| 研究課題/領域番号 |
20K07458
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| 研究種目 |
基盤研究(C)
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| 配分区分 | 基金 |
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| 応募区分 | 一般 |
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| 審査区分 |
小区分49030:実験病理学関連
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| 研究機関 | 沖縄科学技術大学院大学 |
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研究代表者 |
TERENZIO Marco 沖縄科学技術大学院大学, 分子神経科学ユニット, 准教授 (60867513)
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| 研究期間 (年度) |
2020-04-01 – 2024-03-31
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| 研究課題ステータス |
完了 (2023年度)
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| 配分額 *注記 |
4,290千円 (直接経費: 3,300千円、間接経費: 990千円)
2022年度: 1,430千円 (直接経費: 1,100千円、間接経費: 330千円)
2021年度: 1,040千円 (直接経費: 800千円、間接経費: 240千円)
2020年度: 1,820千円 (直接経費: 1,400千円、間接経費: 420千円)
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| キーワード | ALS / biomarkers / IPSC / motor neurons / RNA sequencing / axonal translation / disease onset / protein translation / human IPSC / hIPSC / MNs |
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| 研究開始時の研究の概要 |
To overcome the limitations of murine models, we plan to directly derive motor neurons (MNs) from Amyotrophic Lateral Sclerosis (ALS) patient’s fibroblasts to assess changes in axonal local translation. Axonal degeneration is a primary pathological event determining the demise of MNs in ALS and defects of axonal translation contribute to the axonal pathology. We will investigate differences in axonal translation and test any abnormal protein expression as a possible new biomarker of disease and/or target of new therapies in biological fluids from well characterized cohorts of ALS patients.
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| 研究実績の概要 |
1) We acquired a library of 6 ALS and 4 healthy control human iPSC lines from Cedar Sinai Cell Bank. 2) We established a protocol of differentiation of motor neurons (MNs) from these human IPSC lines. 3) We characterized axonal degeneration, protein aggregation and neuronal cell death in our human MNs cultures. 4) We looked at the role of mitochondria and ATP production in correlation with local translation and phase separation for all the lines. 5) We collected cell pellets and culture medium from pre-symptomatic, early onset and late stage of disease MNs in culture. 6) We optimized a protocol for isolation of extracellular vesicles (EVs).
FINAL YEAR
7) We characterized the retrieved EVs biochemically and by electron microscopy. 8) We optimized the condition from proteomics analysis of MN culture media and extracted EVs. 9) We performed proteomics with the samples described in 3-4 to identify new ALS biomarkers. 10) We conducted a bioinformatic analysis to compare the different cell lines for each time point. 11) We are proceeding with candidate biomarker validation both in culture and from patient fluids.
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