Functional consequences of sphingomyelinase-induced changes in erythrocyte membrane structure.

Inflammation enhances the secretion of sphingomyelinases (SMases). SMases catalyze the hydrolysis of sphingomyelin into phosphocholine and ceramide. In erythrocytes, ceramide formation leads to exposure of the removal signal phosphatidylserine (PS), creating a potential link between SMase activity and anemia of inflammation. Therefore, we studied the effects of SMase on various pathophysiologically relevant parameters of erythrocyte homeostasis. Time-lapse confocal microscopy revealed a SMase-induced transition from the discoid to a spherical shape, followed by PS exposure, and finally loss of cytoplasmic content. Also, SMase treatment resulted in ceramide-associated alterations in membrane-cytoskeleton interactions and membrane organization, including microdomain formation. Furthermore, we observed increases in membrane fragility, vesiculation and invagination, and large protein clusters. These changes were associated with enhanced erythrocyte retention in a spleen-mimicking model. Erythrocyte storage under blood bank conditions and during physiological aging increased the sensitivity to SMase. A low SMase activity already induced morphological and structural changes, demonstrating the potential of SMase to disturb erythrocyte homeostasis. Our analyses provide a comprehensive picture in which ceramide-induced changes in membrane microdomain organization disrupt the membrane-cytoskeleton interaction and membrane integrity, leading to vesiculation, reduced deformability, and finally loss of erythrocyte content. Understanding these processes is highly relevant for understanding anemia during chronic inflammation, especially in critically ill patients receiving blood transfusions

Working Papers, Open Access and Cyber-Infrastructure in Classical Studies

Princeton–Stanford Working Papers in Classics is a web-based series of work-in-progress scripts by members of two leading departments of classics. It introduces the humanities to a new form of scholarly communication and represents a major advance in the free availability of classical-studies scholarship in cyberspace. This article both reviews the initial performance of this open-access experiment and the benefits and challenges of working papers more generally for classical studies. After two years of operation Princeton–Stanford Working Papers in Classics has proven to be a clear success. This series has built up a large international readership and a sizeable body of preprints and performs important scholarly and community-outreach functions. As this performance is largely due to its congruency with the working arrangements of ancient historians and classicists and the global demand for open-access scholarship, the series confirms the viability of this means of scholarly communication and the likelihood of its expansion in our discipline. But modifications are required to increase the benefits this series brings and the amount of scholarship it makes freely available online. Finally departments wishing to replicate its success will have to consider other important developments, such as the increasing availability of postprints, the linking of research funding to open access, and the emergence of new cyber-infrastructure

Colored spanning graphs for set visualization

We study an algorithmic problem that is motivated by ink minimization for sparse set visualizations. Our input is a set of points in the plane which are either blue, red, or purple. Blue points belong exclusively to the blue set, red points belong exclusively to the red set, and purple points belong to both sets. A red-blue-purple spanning graph (RBP spanning graph) is a set of edges connecting the points such that the subgraph induced by the red and purple points is connected, and the subgraph induced by the blue and purple points is connected.We study the geometric properties of minimum RBP spanning graphs and the algorithmic problems associated with computing them. Specifically, we show that the general problem can be solved in polynomial time using matroid techniques. In addition, we discuss more efficient algorithms for the case in which points are located on a line or a circle, and also describe a fast (12¿+1)-approximation algorithm, where ¿ is the Steiner ratio.Peer ReviewedPostprint (author's final draft

Accelerated apoptotic death and <i>in vivo</i> turnover of erythrocytes in mice lacking functional mitogen- and stress-activated kinase MSK1/2

The mitogen- and stress-activated kinase MSK1/2 plays a decisive role in apoptosis. In analogy to apoptosis of nucleated cells, suicidal erythrocyte death called eryptosis is characterized by cell shrinkage and cell membrane scrambling leading to phosphatidylserine (PS) externalization. Here, we explored whether MSK1/2 participates in the regulation of eryptosis. To this end, erythrocytes were isolated from mice lacking functional MSK1/2 (msk−/−) and corresponding wild-type mice (msk+/+). Blood count, hematocrit, hemoglobin concentration and mean erythrocyte volume were similar in both msk−/− and msk+/+ mice, but reticulocyte count was significantly increased in msk−/− mice. Cell membrane PS exposure was similar in untreated msk−/− and msk+/+ erythrocytes, but was enhanced by pathophysiological cell stressors ex vivo such as hyperosmotic shock or energy depletion to significantly higher levels in msk−/− erythrocytes than in msk+/+ erythrocytes. Cell shrinkage following hyperosmotic shock and energy depletion, as well as hemolysis following decrease of extracellular osmolarity was more pronounced in msk−/− erythrocytes. The in vivo clearance of autologously-infused CFSE-labeled erythrocytes from circulating blood was faster in msk−/− mice. The spleens from msk−/− mice contained a significantly greater number of PS-exposing erythrocytes than spleens from msk+/+ mice. The present observations point to accelerated eryptosis and subsequent clearance of erythrocytes leading to enhanced erythrocyte turnover in MSK1/2-deficient mice

Bodemtoets voor wortelknobbels gereed

Agrobacterium, de veroorzaker van wortelknobbels, vormt een groot probleem in de boomkwekerij. Er is nu een mogelijkheid om de grond vóór de teelt te toetsen op Agrobacteriu

Classification and prediction of Mycobacterium Avium subsp. Paratuberculosis (MAP) shedding severity in cattle based on young stock heifer faecal microbiota composition using random forest algorithms

BACKGROUND: Bovine paratuberculosis is a devastating infectious disease caused by Mycobacterium avium subsp. paratuberculosis (MAP). The development of the paratuberculosis in cattle can take up to a few years and vastly differs between individuals in severity of the clinical symptoms and shedding of the pathogen. Timely identification of high shedding animals is essential for paratuberculosis control and minimization of economic losses. Widely used methods for detection and quantification of MAP, such as culturing and PCR based techniques rely on direct presence of the pathogen in a sample and have little to no predictive value concerning the disease development. In the current study, we investigated the possibility of predicting MAP shedding severity in cattle based on the faecal microbiota composition. Twenty calves were experimentally infected with MAP and faecal samples were collected biweekly up to four years of age. All collected samples were subjected to culturing on selective media to obtain data about shedding severity. Faecal microbiota was profiled in a subset of samples (n = 264). Using faecal microbiota composition and shedding intensity data a random forest classifier was built for prediction of the shedding status of the individual animals. RESULTS: The results indicate that machine learning approaches applied to microbial composition can be used to classify cows into groups by severity of MAP shedding. The classification accuracy correlates with the age of the animals and use of samples from older individuals resulted in a higher classification precision. The classification model based on samples from the first 12 months of life showed an AUC between 0.78 and 0.79 (95% CI), while the model based on samples from animals older than 24 months showed an AUC between 0.91 and 0.92 (95% CI). Prediction for samples from animals between 12 and 24 month of age showed intermediate accuracy [AUC between 0.86 and 0.87 (95% CI)]. In addition, the results indicate that a limited number of microbial taxa were important for classification and could be considered as biomarkers. CONCLUSIONS: The study provides evidence for the link between microbiota composition and severity of MAP infection and shedding, as well as lays ground for the development of predictive diagnostic tools based on the faecal microbiota composition

eXamine : a Cytoscape app for exploring annotated modules in networks

Background. Biological networks have growing importance for the interpretation of high-throughput "omics" data. Statistical and combinatorial methods allow to obtain mechanistic insights through the extraction of smaller subnetwork modules. Further enrichment analyses provide set-based annotations of these modules. Results. We present eXamine, a set-oriented visual analysis approach for annotated modules that displays set membership as contours on top of a node-link layout. Our approach extends upon Self Organizing Maps to simultaneously lay out nodes, links, and set contours. Conclusions. We implemented eXamine as a freely available Cytoscape app. Using eXamine we study a module that is activated by the virally-encoded G-protein coupled receptor US28 and formulate a novel hypothesis about its functioning

Temsirolimus ameliorates lamin status and nuclear shape in HGPS cells.

<p>(A) Representative Western blots of lamin A/C, progerin, and β-actin in control and HGPS total cell extracts isolated from either mock-treated cells or cells treated with 1.0 μM Temsirolimus daily for a period of 9 days. (B) Fold-expression of lamin A, progerin, and lamin C was determined for each sample analyzed by western blotting with anti-lamin A/C antibody in panel (A) and normalized to βan -Actin (*p-value ≤ 0.05; n = 5). (C) The frequency of misshapen nuclei (dysmorphic) after 9 days of treatment with either the vehicle or 1.0 μM Temsirolimus. An average of 900 nuclei were examined for each condition, and each experiment was repeated 3 times. (D) Immunochemistry was performed on mock-treated or Temsirolimus-treated control (GMO3349C) and HGPS (HGADFN003) fibroblasts after 9 days using antibodies against the indicated proteins (lamin A/C, progerin, and lamin B1). Representative images are shown (n = 4). Scale-bar: 20 μm.</p