A novel method for high-throughput detection and quantification of neutrophil extracellular traps reveals ROS-independent NET release with immune complexes

AbstractA newly-described first-line immune defence mechanism of neutrophils is the release of neutrophil extracellular traps (NETs). Immune complexes (ICxs) induce low level NET release. As such, the in vitro quantification of NETs is challenging with current methodologies. In order to investigate the role of NET release in ICx-mediated autoimmune diseases, we developed a highly sensitive and automated method for quantification of NETs. After labelling human neutrophils with PKH26 and extracellular DNA with Sytox green, cells are fixed and automatically imaged with 3-dimensional confocal laser scanning microscopy (3D-CLSM). NET release is then quantified with digital image analysis whereby the NET amount (Sytox green area) is corrected for the number of imaged neutrophils (PKH26 area). A high sensitivity of the assay is achieved by a) significantly augmenting the area of the well imaged (11%) as compared to conventional assays (0.5%) and b) using a 3D imaging technique for optimal capture of NETs, which are topologically superimposed on neutrophils. In this assay, we confirmed low levels of NET release upon human ICx stimulation which were positive for citrullinated histones and neutrophil elastase. In contrast to PMA-induced NET release, ICx-induced NET release was unchanged when co-incubated with diphenyleneiodonium (DPI). We were able to quantify NET release upon stimulation with serum from RA and SLE patients, which was not observed with normal human serum. To our knowledge, this is the first semi-automated assay capable of sensitive detection and quantification of NET release at a low threshold by using 3D CLSM. The assay is applicable in a high-throughput manner and allows the in vitro analysis of NET release in ICx-mediated autoimmune diseases

Neurodevelopmental evaluation and referral practices in children with congenital heart disease in central South Africa

Introduction: Children with congenital heart disease (CHD) are at higher risk for developmental delays than the general population. The American Heart Association (AHA) published a guideline to address these concerns in 2012. This study determined the neurodevelopmental evaluation and referral practices of practitioners in central South Africa.Method: An online survey was administered to practitioners (n=45) including paediatric cardiologists (n=4), cardiothoracic surgeons (n=4) and general paediatricians (n=37). Information on practitioner characteristics, awareness of the 2012 AHA guideline; and neurodevelopmental evaluation and referral practices was collected.Results: Twenty-one practitioners responded, including paediatric cardiologists (n=4), cardiothoracic surgeons (n=2) and paediatricians (n=15). Data for 20 practitioners was included. Despite most practitioners (n=18) indicating guidelines for the management of development were important, the majority (n=16; 80%) were unaware of the guideline. Most practitioners (n=18; 90%) failed to risk stratify children to identify those to be evaluated. Children with developmental delays were referred for formal developmental evaluation (n=11; 55%) and to intervention therapies (n= 15; 75%).Conclusion: Most practitioners are unaware of the 2012 AHA guideline. Awareness of the developmental risks associated with CHD and implementation of the guideline could promote early identification of developmental delays with referral to intervention therapies

Cognitive emotion regulation strategies and depressive symptoms: differences between males and females.

FSW - Self-regulation models for health behavior and Psychopathology - Ou

Cognitive coping strategies and symptoms of depression and anxiety: A comparison between adolescents and adults.

FSW - Self-regulation models for health behavior and Psychopathology - Ou

The relationship between cognitive emotion regulation strategies and emotional problems: comparison between a clinical and a non-clinical sample.

FSW - Self-regulation models for health behavior and Psychopathology - Ou

Establishing the role of rare coding variants in known Parkinson's disease risk loci

Many common genetic factors have been identified to contribute to Parkinson's disease (PD) susceptibility, improving our understanding of the related underlying biological mechanisms. The involvement of rarer variants in these loci has been poorly studied. Using International Parkinson's Disease Genomics Consortium data sets, we performed a comprehensive study to determine the impact of rare variants in 23 previously published genome-wide association studies (GWAS) loci in PD. We applied Prix fixe to select the putative causal genes underneath the GWAS peaks, which was based on underlying functional similarities. The Sequence Kernel Association Test was used to analyze the joint effect of rare, common, or both types of variants on PD susceptibility. All genes were tested simultaneously as a gene set and each gene individually. We observed a moderate association of common variants, confirming the involvement of the known PD risk loci within our genetic data sets. Focusing on rare variants, we identified additional association signals for LRRK2, STBD1, and SPATA19. Our study suggests an involvement of rare variants within several putatively causal genes underneath previously identified PD GWAS peaks

TNO at TRECVID 2013 : multimedia event detection and instance search

We describe the TNO system and the evaluation results for TRECVID 2013 Multimedia Event Detection (MED) and instance search (INS) tasks. The MED system consists of a bag-of-word (BOW) approach with spatial tiling that uses low-level static and dynamic visual features, an audio feature and high-level concepts. Automatic speech recognition (ASR) and optical character recognition (OCR) are not used in the system. In the MED case with 100 example training videos, support-vector machines (SVM) are trained and fused to detect an event in the test set. In the case with 0 example videos, positive and negative concepts are extracted as keywords from the textual event description and events are detected with the high-level concepts. The MED results show that the SIFT keypoint descriptor is the one which contributes best to the results, fusion of multiple low-level features helps to improve the performance, and the textual event-description chain currently performs poorly. The TNO INS system presents a baseline open-source approach using standard SIFT keypoint detection and exhaustive matching. In order to speed up search times for queries a basic map-reduce scheme is presented to be used on a multi-node cluster. Our INS results show above-median results with acceptable search times.This research for the MED submission was performed in the GOOSE project, which is jointly funded by the enabling technology program Adaptive Multi Sensor Networks (AMSN) and the MIST research program of the Dutch Ministry of Defense. The INS submission was partly supported by the MIME project of the creative industries knowledge and innovation network CLICKNL.peer-reviewe

Cognitive, Behavioral and Goal Adjustment Coping and Depressive Symptoms in Young People with Diabetes: A Search for Intervention Targets for Coping Skills Training

The aim of the present study was to find relevant coping factors for the development of psychological intervention programs for young people with Type 1 (T1) diabetes. A wide range of coping techniques was studied, including cognitive coping, behavioral coping and goal adjustment coping. A total of 78 young people with T1 diabetes participated. They were contacted through a social networking website, several Internet sites, and flyers. A wide range of coping techniques appeared to be related to depressive symptoms. Especially the cognitive coping strategies self-blame, rumination, refocus positive, and other-blame, together with goal adjustment coping, were of importance. A large proportion of the variance of depressive symptoms could be explained (65 %). These findings suggest that these specific coping strategies should be part of coping skills trainings for young people with T1 diabetes

Different Aspects of Classical Pathway Overactivation in Patients With C3 Glomerulopathy and Immune Complex-Mediated Membranoproliferative Glomerulonephritis

The rare and heterogeneous kidney disorder C3 glomerulopathy (C3G) is characterized by dysregulation of the alternative pathway (AP) of the complement system. C3G is often associated with autoantibodies stabilizing the AP C3 convertase named C3 nephritic factors (C3NeF). The role of classical pathway (CP) convertase stabilization in C3G and related diseases such as immune complex-mediated membranoproliferative glomerulonephritis (IC-MPGN) remains largely unknown. Here, we investigated the CP convertase activity in patients with C3G and IC-MPGN. Using a refined two-step hemolytic assay, we measured the stability of CP convertases directly in the serum of 52 patients and 17 healthy controls. In four patients, CP convertase activity was prolonged compared to healthy controls, i.e. the enzymatic complex was stabilized. In three patients (2 C3G, 1 IC-MPGN) the convertase stabilization was caused by immunoglobulins, indicating the presence of autoantibodies named C4 nephritic factors (C4NeFs). Importantly, the assay also enabled detection of non-immunoglobulin-mediated stabilization of the CP convertase in one patient with C3G. Prolonged CP convertase activity coincided with C3NeF activity in all patients and for up to 70 months of observation. Crucially, experiments with C3-depleted serum showed that C4NeFs stabilized the CP C3 convertase (C4bC2a), that does not contain C3NeF epitopes. All patients with prolonged CP convertase activity showed clear signs of complement activation, i.e. lowered C3 and C5 levels and elevated levels of C3d, C3bc, C3bBbP, and C5b-9. In conclusion, this work provides new insights into the diverse aspects and (non-)immunoglobulin nature of factors causing CP convertase overactivity in C3G/IC-MPGN.</p

The NET-effect of combining rituximab with belimumab in severe systemic lupus erythematosus

Afdeling Klinische Chemie en Laboratoriumgeneeskunde (AKCL