A new strategy for faster urinary biomarkers identification by Nano-LC-MALDI-TOF/TOF mass spectrometry

<p>Abstract</p> <p>Background</p> <p>LC-MALDI-TOF/TOF analysis is a potent tool in biomarkers discovery characterized by its high sensitivity and high throughput capacity. However, methods based on MALDI-TOF/TOF for biomarkers discovery still need optimization, in particular to reduce analysis time and to evaluate their reproducibility for peak intensities measurement. The aims of this methodological study were: (i) to optimize and critically evaluate each step of urine biomarker discovery method based on Nano-LC coupled off-line to MALDI-TOF/TOF, taking full advantage of the dual decoupling between Nano-LC, MS and MS/MS to reduce the overall analysis time; (ii) to evaluate the quantitative performance and reproducibility of nano-LC-MALDI analysis in biomarker discovery; and (iii) to evaluate the robustness of biomarkers selection.</p> <p>Results</p> <p>A pool of urine sample spiked at increasing concentrations with a mixture of standard peptides was used as a specimen for biological samples with or without biomarkers. Extraction and nano-LC-MS variabilities were estimated by analyzing in triplicates and hexaplicates, respectively. The stability of chromatographic fractions immobilised with MALDI matrix on MALDI plates was evaluated by successive MS acquisitions after different storage times at different temperatures.</p> <p>Low coefficient of variation (CV%: 10–22%) and high correlation (R²> 0.96) values were obtained for the quantification of the spiked peptides, allowing quantification of these peptides in the low fentomole range, correct group discrimination and selection of "specific" markers using principal component analysis. Excellent peptide integrity and stable signal intensity were found when MALDI plates were stored for periods of up to 2 months at +4°C. This allowed storage of MALDI plates between LC separation and MS acquisition (first decoupling), and between MS and MSMS acquisitions while the selection of inter-group discriminative ions is done (second decoupling). Finally the recording of MSMS spectra to obtain structural information was focused only on discriminative ions in order to minimize analysis time.</p> <p>Conclusion</p> <p>Contrary to other classical approaches with direct online coupling of chromatographic separation and on the flight MS and/or MSMS data acquisition for all detected analytes, our dual decoupling strategy allowed us to focus on the most discriminative analytes, giving us more time to acquire more replicates of the same urine samples thus increasing detection sensitivity and mass precision.</p

Heat-acclimatization and pre-cooling: a further boost for endurance performance?

© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd To determine if pre-cooling (PC) following heat-acclimatization (HA) can further improve self-paced endurance performance in the heat, 13 male triathletes performed two 20-km cycling time-trials (TT) at 35 °C, 50% relative humidity, before and after an 8-day training camp, each time with (PC) or without (control) ice vest PC. Pacing strategies, physiological and perceptual responses were assessed during each TT. PC and HA induced moderate (+10 ± 18 W; effect size [ES] 4.4 ± 4.6%) and very large (+28 ± 19 W; ES 11.7 ± 4.1%) increases in power output (PO), respectively. The overall PC effect became unclear after HA (+4 ± 14 W; ES 1.4 ± 3.0%). However, pacing analysis revealed that PC remained transiently beneficial post-HA, i.e., during the first half of the TT. Both HA and PC pre-HA were characterized by an enhanced PO without increased cardio-thermoregulatory or perceptual disturbances, while post-HA PC only improved thermal comfort. PC improved 20-km TT performance in unacclimatized athletes, but an 8-day HA period attenuated the magnitude of this effect. The respective converging physiological responses to HA and PC may explain the blunting of PC effectiveness. However, perceptual benefits from PC can still account for the small alterations to pacing noted post-HA

The 2mrad crossing angle scheme for the international linear collider

http://cern.ch/AccelConf/e08/papers/mopp005.pdfInternational audienceThe present baseline configuration of the ILC has a 14 mrad crossing angle between the beams at the interaction point. This allows easier extraction of the beams after col- lisions, but imposes on the other hand more constraints on the control of the beams prior to colliding them. More- over, some limitations to physics capabilities arise, in par- ticular because of the degraded very forward electromag- netic detector hermeticity and because calibration proce- dures for (gaseous) tracking detectors become more com- plex. To mitigate these problems, alternative configurations with very small crossing angles are studied. A new version of the 2 mrad layout was designed last year, based on sim- pler concepts and assumptions. The emphasis of this new scheme was to satisfy specifications with as few and feasi- ble magnets as possible, in order to reduce costs

Tacrolimus Population Pharmacokinetic-Pharmacogenetic Analysis and Bayesian Estimation in Renal Transplant Recipients

Objectives: The aims of this study were (i) to investigate the population pharmacokinetics of tacrolimus in renal transplant recipients, including the influence of biological and pharmacogenetic covariates; and (ii) to develop a Bayesian estimator able to reliably estimate the individual pharmacokinetic parameters and inter-dose area under the blood concentration-time curve (AUC) from 0 to 12 hours (AUC12) in renal transplant patients. Methods: Full pharmacokinetic profiles were obtained from 32 renal transplant patients at weeks 1 and 2, and at months 1, 3 and 6 post-transplantation. The population pharmacokinetic analysis was performed using the nonlinear mixed-effect modelling software NONMEM® version VI. Patients’ genotypes were characterized by allelic discrimination for PXR −25385C&gt;T genes. Results: Tacrolimus pharmacokinetics were well described by a two-compartment model combined with an Erlang distribution to describe the absorption phase, with low additive and proportional residual errors of 1.6 ng/mL and 9%, respectively. Both the haematocrit and PXR −25385C&gt;T single nucleotide polymorphism (SNP) were identified as significant covariates for apparent oral clearance (CL/F) of tacrolimus, which allowed improvement of prediction accuracy. Specifically, CL/F decreased gradually with the number of mutated alleles for the PXR −25385C&gt;T SNP and was inversely proportional to the haematocrit value. However, clinical criteria of relevance, mainly the decrease in interindividual variability and residual error, led us to retain only the haematocrit in the final model. Maximum a posteriori Bayesian forecasting allowed accurate prediction of the tacrolimus AUC12 using only three sampling times (at 0 hour [predose] and at 1 and 3 hours postdose) in addition to the haematocrit value, with a nonsignificant mean AUC bias of 2% and good precision (relative mean square error = 11%). Conclusion: Population pharmacokinetic analysis of tacrolimus in renal transplant recipients showed a significant influence of the haematocrit on its CL/F and led to the development of a Bayesian estimator compatible with clinical practice and able to accurately predict tacrolimus individual pharmacokinetic parameters and the AUC12

An artificial intelligence generated automated algorithm to measure total kidney volume in ADPKD

Introduction Accurate tools to inform individual prognosis in patients with autosomal dominant polycystic kidney disease (ADPKD) are lacking. Here, we report an artificial intelligence (AI) generated method for routinely measuring total kidney volume (TKV). Methods An ensemble U-net algorithm was created using the nnUNet approach. The training and internal cross-validation cohort consisted of all 1.5T MRI data acquired using 5 different MRI scanners (454 kidneys, 227 scans) in the CYSTic consortium which was first manually segmented by a single human operator. As an independent validation cohort, we utilised 48 sequential clinical MRI scans with reference results of manual segmentation acquired by 6 individual analysts at a single centre. The tool was then implemented for clinical use and its performance analysed. Results The training / internal validation cohort was younger (mean age 44.0 vs 51.5 years) and the female-male ratio higher (1.2 v 0.94) compared to the clinical validation cohort. The majority of CYSTic patients had PKD1 mutations (79%) and typical disease (Mayo Imaging Class 1, 86%). The median DICE score on the clinical validation dataset between the algorithm and human analysts was 0.96 for left and right kidneys with a median TKV error of -1.8%. The time taken to manually segment kidneys in the CYSTic dataset was 56 (±28) min whereas manual corrections of the algorithm output took 8.5 (±9.2) min per scan. Conclusions Our AI-based algorithm demonstrates performance comparable to manual segmentation. Its rapidity and precision in real-world clinical cases demonstrate its suitability for clinical application

Clinical spectrum, prognosis and estimated prevalence of DNAJB11-kidney disease

Monoallelic mutations of DNAJB11 were recently described in seven pedigrees with atypical clinical presentations of autosomal dominant polycystic kidney disease. DNAJB11 encodes one of the main cofactors of the endoplasmic reticulum chaperon BiP, a heat-shock protein required for efficient protein folding and trafficking. Here we conducted an international collaborative study to better characterize the DNAJB11-associated phenotype. Thirteen different loss-of-function variants were identified in 20 new pedigrees (54 affected individuals) by targeted next-generation sequencing, whole-exome sequencing or whole-genome sequencing. Amongst the 77 patients (27 pedigrees) now in total reported, 32 reached end stage kidney disease (range, 55-89 years, median age 75); without a significant difference between males and females. While a majority of patients presented with non-enlarged polycystic kidneys, renal cysts were inconsistently identified in patients under age 45. Vascular phenotypes, including intracranial aneurysms, dilatation of the thoracic aorta and dissection of a carotid artery were present in four pedigrees. We accessed Genomics England 100,000 genomes project data, and identified pathogenic variants of DNAJB11 in nine of 3934 probands with various kidney and urinary tract disorders. The clinical diagnosis was cystic kidney disease for eight probands and nephrocalcinosis for one proband. No additional pathogenic variants likely explaining the kidney disease were identified. Using the publicly available GnomAD database, DNAJB11 genetic prevalence was calculated at 0.85/10.000 individuals. Thus, establishing a precise diagnosis in atypical cystic or interstitial kidney disease is crucial, with important implications in terms of follow-up, genetic counseling, prognostic evaluation, therapeutic management, and for selection of living kidney donors

How to look next? A data-driven approach for scanpath prediction

By and large, current visual attention models mostly rely, when considering static stimuli, on the following procedure. Given an image, a saliency map is computed, which, in turn, might serve the purpose of predicting a sequence of gaze shifts, namely a scanpath instantiating the dynamics of visual attention deployment. The temporal pattern of attention unfolding is thus confined to the scanpath generation stage, whilst salience is conceived as a static map, at best conflating a number of factors (bottom-up information, top-down, spatial biases, etc.). In this note we propose a novel sequential scheme that consists of a three-stage processing relying on a center-bias model, a context/layout model, and an object-based model, respectively. Each stage contributes, at different times, to the sequential sampling of the final scanpath. We compare the method against classic scanpath generation that exploits state-of-the-art static saliency model. Results show that accounting for the structure of the temporal unfolding leads to gaze dynamics close to human gaze behaviour