組織因子惹起-トロンボモジュリン添加凝固波形解析を用いたプロテインC経路異常を伴う血栓性素因のスクリーニング法

Objectives: Absolute or relative protein (P)C pathway abnormalities (PC deficiency, PS deficiency, antiphospholipid syndrome (APS), factor (F)V-abnormality, and high FVIII level) cause thrombophilia. Although screening assays for these thrombophilias are available, one utilizing clot waveform analysis (CWA) remains unknown. We aimed to establish a CWA-based screening assay to distinguish PC pathway abnormality-related thrombophilia. Methods: Samples were reacted with tissue factor (TF)/phospholipids and recombinant thrombomodulin (rTM; optimal 20 nM), followed by CWA measurement. The peak ratio (with/without rTM) of the first derivative curve of clot waveform was calculated. Results: The peak ratio in healthy plasmas (n = 35) was 0.36 ± 0.13; hence, the cutoff value was set to 0.49. The peak ratios in plasmas with PC deficiency, PS deficiency, high-FVIII (spiked 300 IU/dl), and APS were higher than the cutoff values (0.79/0.97/0.50/0.93, respectively). PC-deficient plasma or PS-deficient plasma mixed with normal plasma (25%/50%/75%/100% PC or PS level) showed dose-dependent decreases in the peak ratios (PC deficient: 0.85/0.64/0.44/0.28; PS deficient: 0.69/0.53/0.40/0.25), suggesting that the peak ratio at ≤50% of PC or PS level exceeded the cutoff value. The peak ratio in FV deficiency with FV ≤25% was higher than the cutoff value. FV-deficient plasma spiked with 40 IU/dl rFV-R506Q (FVLeiden ) or rFV-W1920R (FVNara ) showed >90% peak ratios. Conclusions: rTM-mediated TF-triggered CWA might be useful for screening PC pathway abnormality-related thrombophilia.博士（医学）・乙第1527号・令和4年9月28日© 2022 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.This is the peer reviewed version of the following article: [https://onlinelibrary.wiley.com/doi/10.1111/ejh.13777], which has been published in final form at [https://doi.org/10.1111/ejh.13777]. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions. This article may not be enhanced, enriched or otherwise transformed into a derivative work, without express permission from Wiley or by statutory rights under applicable legislation. Copyright notices must not be removed, obscured or modified. The article must be linked to Wiley’s version of record on Wiley Online Library and any embedding, framing or otherwise making available the article or pages thereof by third parties from platforms, services and websites other than Wiley Online Library must be prohibited.発行元が定める登録猶予期間終了の後、本文を登録予定（2023.07

慢性肺疾患の早産児におけるプロテインC経路 : 前向き研究

Background: Chronic lung disease (CLD) is a major neonatal pulmonary disorder associated with inflammation. Recent studies have shown that protein C anticoagulant pathways, such as those for protein C (PC), protein S (PS), and thrombomodulin (TM), could be useful indices for reflecting pulmonary injury. However, the involvement of these factors in preterm infants with very low birthweight (VLBW) who have developed CLD remains to be investigated. Here, we investigated whether PC pathway-related factors could predict the development of CLD in preterm infants with VLBW. Methods: We collected plasma samples from 26 preterm infants with VLBW (13 each from those with and without CLD) at the time of birth and measured TM, PC, and PS levels in their plasmas. We analyzed prospectively the relationship between these factors in infants with and without CLD. Results: There were significant differences in gestational age, birthweight, Apgar score (5 min), and duration of mechanical ventilation between the CLD and non-CLD groups. No significant differences in the PC and PS levels at birth were observed between the two groups, whereas the TM levels in the CLD group were significantly higher than those in the non-CLD group (P = 0.013). The TM levels correlated with gestational age and duration of mechanical ventilation. However, covariance analysis demonstrated that gestational age was significantly associated with TM levels, and consequently, development of CLD was not associated with TM level at birth. Conclusions: Thrombomodulin, PC, and PS levels at birth could not predict the development of CLD in preterm infants with VLBW.博士（医学）・甲第850号・令和4年9月28日© 2022 Japan Pediatric Society.This is the peer reviewed version of the following article: [https://onlinelibrary.wiley.com/doi/10.1111/ped.15221], which has been published in final form at [https://doi.org/10.1111/ped.15221]. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions. This article may not be enhanced, enriched or otherwise transformed into a derivative work, without express permission from Wiley or by statutory rights under applicable legislation. Copyright notices must not be removed, obscured or modified. The article must be linked to Wiley’s version of record on Wiley Online Library and any embedding, framing or otherwise making available the article or pages thereof by third parties from platforms, services and websites other than Wiley Online Library must be prohibited.発行元が定める登録猶予期間終了の後、本文を登録予定（2023.01

第VIII因子A1ドメインC末端酸性領域は、トロンビン惹起活性化とArg³⁷²開裂を制御する

Background: Factor VIII (FVIII) is activated by thrombin-catalyzed cleavage at three sites. Previous reports indicated that the A2 domain contained thrombin-interactive sites responsible for cleavage at Arg³⁷². We have also found that the A1 domain of FVIII bound to the anion-binding exosite I of thrombin. The present study focused, therefore, on thrombin interaction with A1 residues 337-372 containing clustered acidic and hirugen-like sequences. Aim: To identify specific thrombin-interactive site(s) within the A1 acidic region of FVIII. Methods and results: The synthetic peptide of residues 337-353 with sulfated Tyr³⁴⁶ (337-353S) significantly blocked thrombin-catalyzed FVIII activation and cleavage at Arg³⁷², while a corresponding peptide of residues 354-372 had no significant effect. Treatment with 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide to cross-link thrombin and 340-350S suggested that the 344-349 clustered acidic region was involved in thrombin interaction. Alanine-substituted FVIII mutants, Y346A and D347A/D348A/D349A, depressed thrombin-catalyzed activation and cleavage at Arg³⁷², with peak activation at ~ 50% and cleavage rates of ~ 10% to 20% compared to wild type (WT). The peak level of thrombin-catalyzed activation and the cleavage rate at Arg³⁷² using FVIII mutants with 337-346 residues substituted with hirugen-sequences (MKNNEEAEDY337-346GDFEEIPEEY) were ~ 1.5- and ~ 2.5-fold of WT, respectively. Surface plasmon resonance-based analysis demonstrated that the Kd for active-site modified thrombin interactions using Y346A and D347A/D348A/D349A mutants was ~ 3- to 6-fold higher than that of WT, and that the hirugen-hybrid mutant facilitated association kinetics ~ 1.8-fold of WT. Conclusion: Residues 346-349 with sulfated Tyr provided a thrombin-interactive site responsible for activation and cleavage at Arg³⁷². A hirugen-hybrid A1 mutant showed more efficient thrombin-catalyzed cleavage at Arg³⁷².博士（医学）・甲第788号・令和3年3月15日© 2020 International Society on Thrombosis and Haemostasis.This is the peer reviewed version of the following article: https://onlinelibrary.wiley.com/doi/10.1111/jth.15201, which has been published in final form at https://doi.org/10.1111/jth.15201. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions

ROTEMを用いた小児特発性ネフローゼ症候群患者の急性期における包括的全血凝固線溶能

Background: Venous thromboembolism is a rare, serious complication of idiopathic nephrotic syndrome (INS) in childhood. The mechanisms responsible for the hypercoagulable state in the acute phase of INS are poorly understood, however. This study aimed to assess overall coagulation and fibrinolytic function in pediatric patients with INS. Methods: Global coagulation and fibrinolysis were examined in whole blood samples from 22 children with initial onset INS (initial-group), 22 children with relapsed INS (relapse-group), and 15 control pediatric patients using rotational thromboelastometry (ROTEM®). In the initial-group, blood samples were obtained before (week 0) and 1-4 weeks after initiation of corticosteroid therapy. EXTEM and FIBTEM were used to assess coagulation and fibrinolysis, respectively. Clot time (CT), clot formation time (CFT), maximum clot firmness (MCF), and α-angle were determined as coagulation parameters, and lysis index at 30 and 60 min (LI30 and LI60, respectively) were assessed as fibrinolytic parameters. Results: CT was significantly shortened, and MCF and α-angle were significantly greater than controls at week 0 and week 1 both in the initial-group and the relapse-group. MCF correlated with serum albumin (r = 0.70, p < 0.001) and fibrinogen level (r = 0.68, p < 0.001). The fibrinolytic parameters (LI30 and LI60) in the initial-group were stable and higher than those in controls at all time points (p < 0.01). Conclusions: We have shown that the hypofibrinolytic defect did not improve with effective NS treatment at the early 4-week time-point. Additionally, a likely pre-thrombotic state was evident in the period before initial onset and 1 week after corticosteroid therapy in pediatric INS.博士（医学）・乙第1522号・令和4年3月15日© 2021. The Author(s), under exclusive licence to International Pediatric Nephrology Association.This version of the article has been accepted for publication, after peer review (when applicable) and is subject to Springer Nature’s AM terms of use, but is not the Version of Record and does not reflect post-acceptance improvements, or any corrections. The Version of Record is available online at: http://dx.doi.org/10.1007/s00467-021-05366-4.発行元が定める登録猶予期間終了の後、本文を登録予定（2023.01

Camera Calibration of the CTA-LST prototype

International audienceThe Cherenkov Telescope Array (CTA) is the next-generation gamma-ray observatory that is expected to reach one order of magnitude better sensitivity than that of current telescope arrays. The Large-Sized Telescopes (LSTs) have an essential role in extending the energy range down to 20 GeV. The prototype LST (LST-1) proposed for CTA was built in La Palma, the northern site of CTA, in 2018. LST-1 is currently in its commissioning phase and moving towards scientific observations. The LST-1 camera consists of 1855 photomultiplier tubes (PMTs) which are sensitive to Cherenkov light. PMT signals are recorded as waveforms sampled at 1 GHz rate with Domino Ring Sampler version 4 (DRS4) chips. Fast sampling is essential to achieve a low energy threshold by minimizing the integration of background light from the night sky. Absolute charge calibration can be performed by the so-called F-factor method, which allows calibration constants to be monitored even during observations. A calibration pipeline of the camera readout has been developed as part of the LST analysis chain. The pipeline performs DRS4 pedestal and timing corrections, as well as the extraction and calibration of charge and time of pulses for subsequent higher-level analysis. The performance of each calibration step is examined, and especially charge and time resolution of the camera readout are evaluated and compared to CTA requirements. We report on the current status of the calibration pipeline, including the performance of each step through to signal reconstruction, and the consistency with Monte Carlo simulations

Calibration and performance of the readout system based on switched capacitor arrays for the Large-Sized Telescope of the Cherenkov Telescope Array

International audienceThe Cherenkov Telescope Array1 (CTA) is the next-generation ground-based observatory for very-high-energy gamma rays. The CTA consists of three types of telescopes with different mirror areas to cover a wide energy range (20 GeV–300 TeV) with an order of magnitude higher sensitivity than the predecessors. Among those telescopes, the Large-Sized Telescope (LST) is designed to detect low-energy gamma rays between 20 GeV and a few TeV with a 23 m diameter mirror. To make the most of such a large light collection area (about 400 m2), the focal plane camera must detect as much reflected Cherenkov light as possible. We have developed each camera component to meet the CTA performance requirements for more than ten years and performed quality-control tests before installing the camera to the telescope.2, 3 The first LST (LST-1) was inaugurated in October 2018 in La Palma, Spain (Figure 1).4 After the inauguration, various calibration tests were performed to adjust hardware parameters and verify the camera performance. In parallel, we have been developing the analysis software to extract physical parameters from low-level data, taking into account some intrinsic characteristics of the switched capacitor arrays, Domino Ring Sampler version 4 (DRS4), used for sampling the waveform of a Cherenkov signal. In this contribution, we describe the hard- ware design of the LST camera in Section 2, a procedure for low-level calibration in Section 3, and the readout e of the LST camera after the hardware calibration with a dedicated analysis chain in Section 4