出版者・発行元：Cold Spring Harbor Laboratory  

Abstract

Antibodies are extensively used in biomedical research, clinical fields, and disease treatment. However, to enhance the reproducibility and reliability of antibody-based experiments, it is crucial to have a detailed understanding of the antibody’s target specificity and epitope. In this study, we developed a high-throughput and precise epitope analysis method, DECODE (Decoding Epitope Composition by Optimized-mRNA-display, Data analysis, and Expression sequencing). This method allowed identifying patterns of epitopes recognized by monoclonal or polyclonal antibodies at single amino acid resolution and predicted cross-reactivity against the entire protein database. By applying the obtained epitope information, it has become possible to develop a new 3D immunostaining method that increases the penetration of antibodies deep into tissues. Furthermore, to demonstrated the applicability of DECODE to more complex blood antibodies, we performed epitope analysis using serum antibodies from mice with experimental autoimmune encephalomyelitis (EAE). As a result, we were able to successfully identify an epitope that matched the sequence of the peptide inducing the disease model without relying on existing antigen information. These results demonstrate that DECODE can provide high-quality epitope information, improve the reproducibility of antibody-dependent experiments, diagnostics and therapeutics, and contribute to discover pathogenic epitopes from antibodies in the blood.

DOI： 10.1101/2024.06.13.598778

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担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/s41592-024-02307-5

PubMed

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担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Elsevier BV  

DOI： 10.1016/j.ibneur.2023.08.873

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掲載種別：研究論文（学術雑誌）   出版者・発行元：Public Library of Science (PLoS)  

The reduced sleep duration previously observed in Camk2b knockout mice revealed a role for Ca²⁺/calmodulin-dependent protein kinase II (CaMKII)β as a sleep-promoting kinase. However, the underlying mechanism by which CaMKIIβ supports sleep regulation is largely unknown. Here, we demonstrate that activation or inhibition of CaMKIIβ can increase or decrease sleep duration in mice by almost 2-fold, supporting the role of CaMKIIβ as a core sleep regulator in mammals. Importantly, we show that this sleep regulation depends on the kinase activity of CaMKIIβ. A CaMKIIβ mutant mimicking the constitutive-active (auto)phosphorylation state promotes the transition from awake state to sleep state, while mutants mimicking subsequent multisite (auto)phosphorylation states suppress the transition from sleep state to awake state. These results suggest that the phosphorylation states of CaMKIIβ differently control sleep induction and maintenance processes, leading us to propose a “phosphorylation hypothesis of sleep” for the molecular control of sleep in mammals.

DOI： 10.1371/journal.pbio.3001813

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

BACKGROUND & AIMS: Tissue-clearing and three-dimensional (3D) imaging techniques aid clinical histopathological evaluation; however, further methodological developments are required prior to use in clinical practice. METHODS: We sought to develop a novel fluorescence staining method based on the classical PAS stain. We further attempted to develop a 3D imaging system based on this staining method and evaluated whether the system can be employed for quantitative 3D pathological evaluation and deep learning-based automatic diagnosis of inflammatory bowel diseases. RESULTS: We successfully developed a novel periodic acid-FAM hydrazide (PAFhy) staining method for 3D imaging when combined with a tissue-clearing technique (PAFhy-3D). This strategy enabled clear and detailed imaging of the 3D architectures of crypts in human colorectal mucosa. PAFhy-3D imaging also revealed abnormal architectural changes in crypts in ulcerative colitis tissues and identified the distributions of neutrophils in cryptitis and crypt abscesses. PAFhy-3D revealed novel pathological findings including "spiral staircase-like crypts" specific to inflammatory bowel diseases. Quantitative analysis of crypts based on 3D morphological changes enabled differential diagnosis of ulcerative colitis, Crohn's disease, and non-inflammatory bowel disease; such discrimination could not be achieved by pathologists. Furthermore, a deep learning-based system employing PAFhy-3D images was used to distinguish these diseases The accuracies were excellent (macro-average AUC = 0.94; F1 scores = 0.875 for ulcerative colitis, 0.717 for Crohn's disease, and 0.819 for non-inflammatory bowel disease). CONCLUSIONS: PAFhy staining and PAFhy-3D imaging are promising approaches for next-generation experimental and clinical histopathology.

DOI： 10.1016/j.jcmgh.2022.07.001

PubMed

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出版者・発行元：Cold Spring Harbor Laboratory  

ABSTRACT

The reduced sleep duration observed in Camk2a and Camk2b knockout mice revealed the role of Ca²⁺/calmodulin-dependent protein kinase II (CaMKII)α/CAMKIIβ as sleep-promoting kinases and lead to the phosphorylation hypothesis of sleep. However, the underlying mechanism of sleep regulation by kinases and protein phosphorylation is largely unknown. Here, we demonstrate that the phosphorylation states of CaMKIIβ regulates sleep duration and sleep needs. Importantly, the activation or inhibition of CaMKIIβ can increase or decrease sleep duration by almost two-fold, supporting the role of CaMKIIβ as a core sleep regulator in mammals. This sleep regulation depends on the kinase activity of CaMKIIβ in excitatory neurons. Furthermore, CaMKIIβ mutants mimicking different phosphorylation states can regulate various sleep steps including sleep induction, sleep maintenance, and sleep cancelation. Key CaMKIIβ residues responsible for the mode switch undergo ordered (auto-)phosphorylation. We thus propose that ordered multi-site phosphorylation of CaMKIIβ underlies multi-step sleep regulation in mammals.

DOI： 10.1101/2021.10.11.463945

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担当区分：筆頭著者   記述言語：英語  

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease; transactivation response DNA-binding protein of 43 kDa (TDP-43) and iron accumulation are supposed to play a crucial role in the pathomechanism of the disease. Here, we report an unusual case of a patient with ALS who presented with speech apraxia as an initial symptom and upper motor neuron deficiencies. In the early clinical stages, single-photon emission computed tomography visualized focal hypoperfusion of the right frontal operculum, and magnetic resonance imaging identified a hypointense area along the frontal lobe on T2-weighted images. Neuropathological examination revealed that neuronophagia of Betz cells, gliosis, appearance of phosphorylated TDP-43 (p-TDP-43)-positive glial and neuronal inclusions, and prominent iron accumulation were frequently visible in the precentral gyrus. TDP-43 pathology and focal iron accumulation were also visible in the frontal operculum, but only a mild neuronal loss and a few p-TDP-43-positive neuronal and glial inclusions were found in the hypoglossal nucleus of the medulla oblongata and anterior horn of the spinal cord. Immunoblot analysis revealed an atypical band pattern for ALS. In our case, abnormal TDP-43 and iron accumulation might possibly have caused neurodegeneration of the frontal operculum, in tandem or independently; it might then have spread into the primary motor area. Our results suggest a causative association between TDP-43 and iron accumulation in the pathomechanisms of ALS presenting with upper motor neuron signs.

DOI： 10.1111/neup.12763

PubMed

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担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1038/s41596-019-0240-9

Web of Science

PubMed

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その他リンク： http://www.nature.com/articles/s41596-019-0240-9

担当区分：筆頭著者   記述言語：日本語   掲載種別：研究論文（学術雑誌）  

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開催年月日： 2025年7月

記述言語：英語   会議種別：口頭発表（招待・特別）  

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開催年月日： 2023年12月

記述言語：英語   会議種別：ポスター発表  

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開催年月日： 2023年11月

記述言語：英語   会議種別：ポスター発表  

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開催年月日： 2023年9月

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開催年月日： 2023年1月

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開催年月日： 2022年11月

記述言語：英語   会議種別：ポスター発表  

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開催年月日： 2022年6月 - 2022年7月

記述言語：英語   会議種別：口頭発表（一般）  

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開催年月日： 2022年5月

記述言語：日本語   会議種別：シンポジウム・ワークショップ パネル（指名）  

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開催年月日： 2020年12月

記述言語：日本語   会議種別：ポスター発表  

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記述言語：日本語   会議種別：シンポジウム・ワークショップ パネル（指名）  

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記述言語：英語   会議種別：シンポジウム・ワークショップ パネル（指名）  

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会議種別：口頭発表（招待・特別）  

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記述言語：英語   会議種別：ポスター発表  

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記述言語：英語   会議種別：ポスター発表  

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記述言語：日本語   会議種別：公開講演，セミナー，チュートリアル，講習，講義等  

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記述言語：英語   会議種別：公開講演，セミナー，チュートリアル，講習，講義等  

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記述言語：英語   会議種別：公開講演，セミナー，チュートリアル，講習，講義等  

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記述言語：英語   会議種別：ポスター発表  

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記述言語：英語   会議種別：口頭発表（一般）  

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記述言語：英語   会議種別：口頭発表（一般）  

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記述言語：英語   会議種別：ポスター発表  

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記述言語：日本語   会議種別：口頭発表（招待・特別）  

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記述言語：英語   会議種別：ポスター発表  

ポスター発表

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記述言語：英語   会議種別：口頭発表（招待・特別）  

selected oral speaker

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記述言語：日本語   会議種別：口頭発表（一般）  

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