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Neutrophils, the most abundant human immune cells, are rapidly recruited to sites of infection, where they fulfill their life-saving antimicrobial functions. While traditionally regarded as short-lived phagocytes, recent findings on long-term survival, neutrophil extracellular trap (NET) formation, heterogeneity and plasticity, suppressive functions, and tissue injury have expanded our understanding of their diverse role in infection and inflammation. This review summarises our current understanding of neutrophils in host-pathogen interactions and disease involvement, illustrating the versatility and plasticity of the neutrophil, moving between host defence, immune modulation, and tissue damage. [ABSTRACT FROM AUTHOR]

Posttranslational modification of the neural cell adhesion molecule (NCAM) by polysialic acid (polySia) is well studied in the nervous system and described as a dynamic modulator of plastic processes like precursor cell migration, axon fasciculation, and synaptic plasticity. Here, we describe a novel function of polysialylated NCAM (polySia-NCAM) in innate immunity of the lung. In mature lung tissue of healthy donors, polySia was exclusively attached to the transmembrane isoform NCAM-140 and located to intracellular compartments of epithelial cells. In patients with chronic obstructive pulmonary disease, however, increased polySia levels and processing of the NCAM carrier were observed. Processing of polysialylated NCAM was reproduced in a mouse model by bleomycin administration leading to an activation of the inflammasome and secretion of interleukin (IL)-1β. As shown in a cell culture model, polySia-NCAM-140 was kept in the late trans-Golgi apparatus of lung epithelial cells and stimulation by IL-1β or lipopolysaccharide induced metalloprotease-mediated ectodomain shedding, resulting in the secretion of soluble polySia-NCAM. Interestingly, polySia chains of secreted NCAM neutralized the cytotoxic activity of extracellular histones as well as DNA/histone-network-containing 'neutrophil extracellular traps', which are formed during invasion of microorganisms. Thus, shedding of polySia-NCAM by lung epithelial cells may provide a host-protective mechanism to reduce tissue damage during inflammatory processes. [ABSTRACT FROM AUTHOR]

Neutrophils play an important role in innate immunity by defending the host organism against invading microorganisms. Antimicrobial activity of neutrophils is mediated by release of antimicrobial peptides, phagocytosis as well as formation of neutrophil extracellular traps (NET). These structures are composed of DNA, histones and granular proteins such as neutrophil elastase and myeloperoxidase. This study focused on the influence of NET on the host cell functions, particularly on human alveolar epithelial cells as the major cells responsible for gas exchange in the lung. Upon direct interaction with epithelial and endothelial cells, NET induced cytotoxic effects in a dose-dependent manner, and digestion of DNA in NET did not change NET-mediated cytotoxicity. Pre-incubation of NET with antibodies against histones, with polysialic acid or with myeloperoxidase inhibitor but not with elastase inhibitor reduced NET-mediated cytotoxicity, suggesting that histones and myeloperoxidase are responsible for NET-mediated cytotoxicity. Although activated protein C (APC) did decrease the histone-induced cytotoxicity in a purified system, it did not change NET-induced cytotoxicity, indicating that histonedependent cytotoxicity of NET is protected against APC degradation. Moreover, in LPS-induced acute lung injury mouse model, NET formation was documented in the lung tissue as well as in the bronchoalveolar lavage fluid. These data reveal the important role of protein components in NET, particularly histones, which may lead to host cell cytotoxicity and may be involved in lung tissue destruction. [ABSTRACT FROM AUTHOR]

We have evaluated the effects of acetonitrile on the structure and function of bovine carbonic anhydrase II. The potential structural and functional changes in carbonic anhydrase in the presence of different acetonitrile/buffer ratios (0%, 17.5% and 47.5% v/v) were determined using a variety of methods. These included simple spectrophotometric methods to record enzyme velocity, fluorescence measurements and calculation of accessible surface area (ASA) to identify possible alterations in tertiary structure of the protein, CD measurements to search for secondary structure conversions, and thermal scanning to determine structural stability of the protein in different media. The Far-UV CD studies indicated that carbonic anhydrase, for the most part, retains its secondary structure in the presence of acetonitrile. Fluorescence measurements using iodide ion and ANS along with ASA calculations revealed that in the presence of acetonitrile some degree of conformational change occurs in the carbonic anhydrase structure. In addition to the hydrophobic pockets, two additional tryptophanyl residues become exposed to the solvent, thereby increasing the surface hydrophobicity of the protein. These alterations dramatically reduce the catalytic activity, thermal stability, and aggregation velocity of the enzyme. Thus, our results support a molten globule-like structure of carbonic anhydrase in the presence of acetonitrile. [ABSTRACT FROM PUBLISHER]

In this article, the author shares her experience of attending the European Medical Writers Association (EMWA) conferences which involved topics like regulatory writing, proofreading techniques, and writing for the internet.

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. [ABSTRACT FROM AUTHOR]

Abstract: Ribonuclease (RNase) treatment represents a novel mechanism based approach to anticancer therapy as an alternative to the DNA damaging drugs commonly used in clinical practice. Apart from their ribonucleolytic activity, cytotoxic effects have attracted a considerable attention to RNases because of their potential as selective agents for treatment of certain malignancies. Among these enzymes, Binase, an RNase from Bacillus intermedius, has shown promising results. Here, we have found that binase selectively attacked human A549 alveolar adenocarcinoma cells to trigger an apoptotic response, whereas normal lung epithelial cells LEK were not affected by the ribonuclease. The tumor transformation led to the modification of certain cellular characteristics causing cell sensitivity to binase. Although a general mode for RNases cytotoxicity includes their penetration into the cell, translocation to the cytosol and degradation of ribonucleic acid, many aspects of this process have not been fully elucidated. Our data revealed the following time-dependent changes induced by binase in A549 cells: (a) fast permanent internalization of the enzyme during the first hours of treatment; (b) temporary increase in cellular permeability for macromolecules during the 4–6 h of treatment; (c) apoptotic alterations in population after 24 h and (d) DNA fragmentation and cell death after 72 h of treatment with binase. Elucidation of these molecular strategies used by this promising toxin provides us essential information for the development of new anticancer drugs. [Copyright &y& Elsevier]

The goals of this study are to compare the lumbar spine response variance between the hybrid III, test device for human occupant restraint (THOR), and global human body models consortium simplified 50th percentile (GHBMC M50-OS) finite element models and evaluate the sensitivity of lumbar spine injury metrics to multidirectional acceleration pulses for spaceflight landing conditions. The hybrid III, THOR, and GHBMC models were positioned in a baseline posture within a generic seat with side guards and a five-point restraint system. Thirteen boundary conditions, which were categorized as loading condition variables and environmental variables, were included in the parametric study using a Latin hypercube design of experiments. Each of the three models underwent 455 simulations for a total of 1365 simulations. The hybrid III and THOR models exhibited similar lumbar compression forces. The average lumbar compression force was 45% higher for hybrid III (2.2±1.5kN) and 51% higher for THOR (2.0±1.6kN) compared to GHBMC (1.3±0.9kN). Compared to hybrid III, THOR sustained an average 64% higher lumbar flexion moment and an average 436% higher lumbar extension moment. The GHBMC model sustained much lower bending moments compared to hybrid III and THOR. Regressions revealed that lumbar spine responses were more sensitive to loading condition variables than environmental variables across all models. This study quantified the intermodel lumbar spine response variations and sensitivity between hybrid III, THOR, and GHBMC. Results improve the understanding of lumbar spine response in spaceflight landings. [ABSTRACT FROM AUTHOR]

The use of anthropomorphic test devices (ATDs) for calculating injury risk of occupants in spaceflight scenarios is crucial for ensuring the safety of crewmembers. Finite element (FE) modeling of ATDs reduces cost and time in the design process. The objective of this study was to validate a Hybrid III ATD FE model using a multidirection test matrix for future spaceflight configurations. Twenty-five Hybrid III physical tests were simulated using a 50th percentile male Hybrid III FE model. The sled acceleration pulses were approximately half-sine shaped, and can be described as a combination of peak acceleration and time to reach peak (rise time). The range of peak accelerations was 10-20 G, and the rise times were 30-110 ms. Test directions were frontal (-GX), rear (GX), vertical (GZ), and lateral (GY). Simulation responses were compared to physical tests using the correlation and analysis (CORA) method. Correlations were very good to excellent and the order of best average response by direction was -GX (0.916±0.054), GZ (0.841±0.117), GX (0.792±0.145), and finally GY (0.775±0.078). Qualitative and quantitative results demonstrated the model replicated the physical ATD well and can be used for future spaceflight configuration modeling and simulation. [ABSTRACT FROM AUTHOR]

Expanding evidence indicates multiple interactions between the hemostatic system and innate immunity, and the coagulation and complement cascades. Here we show in a tissue factor (TF)-dependent model of flow restriction-induced venous thrombosis that complement factors make distinct contributions to platelet activation and fibrin deposition. Complement factor 3 (C3) deficiency causes prolonged bleeding, reduced thrombus incidence, thrombus size, fibrin and platelet deposition in the ligated inferior vena cava, and diminished platelet activation in vitro. Initial fibrin deposition at the vessel wall over 6 hours in this model was dependent on protein disulfide isomerase (PDI) and TF expression by myeloid cells, but did not require neutrophil extracellular trap formation involving peptidyl arginine deiminase 4. In contrast to C3-/- mice, C5-deficient mice had no apparent defect in platelet activation in vitro, and vessel wall platelet deposition and initial hemostasis in vivo. However, fibrin formation, the exposure of negatively charged phosphatidylserine (PS) on adherent leukocytes, and clot burden after 48 hours were significantly reduced in C5-/- mice compared with wild-type controls. These results delineate that C3 plays specific roles in platelet activation independent of formation of the terminal complement complex and provide in vivo evidence for contributions of complement-dependent membrane perturbations to prothrombotic TF activation on myeloid cells. [ABSTRACT FROM AUTHOR]

Objective To study if neutrophil extracellular traps (NETs) are present in the peritoneal fluid of endometriosis patients. NETs play a crucial role in fighting against microorganisms. However, exaggerated NET production may lead to tissue damage in their vicinity in pathological conditions. Our study evaluates the presence of NETs in endometriosis peritoneal fluid. Study design Peritoneal fluid (PF) was collected in a case-control study from 52 women, who underwent either diagnostic or operative laparoscopy. The control group consisted of 17 women with infertility, chronic pelvic pain, simple or functional cysts or irregular bleeding. The endometriosis group, altogether 35 patients, comprised 19 patients with stage I and II and 16 patients with stage III and IV endometriosis. First we tested whether the PF is able to stimulate NET production. Neutrophils from healthy volunteers were treated with the PF of endometriosis patients and controls and NETs were detected with Sytox orange extracellular DNA dye and immunofluorescence microscopy. Then we evaluated if NETs were already present in the collected PF using the specific myeloperoxidase (MPO)-DNA capture ELISA method, based on the MPO associated with the NET scaffold. Results The PF of endometriosis patients did not stimulate NET release from healthy granulocytes. However, pre-existent NETs could be detected in 17 endometriosis patients out of 35 (49%). In contrary, in the control group NETs were present in only 3 patients out of 17 (18%), ( p = 0.03, OR: 4.4). Moreover, the quantification of NETs showed a significantly higher amount of NETs in endometriosis compared to the controls (0.097 vs. 0.02, p = 0.04). Conclusion This is the first study, which evaluated and described the presence of NETs in the PF of endometriosis patients. Our study shows, that NETs may be involved in the complex pathophysiology of endometriosis. [ABSTRACT FROM AUTHOR]