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Background and Aims Glycoprotein VI (GPVI) is a platelet collagen/fibrin(ogen) receptor and an emerging pharmacological target for the treatment of thrombotic and thrombo-inflammatory diseases, notably ischaemic stroke. A first anti-human GPVI (hGPVI) antibody Fab-fragment (ACT017/glenzocimab, K D: 4.1 nM) recently passed a clinical phase 1b/2a study in patients with acute ischaemic stroke and was found to be well tolerated, safe, and potentially beneficial. In this study, a novel humanized anti-GPVI antibody Fab-fragment (EMA601; K D: 0.195 nM) was developed that inhibits hGPVI function with very high potency in vitro and in vivo. Methods Fab-fragments of the mouse anti-hGPVI IgG Emf6.1 were tested for functional GPVI inhibition in human platelets and in hGPVI expressing (hGP6tg/tg) mouse platelets. The in vivo effect of Emf6.1Fab was assessed in a tail bleeding assay, an arterial thrombosis model and the transient middle cerebral artery occlusion (tMCAO) model of ischaemic stroke. Using complementary-determining region grafting, a humanized version of Emf6.1Fab (EMA601) was generated. Emf6.1Fab/EMA601 interaction with hGPVI was mapped in array format and kinetics and quantified by bio-layer interferometry. Results Emf6.1Fab (K D: 0.427 nM) blocked GPVI function in human and hGP6tg/tg mouse platelets in multiple assays in vitro at concentrations ≥5 µg/mL. Emf6.1Fab (4 mg/kg)-treated hGP6tg/tg mice showed potent hGPVI inhibition ex vivo and were profoundly protected from arterial thrombosis as well as from cerebral infarct growth after tMCAO, whereas tail-bleeding times remained unaffected. Emf6.1Fab binds to a so far undescribed membrane proximal epitope in GPVI. The humanized variant EMA601 displayed further increased affinity for hGPVI (K D: 0.195 nM) and fully inhibited the receptor at 0.5 µg/mL, corresponding to a >50-fold potency compared with ACT017. Conclusions EMA601 is a conceptually novel and promising anti-platelet agent to efficiently prevent or treat arterial thrombosis and thrombo-inflammatory pathologies in humans at risk. [ABSTRACT FROM AUTHOR]

Introduction: People with haemophilia (PwH) suffer from haemophilic arthropathy which is accompanied by acute and chronic inflammation. The aim of this study was to examine the neuroinflammatory network operative in PwH and to compare it to healthy controls. Material and Methods: Blood samples were collected from 41 PwH (age 54.7 ± 11.7 years) and 33 healthy controls (age 50.9 ± 10.5 years) and the levels of 13 neuroinflammatory markers were analyzed by applying an antibody‐based detection kit in a flow cytometer. Results: From 13 analyzed markers, three—ß‐nerve growth factor (ß‐NGF), soluble receptor for advanced glycation endproducts (sRAGE) and Interleukin‐18 (IL‐18) differed significantly between the groups (ß‐NGF p =.045; sRAGE p =.003; IL‐18 p =.007). While ß‐NGF was downregulated in PwH, sRAGE and IL‐18 were upregulated. None of the analyzed markers corelated to the joint status of PwH while CCL2 (C‐C motif ligand 2 chemokine) correlated to HIV infections in PwH (r =.313, p =.007). Correlation analyses of the markers studied also revealed many differences between PwH and controls suggesting a number of deregulations in PwH. Conclusion: The altered levels of sRAGE and ß‐NGF in PwH, which have not been analyzed in PwH before, may help to understand the neuroinflammatory network operative in PwH. The general inflammatory processes in PwH and the involved biomarkers in PwH remain poorly understood. PwH could benefit from new therapies against neuroinflammation which may help to reduce inflammation or also chronic pain. [ABSTRACT FROM AUTHOR]

Bronchopulmonary dysplasia (BPD) is a chronic lung disease commonly seen in preterm infants, and is triggered by infection, mechanical ventilation, and oxygen toxicity. Among other problems, lifelong limitations in lung function and impaired psychomotor development may result. Despite major advances in understanding the disease pathologies, successful interventions are still limited to only a few drug therapies with a restricted therapeutic benefit, and which sometimes have significant side effects. As a more promising therapeutic option, mesenchymal stem cells (MSCs) have been in focus for several years due to their anti-inflammatory effects and their secretion of growth and development promoting factors. Preclinical studies provide evidence in that MSCs have the potential to contribute to the repair of lung injuries. This review provides an overview of MSCs, and other stem/progenitor cells present in the lung, their identifying characteristics, and their differentiation potential, including cytokine/growth factor involvement. Furthermore, animal studies and clinical trials using stem cells or their secretome are reviewed. To bring MSC-based therapeutic options further to clinical use, standardized protocols are needed, and upcoming side effects must be critically evaluated. To fill these gaps of knowledge, the MSCs' behavior and the effects of their secretome have to be examined in more (pre-) clinical studies, from which only few have been designed to date. [ABSTRACT FROM AUTHOR]

iNKT (invariant natural killer T) cells are unconventional immunoregulatory T cells which contribute to B cell maturation, antibody and cytokine production. iNKT cells are implicated in the control of autoimmune inflammation in different disorders. For bullous pemphigoid (BP), the most frequent bullous autoimmune dermatosis, the role of iNKT cells has not yet been studied. We, therefore, aimed at investigating the frequency of iNKT cells in peripheral blood and biopsies from lesional and non-lesional skin from patients with BP and controls. Circulating CD3+iTCR+ iNKT cells were assessed by flow cytometry in peripheral blood from 30 patients with BP and from 29 controls (19 patients with skin tumors and 10 healthy controls). In 34 lesional and 13 non-lesional skin biopsies from BP patients and 17 biopsies from control individuals the number of Vα24+Vβ11+ iNKT cells was investigated by immunofluorescence staining. BP patients showed a significantly lower frequency of circulating iNKT cells compared to the control group. Patients with severe disseminated blistering tended to display lower iNKT cell numbers than patients with moderate disease severity. In lesional skin of BP patients, an enrichment of iNKT cells was detected compared to skin biopsies from controls. Similarly to control biopsies, non-lesional biopsies of BP patients contained only few iNKT cells. In conclusion, the deficiency of circulating iNKT cells associated with enrichment at the site of cutaneous inflammation suggests that iNKT cells may play a pathophysiologically relevant role in BP. [ABSTRACT FROM AUTHOR]

Background: Is the patient really suffering from acute appendicitis? Right lower quadrant pain is the most common sign of acute appendicitis. However, many other bowels pathologies might mimic acute appendicitis. Due to fear of the consequences of delayed or missed diagnosis, the indication for emergency appendectomy is liberally made. This has been shown to be associated with high rates of negative appendectomy with risk of potentially serious or lethal complications. Thus there is need for a better preoperative screening of patients with suspected appendicitis. Methods: This prospective single center single-blinded pilot study was conducted in the Department of surgery at the HELIOS Universitätsklinikum Wuppertal, Germany. Calprotectin was measured in pre-therapeutic stool samples of patients presenting in the emergency department with pain to the right lower quadrant. Fecal calprotectin (FC) values were analyzed using commercially available ELISA kits. Cut-off values for FC were studied using the receiver-operator characteristic (ROC) curve. The Area under the curve (AUC) was reported for each ROC curve. Results: The mean FC value was 51.4 ± 118.8 μg/g in patients with AA, 320.9 ± 416.6 μg/g in patients with infectious enteritis and 24.8 ± 27.4 μg/g in the control group. ROC curve showed a close to 80% specificity and sensitivity of FC for AA at a cut-off value of 51 μg/g, AUC = 0.7. The sensitivity of FC at this cut-off value is zero for enteritis with a specificity of 35%. Conclusion: Fecal calprotectin could be helpful in screening patients with pain to the right lower quadrant for the presence of acute appendicitis or infectious enteritis with the aim of facilitating clinical decision-making and reducing the rate of negative appendectomy. [ABSTRACT FROM AUTHOR]

Background: Epigenetic signatures are highly cell type specific. Separation of distinct cell populations is therefore desirable for all epigenetic studies. However, to date little information is available on whether separation protocols might influence epigenetic and/or gene expression signatures and hence might be less beneficial. We investigated the influence of two frequently used protocols to isolate intestinal epithelium cells (IECs) from 6 healthy individuals. Materials and Methods: Epithelial cells were isolated from small bowel (i.e. terminal ileum) biopsies using EDTA/DTT and enzymatic release followed by magnetic bead sorting via EPCAM labeled microbeads. Effects on gene/mRNA expression were analyzed using a real time PCR based expression array. DNA methylation was assessed by pyrosequencing of bisulfite converted DNA and methylated DNA immunoprecipitation (MeDIP). Results: While cell purity was >95% using both cell separation approaches, gene expression analysis revealed significantly higher mRNA levels of several inflammatory genes in EDTA/DTT when compared to enzymatically released cells. In contrast, DNA methylation of selected genes was less variable and only revealed subtle differences. Comparison of DNA methylation of the epithelial cell marker EPCAM in unseparated whole biopsy samples with separated epithelium (i.e. EPCAM positive and negative fraction) demonstrated significant differences in DNA methylation between all three tissue fractions indicating cell type specific methylation patterns can be masked in unseparated tissue samples. Conclusions: Taken together, our data highlight the importance of considering the potential effect of cell separation on gene expression as well as DNA methylation signatures. The decision to separate tissue samples will therefore depend on study design and specific separation protocols. [ABSTRACT FROM AUTHOR]

A conference paper about the transcriptome dysregulation and epigenome signature responses in hepatocytes upon hepatitis B virus infection and therapy is presented. It informs that the infection with hepatitis B virus (HBV) induces transregulation of the host cell gene expression and eventually malignant transformation. It further informs that the HBV X protein acts as a transactivator of cellular promoters.

A conference paper on a study on newborns related to placental insufficiency and consecutive dystrophy. It mentions that this study involved 42 patients and endothelial nitric oxide synthase (eNOS) copy number was high in infants with impaired perfusion in the uterine. It further states that eNOS expression in umbilical cord arterial endothelial cells are triggered by adaptive chromatin signatures.