RIP3, a kinase promoting necroptotic cell death, mediates adverse remodelling after myocardial infarction

Aims Programmed necrosis (necroptosis) represents a newly identified mechanism of cell death combining features of both apoptosis and necrosis. Like apoptosis, necroptosis is tightly regulated by distinct signalling pathways. A key regulatory role in programmed necrosis has been attributed to interactions of the receptor-interacting protein kinases, RIP1 and RIP3. However, the specific functional role of RIP3-dependent signalling and necroptosis in the heart is unknown. The aims of this study were thus to assess the significance of necroptosis and RIP3 in the context of myocardial ischaemia. Methods and results Immunoblots revealed strong expression of RIP3 in murine hearts, indicating potential functional significance of this protein in the myocardium. Consistent with a role in promoting necroptosis, adenoviral overexpression of RIP3 in neonatal rat cardiomyocytes and stimulation with TNF-α induced the formation of a complex of RIP1 and RIP3. Moreover, RIP3 overexpression was sufficient to induce necroptosis of cardiomyocytes. In vivo, cardiac expression of RIP3 was up-regulated upon myocardial infarction (MI). Conversely, mice deficient for RIP3 (RIP3−/−) showed a significantly better ejection fraction (45 ± 3.6 vs. 32 ± 4.4%, P < 0.05) and less hypertrophy in magnetic resonance imaging studies 30 days after experimental infarction due to left anterior descending coronary artery ligation. This was accompanied by a diminished inflammatory response of infarcted hearts and decreased generation of reactive oxygen species. Conclusion Here, we show that RIP3-dependent necroptosis modulates post-ischaemic adverse remodelling in a mouse model of MI. This novel signalling pathway may thus be an attractive target for future therapies that aim to limit the adverse consequences of myocardial ischaemi

Intestinal B cells license metabolic T-cell activation in NASH microbiota/antigen-independently and contribute to fibrosis by IgA-FcR signalling

BACKGROUND & AIMS: The progression of non-alcoholic steatohepatitis (NASH) to fibrosis and hepatocellular carcinoma (HCC) is aggravated by auto-aggressive T cells. The gut-liver axis contributes to NASH, but the mechanisms involved and the consequences for NASH-induced fibrosis and liver cancer remain unknown. We investigated the role of gastrointestinal B cells in the development of NASH, fibrosis and NASH-induced HCC. METHODS: C57BL/6J wild-type (WT), B cell-deficient and different immunoglobulin-deficient or transgenic mice were fed distinct NASH-inducing diets or standard chow for 6 or 12 months, whereafter NASH, fibrosis, and NASH-induced HCC were assessed and analysed. Specific pathogen-free/germ-free WT and μMT mice (containing B cells only in the gastrointestinal tract) were fed a choline-deficient high-fat diet, and treated with an anti-CD20 antibody, whereafter NASH and fibrosis were assessed. Tissue biopsy samples from patients with simple steatosis, NASH and cirrhosis were analysed to correlate the secretion of immunoglobulins to clinicopathological features. Flow cytometry, immunohistochemistry and single-cell RNA-sequencing analysis were performed in liver and gastrointestinal tissue to characterise immune cells in mice and humans. RESULTS: Activated intestinal B cells were increased in mouse and human NASH samples and licensed metabolic T-cell activation to induce NASH independently of antigen specificity and gut microbiota. Genetic or therapeutic depletion of systemic or gastrointestinal B cells prevented or reverted NASH and liver fibrosis. IgA secretion was necessary for fibrosis induction by activating CD11b+CCR2+F4/80+CD11c-FCGR1+ hepatic myeloid cells through an IgA-FcR signalling axis. Similarly, patients with NASH had increased numbers of activated intestinal B cells; additionally, we observed a positive correlation between IgA levels and activated FcRg+ hepatic myeloid cells, as well the extent of liver fibrosis. CONCLUSIONS: Intestinal B cells and the IgA-FcR signalling axis represent potential therapeutic targets for the treatment of NASH. IMPACT AND IMPLICATIONS: There is currently no effective treatment for non-alcoholic steatohepatitis (NASH), which is associated with a substantial healthcare burden and is a growing risk factor for hepatocellular carcinoma (HCC). We have previously shown that NASH is an auto-aggressive condition aggravated, amongst others, by T cells. Therefore, we hypothesized that B cells might have a role in disease induction and progression. Our present work highlights that B cells have a dual role in NASH pathogenesis, being implicated in the activation of auto-aggressive T cells and the development of fibrosis via activation of monocyte-derived macrophages by secreted immunoglobulins (e.g., IgA). Furthermore, we show that the absence of B cells prevented HCC development. B cell-intrinsic signalling pathways, secreted immunoglobulins, and interactions of B cells with other immune cells are potential targets for combinatorial NASH therapies against inflammation and fibrosis

The European Forest Plant Species List (EuForPlant): Concept and applications

Question When evaluating forests in terms of their biodiversity, distinctiveness and naturalness, the affinity of the constituent species to forests is a crucial parameter. Here we ask to what extent are vascular plant species associated with forests, and does species' affinity to forests vary between European regions? Location Temperate and boreal forest biome of Northwestern and Central Europe. Methods We compiled EuForPlant, a new extensive list of forest vascular plant species in 24 regions spread across 13 European countries using vegetation databases and expert knowledge. Species were region-specifically classified into four categories reflecting the degree of their affinity to forest habitats: 1.1, species of forest interiors; 1.2, species of forest edges and forest openings; 2.1, species that can be found in forest as well as open vegetation; and 2.2, species that can be found partly in forest, but mainly in open vegetation. An additional "O" category was distinguished, covering species typical for non-forest vegetation. Results EuForPlant comprises 1,726 species, including 1,437 herb-layer species, 159 shrubs, 107 trees, 19 lianas and 4 epiphytic parasites. Across regions, generalist forest species (with 450 and 777 species classified as 2.1 and 2.2, respectively) significantly outnumbered specialist forest species (with 250 and 137 species classified as 1.1 and 1.2, respectively). Even though the degree of shifting between the categories of forest affinity among regions was relatively low (on average, 17.5%), about one-third of the forest species (especially 1.2 and 2.2) swapped categories in at least one of the study regions. Conclusions The proposed list can be used widely in vegetation science and global change ecology related to forest biodiversity and community dynamics. Shifting of forest affinity among regions emphasizes the importance of a continental-scale forest plant species list with regional specificity

HNF4alpha Dysfunction as a Molecular Rational for Cyclosporine Induced Hypertension

Induction of tolerance against grafted organs is achieved by the immunosuppressive agent cyclosporine, a prominent member of the calcineurin inhibitors. Unfortunately, its lifetime use is associated with hypertension and nephrotoxicity. Several mechanism for cyclosporine induced hypertension have been proposed, i.e. activation of the sympathetic nervous system, endothelin-mediated systemic vasoconstriction, impaired vasodilatation secondary to reduction in prostaglandin and nitric oxide, altered cytosolic calcium translocation, and activation of the renin-angiotensin system (RAS). In this regard the molecular basis for undue RAS activation and an increased signaling of the vasoactive oligopeptide angiotensin II (AngII) remain elusive. Notably, angiotensinogen (AGT) is the precursor of AngII and transcriptional regulation of AGT is controlled by the hepatic nuclear factor HNF4alpha. To better understand the molecular events associated with cyclosporine induced hypertension, we investigated the effect of cyclosporine on HNF4alpha expression and activity and searched for novel HNF4alpha target genes among members of the RAS cascade. Using bioinformatic algorithm and EMSA bandshift assays we identified angiotensin II receptor type 1 (AGTR1), angiotensin I converting enzyme (ACE), and angiotensin I converting enzyme 2 (ACE2) as genes targeted by HNF4alpha. Notably, cyclosporine represses HNF4alpha gene and protein expression and its DNA-binding activity at consensus sequences to AGT, AGTR1, ACE, and ACE2. Consequently, the gene expression of AGT, AGTR1, and ACE2 was significantly reduced as evidenced by quantitative real-time RT-PCR. While RAS is composed of a sophisticated interplay between multiple factors we propose a decrease of ACE2 to enforce AngII signaling via AGTR1 to ultimately result in vasoconstriction and hypertension. Taken collectively we demonstrate cyclosporine to repress HNF4alpha activity through calcineurin inhibitor mediated inhibition of nuclear factor of activation of T-cells (NFAT) which in turn represses HNF4alpha that leads to a disturbed balance of RAS

Intestinal B-cells license metabolic T-cell activation in NASH microbiota/antigen-independently and contribute to fibrosis by IgA-FcR signalling

BACKGROUND & AIMS The progression of nonalcoholic steatohepatitis (NASH) to fibrosis and hepatocellular carcinoma (HCC) is aggravated by auto-aggressive T cells. The gut-liver axis contributes to NASH, but the mechanisms involved and the consequences for NASH-induced fibrosis and liver cancer remain unknown. We investigated the role of gastrointestinal B cells in the development of NASH, fibrosis and NASH-induced HCC. METHODS C57BL/6J wild-type (WT), B cell-deficient and different immunoglobulin-deficient or transgenic mice were fed distinct NASH diets (for example, choline-deficient high-fat diet, CD-HFD) or chow diet for 6 or 12 months, whereafter NASH, fibrosis, and NASH-induced HCC were assessed and analysed. Specific pathogen-free/germ-free WT and μMT mice (containing B cells only in the gastrointestinal tract) were fed a CD-HFD, and treated with an anti-CD20 antibody, whereafter NASH and fibrosis were assessed. Tissue biopsy samples from patients with NAFL, NASH and cirrhosis were analysed to correlate the secretion of immunoglobulins to clinicopathological features. Flow cytometry, immunohistochemistry and scRNA-Seq analysis were performed in liver and gastrointestinal tissue for immune cells in mice and humans. RESULTS Activated intestinal B cells were increased in mouse and human NASH samples and licensed metabolic T-cell activation to induce NASH independently of antigen-specificity and gut microbiota. Genetic or therapeutic depletion of systemic or gastrointestinal B cells prevented or reverted NASH and liver fibrosis. IgA secretion was necessary for fibrosis induction by activating CD11b+CCR2+F4/80+CD11c-FCGR1+ hepatic myeloid cells through an IgA-FcR signalling axis. Similarly, patients with NASH had increased numbers of activated intestinal B-cells and showed a positive correlation between IgA levels and activated FcRγ+ hepatic myeloid cells as well extent of liver fibrosis. CONCLUSIONS Intestinal B cells and the IgA-FcR signalling axis represent potential therapeutic targets for treating NASH. IMPACT AND IMPLICATIONS Nonalcoholic steatohepatitis (NASH) is a chronic inflammatory condition on the rise and can lead to hepatocellular carcinoma (HCC), the 3rd most common cause of cancer-related death worldwide. Currently, there is no effective treatment for this progressive disease that correlates with a marked risk of HCC mortality and carries a substantial healthcare burden. To date, among all the solid tumours, especially in HCC, the incidence and mortality rates are almost the same, making it crucial to find curative treatments for chronic diseases, such as NASH, which highly predispose to tumorigenesis. We have previously shown that NASH is an auto-aggressive condition aggravated, amongst others, by T cells. Therefore, we hypothesized that B cells might have a role in disease induction and progression. Our present work highlights that B cells have a dual role in NASH pathogenesis, being implicated in the activation of auto-aggressive T cells and the development of fibrosis via activation of monocyte-derived macrophages by secreted immunoglobulins (e.g., IgA). Furthermore, we could show that the absence of B cells prevented HCC development. B-cell intrinsic signalling pathways, secreted immunoglobulins, and interactions of B cells with other immune cells are potential targets in combinatorial NASH therapies against inflammation and fibrosis

Interprofessional and interdisciplinary simulation-based training leads to safe sedation procedures in the emergency department

BACKGROUND Sedation is a procedure required for many interventions in the Emergency department (ED) such as reductions, surgical procedures or cardioversions. However, especially under emergency conditions with high risk patients and rapidly changing interdisciplinary and interprofessional teams, the procedure caries important risks. It is thus vital but difficult to implement a standard operating procedure for sedation procedures in any ED. Reports on both, implementation strategies as well as their success are currently lacking. This study describes the development, implementation and clinical evaluation of an interprofessional and interdisciplinary simulation-based sedation training concept. METHODS All physicians and nurses with specialised training in emergency medicine at the Berne University Department of Emergency Medicine participated in a mandatory interdisciplinary and interprofessional simulation-based sedation training. The curriculum consisted of an individual self-learning module, an airway skill training course, three simulation-based team training cases, and a final practical learning course in the operating theatre. Before and after each training session, self-efficacy, awareness of emergency procedures, knowledge of sedation medication and crisis resource management were assessed with a questionnaire. Changes in these measures were compared via paired tests, separately for groups formed based on experience and profession. To assess the clinical effect of training, we collected patient and team satisfaction as well as duration and complications for all sedations in the ED within the year after implementation. We further compared time to beginning of procedure, time for duration of procedure and time until discharge after implementation with the one year period before the implementation. Cohen's d was calculated as effect size for all statistically significant tests. RESULTS Fifty staff members (26 nurses and 24 physicians) participated in the training. In all subgroups, there is a significant increase in self-efficacy and knowledge with high effect size (d z  = 1.8). The learning is independent of profession and experience level. In the clinical evaluation after implementation, we found no major complications among the sedations performed. Time to procedure significantly improved after the introduction of the training (d = 0.88). DISCUSSION Learning is independent of previous working experience and equally effective in raising the self-efficacy and knowledge in all professional groups. Clinical outcome evaluation confirms the concepts safety and feasibility. CONCLUSION An interprofessional and interdisciplinary simulation-based sedation training is an efficient way to implement a conscious sedation concept in an ED

Deep Drilling in the Time Domain with DECam: Survey Characterization

This paper presents a new optical imaging survey of four deep drilling fields (DDFs), two Galactic and two extragalactic, with the Dark Energy Camera (DECam) on the 4 meter Blanco telescope at the Cerro Tololo Inter-American Observatory (CTIO). During the first year of observations in 2021, $>$4000 images covering 21 square degrees (7 DECam pointings), with $\sim$40 epochs (nights) per field and 5 to 6 images per night per filter in $g$, $r$, $i$, and/or $z$, have become publicly available (the proprietary period for this program is waived). We describe the real-time difference-image pipeline and how alerts are distributed to brokers via the same distribution system as the Zwicky Transient Facility (ZTF). In this paper, we focus on the two extragalactic deep fields (COSMOS and ELAIS-S1), characterizing the detected sources and demonstrating that the survey design is effective for probing the discovery space of faint and fast variable and transient sources. We describe and make publicly available 4413 calibrated light curves based on difference-image detection photometry of transients and variables in the extragalactic fields. We also present preliminary scientific analysis regarding Solar System small bodies, stellar flares and variables, Galactic anomaly detection, fast-rising transients and variables, supernovae, and active galactic nuclei.Comment: 22 pages, 17 figures, 2 tables. Accepted to MNRA

Differential Impact of Tetratricopeptide Repeat Proteins on the Steroid Hormone Receptors

Tetratricopeptide repeat (TPR) motif containing co-chaperones of the chaperone Hsp90 are considered control modules that govern activity and specificity of this central folding platform. Steroid receptors are paradigm clients of Hsp90. The influence of some TPR proteins on selected receptors has been described, but a comprehensive analysis of the effects of TPR proteins on all steroid receptors has not been accomplished yet.We compared the influence of the TPR proteins FK506 binding proteins 51 and 52, protein phosphatase-5, C-terminus of Hsp70 interacting protein, cyclophillin 40, hepatitis-virus-B X-associated protein-2, and tetratricopeptide repeat protein-2 on all six steroid hormone receptors in a homogeneous mammalian cell system. To be able to assess each cofactor's effect on the transcriptional activity of on each steroid receptor we employed transient transfection in a reporter gene assay. In addition, we evaluated the interactions of the TPR proteins with the receptors and components of the Hsp90 chaperone heterocomplex by coimmunoprecipitation. In the functional assays, corticosteroid and progesterone receptors displayed the most sensitive and distinct reaction to the TPR proteins. Androgen receptor's activity was moderately impaired by most cofactors, whereas the Estrogen receptors' activity was impaired by most cofactors only to a minor degree. Second, interaction studies revealed that the strongly receptor-interacting co-chaperones were all among the inhibitory proteins. Intriguingly, the TPR-proteins also differentially co-precipitated the heterochaperone complex components Hsp90, Hsp70, and p23, pointing to differences in their modes of action.The results of this comprehensive study provide important insight into chaperoning of diverse client proteins via the combinatorial action of (co)-chaperones. The differential effects of the TPR proteins on steroid receptors bear on all physiological processes related to steroid hormone activity