Highly sensitive chiral analysis in capillary electrophoresis with large-volume sample stacking with an electroosmotic flow pump.

To improve the sensitivity in chiral analysis by capillary electrophoresis without loss of optical resolution, application of large-volume sample stacking with an electroosmotic flow pump (LVSEP) was investigated. Effects of the addition of cyclodextrin (CD) into a running solution on the LVSEP preconcentration was theoretically studied, where the preconcentration efficiency and effective separation length would be slightly increased if the effective electrophoretic velocity (v(ep, eff, BGS)) of the analytes was decreased by interacting with CD. In LVSEP-CD-modified capillary zone electrophoresis (CDCZE) and LVSEP-CD electrokinetic chromatography with reduced v(ep, eff, BGS), up to 1000-fold sensitivity increases were achieved with almost no loss of resolution. In LVSEP-CD-modified micellar electrokinetic chromatography of amino acids with increased v(ep, eff, BGS), a 1300-fold sensitivity increase was achieved without much loss of resolution, indicating the versatile applicability of LVSEP to many separation modes. An enantio-excess (EE) assay was also carried out in LVSEP-CDCZE, resulting in successful analyses of up to 99.6% EE. Finally, we analyzed ibuprofen in urine by desalting with a C(18) solid-phase extraction column. As a typical result, 250ppb ibuprofen was well concentrated and optically resolved with 84.0-86.6% recovery in LVSEP-CDCZE, indicating the applicability of LVSEP to real samples containing a large amount of unnecessary background salts

LGM permafrost distribution: how well can the latest PMIP multi-model ensembles perform reconstruction?

Here, global-scale frozen ground distribution from the Last Glacial Maximum (LGM) has been reconstructed using multi-model ensembles of global climate models, and then compared with evidence-based knowledge and earlier numerical results. Modeled soil temperatures, taken from Paleoclimate Modelling Intercomparison Project phase III (PMIP3) simulations, were used to diagnose the subsurface thermal regime and determine underlying frozen ground types for the present day (pre-industrial; 0 kya) and the LGM (21 kya). This direct method was then compared to an earlier indirect method, which categorizes underlying frozen ground type from surface air temperature, applying to both the PMIP2 (phase II) and PMIP3 products. Both direct and indirect diagnoses for 0 kya showed strong agreement with the present-day observation-based map. The soil temperature ensemble showed a higher diversity around the border between permafrost and seasonally frozen ground among the models, partly due to varying subsurface processes, implementation, and settings. The area of continuous permafrost estimated by the PMIP3 multi-model analysis through the direct (indirect) method was 26.0 (17.7) million km2 for LGM, in contrast to 15.1 (11.2) million km2 for the pre-industrial control, whereas seasonally frozen ground decreased from 34.5 (26.6) million km2 to 18.1 (16.0) million km2. These changes in area resulted mainly from a cooler climate at LGM, but from other factors as well, such as the presence of huge land ice sheets and the consequent expansion of total land area due to sea-level change. LGM permafrost boundaries modeled by the PMIP3 ensemble-improved over those of the PMIP2 due to higher spatial resolutions and improved climatology-also compared better to previous knowledge derived from geomorphological and geocryological evidence. Combinatorial applications of coupled climate models and detailed stand-alone physical-ecological models for the cold-region terrestrial, paleo-, and modern climates will advance our understanding of the functionality and variability of the frozen ground subsystem in the global eco-climate system

The whole blood transcriptional regulation landscape in 465 COVID-19 infected samples from Japan COVID-19 Task Force

「コロナ制圧タスクフォース」COVID-19患者由来の血液細胞における遺伝子発現の網羅的解析 --重症度に応じた遺伝子発現の変化には、ヒトゲノム配列の個人差が影響する--. 京都大学プレスリリース. 2022-08-23.Coronavirus disease 2019 (COVID-19) is a recently-emerged infectious disease that has caused millions of deaths, where comprehensive understanding of disease mechanisms is still unestablished. In particular, studies of gene expression dynamics and regulation landscape in COVID-19 infected individuals are limited. Here, we report on a thorough analysis of whole blood RNA-seq data from 465 genotyped samples from the Japan COVID-19 Task Force, including 359 severe and 106 non-severe COVID-19 cases. We discover 1169 putative causal expression quantitative trait loci (eQTLs) including 34 possible colocalizations with biobank fine-mapping results of hematopoietic traits in a Japanese population, 1549 putative causal splice QTLs (sQTLs; e.g. two independent sQTLs at TOR1AIP1), as well as biologically interpretable trans-eQTL examples (e.g., REST and STING1), all fine-mapped at single variant resolution. We perform differential gene expression analysis to elucidate 198 genes with increased expression in severe COVID-19 cases and enriched for innate immune-related functions. Finally, we evaluate the limited but non-zero effect of COVID-19 phenotype on eQTL discovery, and highlight the presence of COVID-19 severity-interaction eQTLs (ieQTLs; e.g., CLEC4C and MYBL2). Our study provides a comprehensive catalog of whole blood regulatory variants in Japanese, as well as a reference for transcriptional landscapes in response to COVID-19 infection

DOCK2 is involved in the host genetics and biology of severe COVID-19

「コロナ制圧タスクフォース」COVID-19疾患感受性遺伝子DOCK2の重症化機序を解明 --アジア最大のバイオレポジトリーでCOVID-19の治療標的を発見--. 京都大学プレスリリース. 2022-08-10.Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge. Here we conducted a genome-wide association study (GWAS) involving 2, 393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3, 289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target

The Changing Landscape for Stroke\ua0Prevention in AF: Findings From the GLORIA-AF Registry Phase 2

Background GLORIA-AF (Global Registry on Long-Term Oral Antithrombotic Treatment in Patients with Atrial Fibrillation) is a prospective, global registry program describing antithrombotic treatment patterns in patients with newly diagnosed nonvalvular atrial fibrillation at risk of stroke. Phase 2 began when dabigatran, the first non\u2013vitamin K antagonist oral anticoagulant (NOAC), became available. Objectives This study sought to describe phase 2 baseline data and compare these with the pre-NOAC era collected during phase 1. Methods During phase 2, 15,641 consenting patients were enrolled (November 2011 to December 2014); 15,092 were eligible. This pre-specified cross-sectional analysis describes eligible patients\u2019 baseline characteristics. Atrial fibrillation disease characteristics, medical outcomes, and concomitant diseases and medications were collected. Data were analyzed using descriptive statistics. Results Of the total patients, 45.5% were female; median age was 71 (interquartile range: 64, 78) years. Patients were from Europe (47.1%), North America (22.5%), Asia (20.3%), Latin America (6.0%), and the Middle East/Africa (4.0%). Most had high stroke risk (CHA2DS2-VASc [Congestive heart failure, Hypertension, Age  6575 years, Diabetes mellitus, previous Stroke, Vascular disease, Age 65 to 74 years, Sex category] score  652; 86.1%); 13.9% had moderate risk (CHA2DS2-VASc = 1). Overall, 79.9% received oral anticoagulants, of whom 47.6% received NOAC and 32.3% vitamin K antagonists (VKA); 12.1% received antiplatelet agents; 7.8% received no antithrombotic treatment. For comparison, the proportion of phase 1 patients (of N = 1,063 all eligible) prescribed VKA was 32.8%, acetylsalicylic acid 41.7%, and no therapy 20.2%. In Europe in phase 2, treatment with NOAC was more common than VKA (52.3% and 37.8%, respectively); 6.0% of patients received antiplatelet treatment; and 3.8% received no antithrombotic treatment. In North America, 52.1%, 26.2%, and 14.0% of patients received NOAC, VKA, and antiplatelet drugs, respectively; 7.5% received no antithrombotic treatment. NOAC use was less common in Asia (27.7%), where 27.5% of patients received VKA, 25.0% antiplatelet drugs, and 19.8% no antithrombotic treatment. Conclusions The baseline data from GLORIA-AF phase 2 demonstrate that in newly diagnosed nonvalvular atrial fibrillation patients, NOAC have been highly adopted into practice, becoming more frequently prescribed than VKA in Europe and North America. Worldwide, however, a large proportion of patients remain undertreated, particularly in Asia and North America. (Global Registry on Long-Term Oral Antithrombotic Treatment in Patients With Atrial Fibrillation [GLORIA-AF]; NCT01468701

キノウセイ マイクロチップ オ モチイタ コウセイノウ デンキ エイドウ ニ カンスル ケンキュウ

京都大学0048新制・課程博士博士(工学)甲第13788号工博第2892号新制||工||1427(附属図書館)UT51-2008-C704京都大学大学院工学研究科材料化学専攻(主査)教授 大塚 浩二, 教授 松原 誠二郎, 教授 小寺 秀俊学位規則第4条第1項該当Doctor of EngineeringKyoto UniversityDA

One-step preparation of amino-PEG modified poly(methyl methacrylate) microchips for electrophoretic separation of biomolecules

A simple method for a chemical surface modification of poly(methyl methacrylate) (PMMA) microchips with amino-poly(ethyleneglycol) (PEG-NH(2)) by nucleophilic addition-elimination reaction was developed to improve the separation efficiency and analytical reproducibility in a microchip electrophoresis (MCE) analysis of biomolecules such as proteins and enantiomers. In our procedure, the PEG chains were robustly immobilized only by introducing an aqueous solution of PEG-NH(2) into the PMMA microchannel. The electroosmotic mobilities on the modified chips remained almost constant during 35 days with 37 runs without any recoating. The PEG-NH(2) modified chip provided a fast, reproducible, efficient MCE separation of proteins with a wide variety of isoelectric points within 15s. Furthermore, the application of the modified chip to affinity electrophoresis using bovine serum albumin gave a good chiral separation of amino acids

Highly sensitive oligosaccharide analysis in capillary electrophoresis using large-volume sample stacking with an electroosmotic flow pump.

To obtain high sensitivity in capillary electrophoresis of oligosaccharide without reducing the high resolution with an easy experimental procedure, large-volume sample stacking with an electroosmotic flow pump (LVSEP) was investigated. As a fundamental study, effect of the conductivity of a sample solution in LVSEP was examined. It was revealed that LVSEP was successfully carried out even in using a sample solution with the ionic strength of 150μM and the conductivity ratio of 20, indicating a good applicability of LVSEP to the analysis of real samples containing salts. When glucose oligomer was analyzed as a model sample in LVSEP-capillary zone electrophoresis (CZE), all peaks were well resolved with decreasing only 5% of the peak-to-peak distance, which suggested 95% of the whole capillary could be used for the effective separation. In the analysis of maltoheptaose, a good calibration line with correlation coefficient of 0.9995 was obtained. The limit of detection was estimated as 2pM, which was 500-fold lower than that in the conventional CZE. N-linked glycans released from three glycoproteins, bovine ribonuclease B, bovine fetuin, and human α(1)-acid glycoprotein were also analyzed by LVSEP-CZE. By the sample purification with a gel filtration column, further sample dilution to reduce the sample conductivity for LVSEP was not needed. All glycan samples were well concentrated and separated with up to a 770-fold sensitivity increase. The run-to-run repeatabilities of the migration time, peak height, and peak area were good with relative standard deviations of 0.1-1.3%, 1.2-1.7%, and 2.8-4.9%, respectively