Modified protocol of harvesting, extraction, and normalization approaches for gas chromatography mass spectrometry-based metabolomics analysis of adherent cells grown under high fetal calf serum conditions

A gas chromatography mass spectrometry (GC-MS) metabolomics protocol was modified for quenching, harvesting, and extraction of metabolites from adherent cells grown under high (20%) fetal calf serum conditions. The reproducibility of using either 50% or 80% methanol for quenching of cells was compared for sample harvest. To investigate the efficiency and reproducibility of intracellular metabolite extraction, different volumes and ratios of chloroform were tested. Additionally, we compared the use of total protein amount versus cell mass as normalization parameters. We demonstrate that the method involving 50% methanol as quenching buffer followed by an extraction step using an equal ratio of methanol:chloroform:water (1:1:1, v/v/v) followed by the collection of 6 mL polar phase for GC-MS measurement was superior to the other methods tested. Especially for large sample sets, its comparative ease of measurement leads us to recommend normalization to protein amount for the investigation of intracellular metabolites of adherent human cells grown under high (or standard) fetal calf serum conditions. To avoid bias, care should be taken beforehand to ensure that the ratio of total protein to cell number are consistent among the groups tested. For this reason, it may not be suitable where culture conditions or cell types have very different protein outputs (e.g., hypoxia vs. normoxia). The full modified protocol is available in the Supplementary Materials

Magnetooptical determination of a topological index

When a Dirac fermion system acquires an energy-gap, it is said to have either trivial (positive energy-gap) or non-trivial (negative energy-gap) topology, depending on the parity ordering of its conduction and valence bands. The non-trivial regime is identified by the presence of topological surface or edge-state dispersing in the energy gap of the bulk and is attributed a non-zero topological index. In this work, we show that such topological indices can be determined experimentally via an accurate measurement of the effective velocity of bulk massive Dirac fermions. We demonstrate this analytically starting from the Bernevig-Hughes-Zhang Hamiltonian (BHZ) to show how the topological index depends on this velocity. We then experimentally extract the topological index in Pb1-xSnxSe and Pb1-xSnxTe using infrared magnetooptical Landau level spectroscopy. This approach is argued to be universal to all material classes that can be described by a BHZ-like model and that host a topological phase transition.Comment: Accepted for publication in Nature Partner Journal Quantum Material

Differentiating Staphylococcus aureus from Escherichia coli mastitis: S. aureus triggers unbalanced immune-dampening and host cell invasion immediately after udder infection

The etiology determines quality and extent of the immune response after udder infection (mastitis). Infections with Gram negative bacteria (e.g. Escherichia coli) will quickly elicit strong inflammation of the udder, fully activate its immune defence via pathogen receptor driven activation of I kappa B/NF-kappa B signaling. This often eradicates the pathogen. In contrast, Gram-positive bacteria (e.g. Staphylococcus aureus) will slowly elicit a much weaker inflammation and immune response, frequently resulting in chronic infections. However, it was unclear which immune regulatory pathways are specifically triggered by S. aureus causing this partial immune subversion. We therefore compared in first lactating cows the earliest (1-3 h) udder responses against infection with mastitis causing pathogens of either species. Global transcriptome profiling, bioinformatics analysis and experimental validation of key aspects revealed as S. aureus infection specific features the (i) failure to activating I kappa B/NF-kappa B signaling;(ii) activation of the wnt/beta-catenin cascade resulting in active suppression of NF-kappa B signaling and (iii) rearrangement of the actin-cytoskeleton through modulating Rho GTPase regulated pathways. This facilitates invasion of pathogens into host cells. Hence, S. aureus mastitis is characterized by eliciting unbalanced immune suppression rather than inflammation and invasion of S. aureus into the epithelial cells of the host causing sustained infection

Collagen type XVIII/endostatin is differentially expressed in primary and metastatic colorectal cancers and ovarian carcinomas

Collagen type XVIII (C18) is a nonfibrillar collagen of basement membranes. Its C-terminal fragment, endostatin, has been identified as an inhibitor of angiogenesis. C18 is predominantly expressed by hepatocytes of normal, cirrhotic and neoplastic liver. We compared the patterns of C18 RNA-expression in colonic adenocarcinoma metastases, which represent the most frequently occurring liver tumours, to normal colon mucosa, to primary colon cancers and to ovarian cancers which are often morphologically similar to colonic cancer or metastasis. Two C18-specific RNA-probes were generated to perform in situ hybridization combined with immunohistochemistry for cytokeratin, vimentin and the endothelial marker CD31, in order to characterize the C18-expressing cells. C18/endostatin protein was localized by immunohistology. In colorectal carcinomas and their liver metastases high levels of C18 transcripts were observed in endothelial cells and fibroblasts/myofibroblasts, whereas C18 RNA was virtually absent from carcinoma cells. Ovarian carcinomas displayed high C18 RNA expression both in carcinoma and stromal cells, indicating that induction of C18 transcription in tumour stromal cells is independent of the ability of carcinoma cells to express C18. While the role of tumour cell derived C18 in cancer growth regulation remains unknown, stimulation of proteolysis of the locally strongly expressed C18 to endostatin could offer an attractive approach for a targeted antineoplastic therapy. © 2001 Cancer Research Campaign   http://www.bjcancer.co

Spectrum radial velocity analyser (SERVAL). High-precision radial velocities and two alternative spectral indicators

Context: The CARMENES survey is a high-precision radial velocity (RV) programme that aims to detect Earth-like planets orbiting low-mass stars. Aims: We develop least-squares fitting algorithms to derive the RVs and additional spectral diagnostics implemented in the SpEctrum Radial Velocity Analyser (SERVAL), a publicly available python code. Methods: We measured the RVs using high signal-to-noise templates created by coadding all available spectra of each star.We define the chromatic index as the RV gradient as a function of wavelength with the RVs measured in the echelle orders. Additionally, we computed the differential line width by correlating the fit residuals with the second derivative of the template to track variations in the stellar line width. Results: Using HARPS data, our SERVAL code achieves a RV precision at the level of 1m/s. Applying the chromatic index to CARMENES data of the active star YZ CMi, we identify apparent RV variations induced by stellar activity. The differential line width is found to be an alternative indicator to the commonly used full width half maximum. Conclusions: We find that at the red optical wavelengths (700--900 nm) obtained by the visual channel of CARMENES, the chromatic index is an excellent tool to investigate stellar active regions and to identify and perhaps even correct for activity-induced RV variations.Comment: 13 pages, 13 figures. A&A in press. Code is available at https://github.com/mzechmeister/serva

Time Triggered Protocol (TTP) for Integration Modular Avionics (IMA)

This viewgraph presentation is a review of the Time Triggered Protocol, designed to work with NASA's Integrated Safety-Critical Advanced Avionics Communication and Control (ISAACC) system. ISAACC is the product of the Propulsion High-Impact Avionics Technologies (PHIAT) project at NASA Marshall Space Flight Center (MSFC) during FY03 to the end of FY05. The goal is an avionics architecture suitable for control and monitoring of safety critical systems of manned spacecraft. It must be scalable to allow its use in robotic vehicles or launch pad and propulsion test stand monitoring and control systems. The developed IMA should have: a common power supply and rugged chassis for a set of modules, many upgradeable software functions on one module (i.e. processing unit Reduced weight, straightforward update and system integration. It is also important that it have Partitioning and a Memory Management Unit (MMU

Magnetic fields in M dwarfs from the CARMENES survey

M dwarfs are known to generate the strongest magnetic fields among main-sequence stars with convective envelopes, but the link between the magnetic fields and underlying dynamo mechanisms, rotation, and activity still lacks a consistent picture. In this work we measure magnetic fields from the high-resolution near-infrared spectra taken with the CARMENES radial-velocity planet survey in a sample of 29 active M dwarfs and compare our results against stellar parameters. We use the state-of-the-art radiative transfer code to measure total magnetic flux densities from the Zeeman broadening of spectral lines and filling factors. We detect strong kG magnetic fields in all our targets. In 16 stars the magnetic fields were measured for the first time. Our measurements are consistent with the magnetic field saturation in stars with rotation periods P<4d. The analysis of the magnetic filling factors reveal two different patterns of either very smooth distribution or a more patchy one, which can be connected to the dynamo state of the stars and/or stellar mass. Our measurements extend the list of M dwarfs with strong surface magnetic fields. They also allow us to better constrain the interplay between the magnetic energy, stellar rotation, and underlying dynamo action. The high spectral resolution and observations at near-infrared wavelengths are the beneficial capabilities of the CARMENES instrument that allow us to address important questions about the stellar magnetism.Comment: 13 pages of main text, 14 pages of online material, 2 table

Using the first European Breeding Bird Atlas for science and perspectives for the new Atlas

Capsule The first European Bird Census Council (EBCC) Atlas of European Breeding Birds has been widely used in scientific publications. Aims To quantify how scientific publications have used data from the first European Bird Census Council (EBCC) Atlas of European Breeding Birds, what the topics of these studies have been, and to identify key aspects in which a second European Breeding Bird Atlas will provide new opportunities for basic and applied science. Methods We searched Google Scholar to find papers published in scientific journals that cited the first atlas. We analysed the contents of a random selection of 100 papers citing this atlas and described the way these papers used information from it. Results The first atlas has been cited in 3150 scientific publications, and can be regarded as a fundamental reference for studies about birds in Europe. It was extensively used as a key reference for the studied bird species. A substantial number of papers re-analysed atlas data to derive new information on species distribution, ecological traits and population sizes. Distribution and ecology were the most frequent topics of studies referring to the atlas, but this source of information was used in a diverse range of studies. In this context, climate change, impact of agriculture and habitat loss were, by order, the most frequently studied environmental pressures. Constraints in the atlas, such as the poor coverage in the east of Europe, the lack of information on distribution change and the coarse resolution were identified as issues limiting the use of the atlas for some purposes. Conclusions This study demonstrates the scientific value of European-wide breeding bird atlases. A second atlas, with its almost complete coverage across Europe, the incorporation of changes in distribution between the two atlases and the inclusion of modelled maps at a resolution of 10 x 10 km will certainly become a key data source and reference for researchers in the near future.Peer reviewe