n = 23 IRIS, n = 73 non-IRIS) and 30 control subjects, we analysed duodenal morphology by histology, measured serum markers of MT, and proinflammatory cytokines in biopsy supernatants, and correlated microbial translocation with T cell reconstitution and activation. RESULTS: Before treatment, duodenal specimens from patients who later developed IRIS exhibited a more pronounced morphological transformation that suggested a disturbed barrier integrity when compared with the non-IRIS group. Villous atrophy was mediated by increased apoptosis of epithelial cells, which was insufficiently counterbalanced by regenerative proliferation of crypt cells. Pretreatment deficiencies in the mucosal secretion of proinflammatory cytokines and chemokines (e.g., IL-6, CCL2) in these patients markedly resolved after therapy induction. High serum levels of lipopolysaccharides (LPS), soluble CD14 (sCD14), and LPS-binding protein (LBP) combined with low endotoxin core antibody (EndoCAb) titres suggested systemic MT in CWD patients developing IRIS. CD4+ T cell count and activation in IRIS CWD patients correlated positively with sCD14 levels and negatively with EndoCAb titres. Furthermore, the degree of intestinal barrier dysfunction and MT was predictive for the onset of IRIS. CONCLUSION: Prolonged MT across a dysfunctional intestinal mucosal barrier due to severe tissue damage favors dysbalanced immune reconstitution and systemic immune activation in IRIS CWD. Therefore, the monitoring of inflammatory and MT markers in CWD patients might be helpful in identifying patients who are at risk of developing IRIS. Therapeutic strategies to reconstitute the mucosal barrier and control inflammation could assist in the prevention of IRIS.

Disease Severity in Moderate-to-Severe COVID-19 Is Associated With Platelet Hyperreactivity and Innate Immune Activation.

Background: Hemostasis and inflammation are both dysregulated in patients with moderate-to-severe coronavirus disease 2019 (COVID-19). Yet, both processes can also be disturbed in patients with other respiratory diseases, and the interactions between coagulation, inflammation, and disease severity specific to COVID-19 are still vague. Methods: Hospitalized patients with acute respiratory symptoms and with severe acute respiratory syndrome coronavirus 2 (SARS-CoV2)-positive (COVpos) and SARS-CoV2-negative (COVneg) status were included. We assessed adenosine diphosphate (ADP)-, thrombin receptor activator peptide 6 (TRAP)-, and arachidonic acid (AA)-induced platelet reactivity by impedance aggregometry, as well as leukocyte subtype spectrum and platelet-leukocyte aggregates by flow cytometry and inflammatory cytokines by cytometric bead array. Results: ADP-, TRAP-, and AA-induced platelet reactivity was significantly higher in COVpos than in COVneg patients. Disease severity, assessed by sequential organ failure assessment (SOFA) score, was higher in COVpos than in COVneg patients and again higher in deceased COVpos patients than in surviving COVpos. The SOFA score correlated significantly with the mean platelet volume and TRAP-induced platelet aggregability. A larger percentage of classical and intermediate monocytes, and of CD4pos T cells (TH) aggregated with platelets in COVpos than in COVneg patients. Interleukin (IL)-1 receptor antagonist (RA) and IL-6 levels were higher in COVpos than in COVneg patients and again higher in deceased COVpos patients than in surviving COVpos. IL-1RA and IL-6 levels correlated with the SOFA score in COVpos but not in COVneg patients. In both respiratory disease groups, absolute levels of B-cell-platelet aggregates and NK-cell-platelet aggregates were correlated with ex vivo platelet aggegation upon stimulation with AA and ADP, respectively, indicating a universal, but not a COVID-19-specific mechanism. Conclusion: In moderate-to-severe COVID-19, but not in other respiratory diseases, disease severity was associated with platelet hyperreactivity and a typical inflammatory signature. In addition to a severe inflammatory response, platelet hyperreactivity associated to a worse clinical outcome in patients with COVID-19, pointing to the importance of antithrombotic therapy for reducing disease severity.