Analysis of doping control samples using supercritical fluid chromatography-tandem mass spectrometry: Ready for routine use

Supercritical fluid chromatography is proving to be a good separation and sample preparation tool for various analytical applications and, as such, has gained the attention of the anti-doping community. Here, the applicability of supercritical fluid chromatography hyphenated to tandem mass spectrometry for routine doping control analysis was tested. A multi-analyte method was developed to cover 197 drugs and metabolites that are prohibited in sport. More than 1000 samples were analyzed by applying a “dilute and inject” approach after hydrolysis of glucuronide metabolites. Additionally, a comparison with routinely used liquid chromatography-mass spectrometry was performed with 250 of the 1000 samples and a number of past positive anti-doping samples. It revealed some features where supercritical fluid chromatography-tandem mass spectrometry was found to be complementary or advantageous to liquid chromatography-mass spectrometry for anti-doping purposes, such as better retention of analytes that are poorly retained in reversed-phase liquid chromatography. Our results suggest that supercritical fluid chromatography-tandem mass spectrometry is sensitive (limit of detection <50% relevant minimum required performance level required by the World Anti-Doping Agency for anti-doping analysis), reproducible, robust, precise (analytes of interest area coefficient of variation <5%; retention time difference coefficient of variation <1%) and complementary to existing techniques currently used for routine analysis in the World Anti-Doping Agency accredited laboratories

Exhaled and nasal nitric oxide in laryngectomized patients

<p>Abstract</p> <p>Background</p> <p>Nitric oxide (NO) shows differing concentrations in lower and upper airways. Patients after total laryngectomy are the only individuals, in whom a complete separation of upper and lower airways is guaranteed. Thus the objective of our study was to assess exhaled and nasal NO in these patients.</p> <p>Methods</p> <p>Exhaled bronchial NO (FE_NO) and nasal nitric oxide (nNO) were measured in patients after total laryngectomy (n = 14) and healthy controls (n = 24). To assess lung function we additionally performed spirometry. Co-factors possibly influencing NO, such as smoking, infections, and atopy were excluded.</p> <p>Results</p> <p>There was a markedly (p < 0.001) lower FE_NOin patients after total laryngectomy (median (range): 4 (1-22) ppb) compared to healthy controls 21 (9-41) ppb). In contrast, nNO was comparable between groups (1368 <it>versus </it>1380 in controls) but showed higher variability in subjects after laryngectomy.</p> <p>Conclusions</p> <p>Our data suggest that either bronchial NO production in patients who underwent laryngectomy is very low, possibly due to alterations of the mucosa or oxidant production/inflammation, or that substantial contributions to FE_NOarise from the larynx, pharynx and mouth, raising FE_NOdespite velum closure. The data fit to those indicating a substantial contribution to FE_NOby the mouth in healthy subjects. The broader range of nNO values found in subjects after laryngectomy may indicate chronic alteration or oligo-symptomatic inflammation of nasal mucosa, as frequently found after total laryngectomy.</p

MicroRNA-200c Attenuates the Tumor-Infiltrating Capacity of Macrophages

Macrophages constitute a major part of the tumor-infiltrating immune cells. Within the tumor microenvironment, they acquire an alternatively activated, tumor-supporting phenotype. Factors released by tumor cells are crucial for the recruitment of tumor-associated macrophages. In the present project, we aimed to understand the role of hsa-miR-200c-3p (miR-200c) in the interplay between tumor cells and macrophages. To this end, we employed a coculture system of MCF7 breast tumor cells and primary human macrophages and observed the transfer of miR-200c from apoptotic tumor cells to macrophages, which required intact CD36 receptor in macrophages. We further comprehensively determined miR-200c targets in macrophages by mRNA-sequencing and identified numerous migration-associated mRNAs to be downregulated by miR-200c. Consequently, miR-200c attenuated macrophage infiltration into 3-dimensional tumor spheroids. miR-200c-mediated reduction in infiltration further correlated with a miR-200c migration signature comprised of the four miR-200c-repressed, predicted targets PPM1F, RAB11FIB2, RDX, and MSN

Routine application of SFC-MS in doping control: Analysis of 3 × 1000 urine samples using three different SFC-MS instruments

Supercritical fluid chromatography-mass spectrometry (SFC-MS) has proved to be a beneficial tool for sample analysis for a wide variety of compounds and, as such, has recently gained the attention of the anti-doping community. We have tested the applicability of SFC-MS for routine doping control analysing approximately 3 × 1000 identical anti-doping samples utilising SFC-MS instruments from three different vendors: Agilent Technologies, Waters Corporation and Shimadzu Corporation. A ‘dilute and inject’ approach either without or after hydrolysis of glucuronide metabolites was applied. Most of the compounds included in our study demonstrated excellent chromatography, whereas some showed co-elution with endogenous interferences requiring MS discrimination. Retention times typically were very stable within batches (%CV ≤ 0.5%), although this appeared to be analyte and column dependent. Chromatographic peak shape was good (symmetrical) and stable over the period of the testing without any change of column. Our results suggest that SFC-MS is a sensitive, reproducible and robust analytical tool ready to be used in anti-doping laboratories alongside the currently applied techniques such as gas and liquid chromatography coupled to mass spectrometry. Even if instruments are designed slightly differently, all three setups demonstrated their fitness for the purpose in anti-doping testing

An experimental study of a virtual reality counselling paradigm using embodied self-dialogue

When faced with a personal problem people typically give better advice to others than to themselves. A previous study showed how it is possible to enact internal dialogue in virtual reality (VR) through participants alternately occupying two different virtual bodies - one representing themselves and the other Sigmund Freud. They could maintain a self-conversation by explaining their problem to the virtual Freud and then from the embodied perspective of Freud see and hear the explanation by their virtual doppelganger, and then give some advice. Alternating between the two bodies they could maintain a self-dialogue, as if between two different people. Here we show that the process of alternating between their own and the Freud body is important for successful psychological outcomes. An experiment was carried out with 58 people, 29 in the body swapping Self-Conversation condition and 29 in a condition where they only spoke to a Scripted Freud character. The results showed that the Self-Conversation method results in a greater perception of change and help compared to the Scripted. We compare this method with the distancing paradigm where participants imagine resolving a problem from a first or third person perspective. We consider the method as a possible strategy for self-counselling

Anaerobic Metazoans: No longer an oxymoron

The sediments of a deep-sea hypersaline and sulfidic Mediterranean basin have yielded an unexpected discovery, the first multicellular animals living entirely without oxygen. Reported by Danovaro et al. in BMC Biology, these three new species of Loricifera add a new and remarkable dimension to anoxic ecosystems previously thought to support only unicellular life

Dissemination of patient blood management practices in Swiss intensive care units: a cross-sectional survey

BACKGROUND Patient blood management (PBM) promotes the routine detection and treatment of anaemia before surgery, optimising the management of bleeding disorders, thus minimising iatrogenic blood loss and pre-empting allogeneic blood utilisation. PBM programmes have expanded from the elective surgical setting to nonsurgical patients, including those in intensive care units (ICUs), but their dissemination in a whole country is unknown. METHODS We performed a cross-sectional, anonymous survey (10 October 2018 to 13 March 2019) of all ordinary medical members of the Swiss Society of Intensive Care Medicine and the registered ICU nurses from the 77 certified adult Swiss ICUs. We analysed PBM-related interventions adopted in Swiss ICUs and related them to the spread of PBM in Swiss hospitals. We explored blood test ordering policies, blood-sparing strategies and red blood cell-related transfusion practices in ICUs. RESULTS A total of 115 medical doctors and 624 nurses (response rates 27% and 30%, respectively) completed the surveys. Hospitals had implemented a PBM programme according to 42% of physicians, more commonly in Switzerland's German-speaking regions (Odds Ratio [OR] 3.39, 95% confidence interval [CI] 1.23-9.35; p = 0.018) and in hospitals with more than 500 beds (OR 3.91, 95% CI 1.48-10.4; p = 0.006). The PBM programmes targeted the detection and correction of anaemia before surgery (79%), minimising perioperative blood loss (94%) and optimising anaemia tolerance (98%). Laboratory tests were ordered in 70.4% by the intensivist during morning rounds; the nurses performed arterial blood gas analyses autonomously in 48.4%. Blood-sparing techniques were used by only 42.1% of nurses (263 of 624, missing: 6) and 47.0% of physicians (54 of 115). Approximately 60% of respondents used an ICU-specific transfusion guideline. The reported haemoglobin threshold for the nonbleeding ICU population was 70 g/l and, therefore, was at the lower limit of current guidelines. CONCLUSIONS Based on this survey, the estimated proportion of the intensivists working in hospitals with a PBM initiative is 42%, with significant variability between regions and hospitals of various sizes. The risk of iatrogenic anaemia is relevant due to liberal blood sample collection practices and the underuse of blood-sparing techniques. The reported transfusion threshold suggests excellent adherence to current international ICU-specific transfusion guidelines

Differential association of GABAB receptors with their effector ion channels in Purkinje cells

Metabotropic GABAB receptors mediate slow inhibitory effects presynaptically and postsynaptically through the modulation of different effector signalling pathways. Here, we analysed the distribution of GABAB receptors using highly sensitive SDS-digested freeze-fracture replica labelling in mouse cerebellar Purkinje cells. Immunoreactivity for GABAB1 was observed on presynaptic and, more abundantly, on postsynaptic compartments, showing both scattered and clustered distribution patterns. Quantitative analysis of immunoparticles revealed a somato-dendritic gradient, with the density of immunoparticles increasing 26-fold from somata to dendritic spines. To understand the spatial relationship of GABAB receptors with two key effector ion channels, the G protein-gated inwardly rectifying K+ (GIRK/Kir3) channel and the voltage-dependent Ca2+ channel, biochemical and immunohistochemical approaches were performed. Co-immunoprecipitation analysis demonstrated that GABAB receptors co-assembled with GIRK and CaV2.1 channels in the cerebellum. Using double-labelling immunoelectron microscopic techniques, co-clustering between GABAB1 and GIRK2 was detected in dendritic spines, whereas they were mainly segregated in the dendritic shafts. In contrast, co-clustering of GABAB1 and CaV2.1 was detected in dendritic shafts but not spines. Presynaptically, although no significant co-clustering of GABAB1 and GIRK2 or CaV2.1 channels was detected, inter-cluster distance for GABAB1 and GIRK2 was significantly smaller in the active zone than in the dendritic shafts, and that for GABAB1 and CaV2.1 was significantly smaller in the active zone than in the dendritic shafts and spines. Thus, GABAB receptors are associated with GIRK and CaV2.1 channels in different subcellular compartments. These data provide a better framework for understanding the different roles played by GABAB receptors and their effector ion channels in the cerebellar network

The Sun at millimeter wavelengths I. Introduction to ALMA Band 3 observations

We present an initial study of one of the first ALMA Band 3 observations of the Sun with the aim to characterise the diagnostic potential of brightness temperatures measured with ALMA on the Sun. The observation covers 48min at a cadence of 2s targeting a Quiet Sun region at disk-centre. Corresponding time series of brightness temperature maps are constructed with the first version of the Solar ALMA Pipeline (SoAP) and compared to simultaneous SDO observations. The angular resolution of the observations is set by the synthesized beam (1.4x2.1as). The ALMA maps exhibit network patches, internetwork regions and also elongated thin features that are connected to large-scale magnetic loops as confirmed by a comparison with SDO maps. The ALMA Band 3 maps correlate best with the SDO/AIA 171, 131 and 304 channels in that they exhibit network features and, although very weak in the ALMA maps, imprints of large-scale loops. A group of compact magnetic loops is very clearly visible in ALMA Band 3. The brightness temperatures in the loop tops reach values of about 8000-9000K and in extreme moments up to 10 000K. ALMA Band 3 interferometric observations from early observing cycles already reveal temperature differences in the solar chromosphere. The weak imprint of magnetic loops and the correlation with the 171, 131, and 304 SDO channels suggests though that the radiation mapped in ALMA Band 3 might have contributions from a larger range of atmospheric heights than previously assumed but the exact formation height of Band 3 needs to be investigated in more detail. The absolute brightness temperature scale as set by Total Power measurements remains less certain and must be improved in the future. Despite these complications and the limited angular resolution, ALMA Band 3 observations have large potential for quantitative studies of the small-scale structure and dynamics of the solar chromosphere.Comment: 14 pages, 9 figures; accepted for publication in A&