OWEZ pelagic fish, progress report and progression after T1

This report presents the current status of pelagic fish research in the Offshore Windfarm Egmond aan Zee (OWEZ) after having finalized the T0 baseline survey in 2003 and the T1 survey after construction of the wind farm in 2007. The T0 study indicated a highly dynamic pelagic fish community along the Dutch coast: species composition were found highly variable on a local scale and clear hot spots appeared abandoned when returning the next day. This picture was confirmed in the T1 survey where the species composition of the catches in the entire coastal zone were found completely different from the ones 4 years earlier. Although no clear and direct effect of the wind farm has been observed yet, the above described information tells us to look for more local effects where we should focus on underlying processes in fish community behaviour. The effects of a single windfarm seem difficult to measure but when the Dutch government allows a drastic increase in such wind farms in the North Sea, the local effect will become a regional effect and might start to influence complete fish stocks. Therefore, complementary research will be executed by studying fish behaviour in the wind farm itself

File forensics for RAW camera image formats

Recent research in multimedia forensics has developed a variety of methods to detect image tampering and to identify the origin of image files. Many of these techniques are based on characteristics in the JPEG format, as it is the most used file format for digital images. In recent years RAW image formats have gained popularity among amateur and professional photographers. This increase in their use and possible misuse makes these file formats an important subject to file forensic examinations. The aim of this paper is to explore to which extend methods previously developed for images in JPEG format can be applied to RAW image formats

Magnetofection potentiates gene delivery to cultured endothelial cells

Modification of cellular functions by overexpression of genes is increasingly practised for research of signalling pathways, but restricted by limitations of low efficiency. We investigated whether the novel technique of magnetofection (MF) could enhance gene transfer to cultured primary endothelial cells. MF of human umbilical vein endothelial cells (HUVEC) increased transfection efficiency of a luciferase reporter gene up to 360-fold compared to various conventional transfection systems. In contrast, there was only an up to 1.6-fold increase in toxicity caused by MF suggesting that the advantages of MF outbalanced the increase in toxicity. MF efficiently increased transfection efficiency using several commercially available cationic lipid transfection reagents and polyethyleneimine (PEI). Using PEI, even confluent HUVEC could be efficiently transfected to express luciferase activity. Using a green fluorescent protein vector maximum percentages of transfected cells amounted up to 38.7% while PEI without MF resulted in only 1.3% transfected cells. Likewise, in porcine aortic endothelial cells MF increased expression of a luciferase or beta-galactosidase reporter, reaching an efficiency of 37.5% of cells. MF is an effective tool for pDNA transfection of endothelial cells allowing high efficiencies. It may be of great use for investigating protein function in cell culture experiments

Nitric oxide enhances de novo formation of endothelial gap junctions

Objective: Gap junctions (formed by connexins, Cx) are important for functional coordination of cells in the vascular wall. However, little is known about their physiological regulation in this tissue. We examined the effects of nitric oxide (NO), an important mediator of vasomotion, wound healing and angiogenesis, on the formation of gap junctions in endothelial cells (human umbilical vein endothelial cells, HUVEC). Methods: Flow cytometry was used to determine dye transfer through newly formed gap junctions between acutely coincubated HUVECs. Parallel experiments in wild-type HeLa cells (no connexins) and transfected HeLa cells exclusively expressing Cx43, Cx40 or Cx37 were performed to determine the specific role of Cx subtypes. The intracellular distribution of Cx40 was examined after fractionation with triton by Western blotting. Intracellular levels of cGMP and cAMP were measured by radioimmunoassay. Results: The NO donor SNAP (1 μM) enhanced gap-junctional coupling in HUVECs by about 40%. This was associated with an enhanced incorporation of Cx40 into the membrane. Both effects were restricted to Cx40 as analyzed in experiments with Cx-selective HeLa cells. The NO-induced increase in cell coupling was elicited by a corresponding rise of cGMP, which secondarily increased intracellular cAMP levels. The latter was an integral part of the signal cascade, since the protein kinase A (PKA) inhibitor H89 blocked the SNAP-induced incorporation of Cx40 into the plasma membrane. Conclusions: We conclude that NO is a potent modulator of gap-junctional coupling in endothelial cells. It enhances de novo formation of endothelial gap junctions by increasing incorporation of Cx40 into the plasma membrane due to PKA activation

Gap-junctional coupling between neutrophils and endothelial cells

Communication between leukocytes and endothelial cells is crucial for inflammatory reactions. Paracrine cross-talk and outside-in signaling (via adhesion molecules) have been characterized as communication pathways to date. As leukocytes and endothelial cells express connexins, we considered intercellular communication via gap junctions an intriguing additional concept. We found that gap-junctional coupling between neutrophils and endothelium occurred in a time-dependent, bidirectional manner and was facilitated by adhesion. After blockade of connexins, transmigration of neutrophils through the endothelial layer was enhanced, and the barrier function of cell monolayers was reduced during transmigration. Tumor necrosis factor α decreased coupling. In the presence of connexins, transmigration of neutrophils did not alter permeability. Thus, neutrophils couple to endothelium via gap junctions, functionally modulating transmigration and leakiness. Gapjunctional coupling may be a novel way of leukocyte- endothelial communication

Shear Stress Induces the Release of an Endothelial Elastase: Role in Integrin alpha(v)beta(3)-Mediated FGF-2 Release

Background/Aims: Laminar shear stress is an important stimulus in the endothelium-dependent control of vascular tone and of vascular remodeling processes. Based on previous studies demonstrating integrin-mediated release of fibroblast growth factor 2 (FGF-2), we investigated whether shear stress-induced integrin activation requires the involvement of an extracellular protease. Methods: Cultured porcine aortic endothelial cells (PAEC) were exposed to laminar shear stress (16 dyn/cm(2)), whereas static cells served as controls. Results: Exposure of PAEC to shear stress led to an increased activity of a protease in supernatants. This protease could be characterized as elastase but was different from neutrophil and pancreatic elastases. The enhanced activity was accompanied by the activation of integrin alpha(v)beta(3) and p38 MAPK, and followed by an increased FGF-2 concentration in the supernatant. Pretreatment with inhibitors of either elastase or integrin alpha(v)beta(3) resulted in a reduction of FGF-2 release. The observed effects of shear stress on integrin alpha(v)beta(3) and p38 MAPK activation, as well as on FGF-2 release could be mimicked by application of pancreatic elastase to static endothelial cells. Conclusion: By inducing the release of an endothelial elastase, shear stress induces an integrin-dependent release of FGF-2 from endothelial cells. Copyright (C) 2011 S. Karger AG, Base

Use of Recycled Crushed Concrete (RCC) Fines for Potential Soil Stabilization

This study evaluated the use of Recycled Crushed Concrete (RCC) Fines for potential soil stabilization. Soil stabilization is the enhancement of subgrade stability to improve the constructability of successive pavement layers. Use of RCC fines may not only provide less costly alternatives for subgrade stabilization, but their use may also alleviate landfill disposal challenges

Sensitive superoxide detection in vascular cells by the new chemiluminescence dye L-012

The detection superoxide production in vascular cells is usually limited by a low sensitivity of available assays, We tested the applicability of the luminol derivate L-012 {[}8-amino-5-chloro-7-phenylpyridol{[}3,4-d]pyridazine-l,4(2H,3H)dione] to measure superoxide production in cultured endothelial cells (human umbilical vein endothelial cells) and rat aortic segments. Following stimulation with the protein kinase stimulator phorbol 12-myristate 13-acetate (PMA, 1 mu M) there was an 2,8-fold increase of L-012 chemiluminescence, whereas incubation with angiotensin II (100 nM) did not result in a measurable increase. Addition of vanadate (100 mu M) considerably increased the chemiluminescence (up to 17-fold) after PMA and made possible the detection of an enhanced superoxide production after stimulation with angiotensin II (by 1.7-fold). This was due to a similar to 9-fold increase in signal intensity of L-012 in the presence of vanadate, Prolonged incubation with vanadate also led to a tyrosine phosphorylation-dependent increase in superoxide formation which was predominantly produced by an NAD(P)H oxidase. Short-Term vanadate-enhanced L-012 chemiluminescence represents a highly sensitive assay making it possible to detect small changes of superoxide formation in intact vascular cells. Copyright(C) 1999 S. Karger AG. Basel

Crucial role of local peroxynitrite formation in neutrophil-induced endothelial cell activation

Introduction and methods: The reaction of superoxide anions and NO not only results in a decreased availability of NO, but also leads to the formation of peroxynitrite, the role of which in the cardiovascular system is still discussed controversially. In cultured human endothelial cells, we studied whether there is a significant interaction between endothelial NO and neutrophil-derived superoxide anions in terms of endothelial peroxynitrite formation. We particularly studied whether a significantly higher redox-stress can be found in those endothelial cells directly adjacent to an activated neutrophil. Results: A considerable part of the 2,7-dihydrodichlorofluoresceine signal in endothelial cells was due to oxidation by peroxynitrite. Providing superoxide radicals by enzymatic source or by the neutrophil respiratory burst increased the fluorescence, which was attenuated by blockade of endothelial NO-synthase, suggesting that peroxynitrite was formed from neutrophil- or extracellular enzyme-derived superoxide and endothelial NO. Considerably higher fluorescence intensity was observed in endothelial cells in direct neighborhood to a neutrophil. This was particularly pronounced in the presence of a NO-donor and was accompanied by a strong activation of NF-κB and increased expression of E-selectin in these cells. Conclusion: Endothelial cells adjacent to neutrophils may have elevated levels of peroxynitrite that result in an increased expression of adhesion molecules. Such cells might represent a preferential site for adhesion and migration of additional neutrophils when simultaneously high concentrations of NO and neutrophil-derived superoxide are present

Provenance analysis for instagram photos

As a feasible device fingerprint, sensor pattern noise (SPN) has been proven to be effective in the provenance analysis of digital images. However, with the rise of social media, millions of images are being uploaded to and shared through social media sites every day. An image downloaded from social networks may have gone through a series of unknown image manipulations. Consequently, the trustworthiness of SPN has been challenged in the provenance analysis of the images downloaded from social media platforms. In this paper, we intend to investigate the effects of the pre-defined Instagram images filters on the SPN-based image provenance analysis. We identify two groups of filters that affect the SPN in quite different ways, with Group I consisting of the filters that severely attenuate the SPN and Group II consisting of the filters that well preserve the SPN in the images. We further propose a CNN-based classifier to perform filter-oriented image categorization, aiming to exclude the images manipulated by the filters in Group I and thus improve the reliability of the SPN-based provenance analysis. The results on about 20, 000 images and 18 filters are very promising, with an accuracy higher than 96% in differentiating the filters in Group I and Group II