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Intestinal inflammation can impair mucosal healing, thereby establishing a vicious cycle leading to chronic inflammatory bowel disease (IBD). However, the signaling networks driving chronic inflammation remain unclear. Here we report that CD4 T cells isolated from patients with IBD produce high levels of interleukin-22 binding protein (IL-22BP), the endogenous inhibitor of the tissue-protective cytokine IL-22. Using mouse models, we demonstrate that IBD development requires T cell-derived IL-22BP. Lastly, intestinal CD4 T cells isolated from IBD patients responsive to treatment with antibodies against tumor necrosis factor-α (anti-TNF-α), the most effective known IBD therapy, exhibited reduced amounts of IL-22BP expression but still expressed IL-22. Our findings suggest that anti-TNF-α therapy may act at least in part by suppressing IL-22BP and point toward a more specific potential therapy for IBD.

The pathogenesis of severe COVID-19 reflects an inefficient immune reaction to SARS-CoV-2. Here we analyze, at the single cell level, plasmablasts egressed into the blood to study the dynamics of adaptive immune response in COVID-19 patients requiring intensive care. Before seroconversion in response to SARS-CoV-2 spike protein, peripheral plasmablasts display a type 1 interferon-induced gene expression signature; however, following seroconversion, plasmablasts lose this signature, express instead gene signatures induced by IL-21 and TGF-β, and produce mostly IgG1 and IgA1. In the sustained immune reaction from COVID-19 patients, plasmablasts shift to the expression of IgA2, thereby reflecting an instruction by TGF-β. Despite their continued presence in the blood, plasmablasts are not found in the lungs of deceased COVID-19 patients, nor does patient IgA2 binds to the dominant antigens of SARS-CoV-2. Our results thus suggest that, in severe COVID-19, SARS-CoV-2 triggers a chronic immune reaction that is instructed by TGF-β, and is distracted from itself.

Objective-Diabetes mellitus involves vascular inflammatory processes and is a main contributor to cardiovascular mortality.Notably, heightened levels of circulating tissue factor (TF) account for the increased thrombogenicity and put those patients at risk for thromboembolic events. Here, we sought to investigate the role of micro-RNA (miR)-driven TF expression and thrombogenicity in diabetes mellitus. Approach and Results-Plasma samples of patients with diabetes mellitus were analyzed for TF protein and activity as well as miR-126 expression before and after optimization of the antidiabetic treatment. We found low miR-126 levels to be associated with markedly increased TF protein and TF-mediated thrombogenicity. Reduced miR-126 expression was accompanied by increased vascular inflammation as evident from the levels of vascular adhesion molecule-1 and fibrinogen, as well as leukocyte counts. With optimization of the antidiabetic treatment miR-126 levels increased and thrombogenicity was reduced. Using a luciferase reporter system, we demonstrated miR-126 to directly bind to the F3-3′-untranslated region, thereby reducing TF expression both on mRNA and on protein levels in human microvascular endothelial cells as well as TF mRNA and activity in monocytes. Conclusions-Circulating

Aims Gut microbiota and their generated metabolites impact the host vascular phenotype. The metaorganismal metabolite trimethylamine N-oxide (TMAO) is both associated with adverse clinical thromboembolic events, and enhances platelet responsiveness in subjects. The impact of TMAO on vascular tissue factor (TF) in vivo is unknown. Here, we explore whether TMAO-enhanced thrombosis potential extends beyond TMAO effects on platelets, and is linked to TF. We also further explore the links between gut microbiota and vascular endothelial TF expression in vivo. Methods and Results In initial exploratory clinical studies, we observed that among sequential stable subjects (n = 2,989) on anti-platelet therapy undergoing elective diagnostic cardiovascular evaluation at a single-site referral center, TMAO levels were associated with an increased incident (3 yr) risk for major adverse cardiovascular events (MACE, myocardial infarction, stroke or death) [4th quartile(Q4) versus Q1 adjusted hazard ratio(95% confidence interval) HR(95%CI), 1.73(1.25-2.38)]. Similar results were observed within subjects on aspirin mono-therapy during follow-up [adjusted HR(95%CI) 1.75(1.25-2.44), n = 2,793). Leveraging access to a second higher risk cohort with previously reported TMAO data and monitoring of anti-platelet medication use, we also observed a strong association between TMAO and incident (1 yr) MACE risk in the multi-site Swiss Acute Coronary Syndromes (ACS) Cohort, focusing on the subset (n = 1,469) on chronic dual anti-platelet therapy during follow-up [adjusted HR(95% CI) 1.70(1.08-2.69)]. These collective clinical data suggest that the thrombosis-associated effects of TMAO may be mediated by cells/factors that are not inhibited by anti-platelet therapy. To test this, we first observed in human microvascular endothelial cells that TMAO dose-dependently induced expression of TF and vascular cell adhesion molecule (VCAM)1. In mouse studies, we observed that TMAO enhanced aortic TF and VCAM1 mRNA and protein expression, which upon immunolocalization studies, was shown to co-localize with vascular endothelial cells. Finally, in arterial injury mouse models, TMAO-dependent enhancement of in vivo TF expression and thrombogenicity were abrogated by either a TF-inhibitory antibody or a mechanism-based microbial choline TMA lyase inhibitor (fluoromethylcholine, FMC). Conclusions Endothelial TF contributes to TMAO-related arterial thrombosis potential, and can be specifically blocked by targeted non-lethal inhibition of gut microbial choline TMA lyase. Translational Perspective The pro-thrombotic effects of the gut microbial TMAO pathway are shown to extend beyond enhancement of platelet responsiveness and include heightened vascular Tissue Factor(TF). In clinical studies, TMAO is shown to predict event risk in patients in the presence of anti-platelet drugs. In animal studies, TMAO elevation is shown to promote vascular endothelial TF expression and a TF-dependent pro-thrombotic effect. Pharmacological targeting of gut microbial choline TMA lyase reduced host TMAO, vascular TF and abrogated the pro-thrombotic TMAO-associated phenotype. These studies suggest inhibiting the TMAO pathway may be a rational target for reducing residual risk in patients on antiplatelet therapy.

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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 marrowderived 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.