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Establishment of cell dispersion and isolation methods from animal tissues to understand the dynamics of minor cells

Research Project

Project/Area Number 16K14764
Research Category

Grant-in-Aid for Challenging Exploratory Research

Allocation TypeMulti-year Fund
Research Field Morphology/Structure
Research InstitutionKindai University (2017-2019)
University of Tsukuba (2016)

Principal Investigator

MATSUDA Manabu  近畿大学, 医学部, 准教授 (30282726)

Project Period (FY) 2016-04-01 – 2020-03-31
Project Status Completed (Fiscal Year 2019)
Budget Amount *help
¥3,640,000 (Direct Cost: ¥2,800,000、Indirect Cost: ¥840,000)
Fiscal Year 2018: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000)
Fiscal Year 2017: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Fiscal Year 2016: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
Keywords細胞分散 / プロテアーゼ / 消化酵素 / 酸性プロテアーゼ / 細胞培養 / ペプシン / 大腸菌発現系 / トランスクリプトーム解析 / RNA解析
Outline of Final Research Achievements

Analysis of the rather rare and minor cells are generally difficult, as they are apt to be masked and hidden by the others, i.e., the majority of the cells. Conventional methods of dissociation of the cells via digestion of live cells with collagenases are not suitable for the minority cells, because the nature of the cells may be altered easily by the environmental changes during the enzymatic digestion. Here, tissue cells are dissociated via digestion with an acidic enzyme after fixation of the tissues. I found acidic conditions that are practically affordably for fixation and digestion without any degradation of RNA. In addition, a fluorescent protein was verified to be available as a reporter molecule in the downstream experiments performed in the acidic conditions. However, degradation of cells by proteases was inevitable in the conditions strict enough to dissociate cells, at least so far. Further experiments are necessary to establish a gold standard method.

Academic Significance and Societal Importance of the Research Achievements

私生物のからだを構成する細胞は、同じように見えてもそれぞれに個性をもっている。個々の細胞を他の細胞から引き離して、発現するRNAを調べることで初期のがん細胞をはじめ細胞の振る舞い、いわば細胞の個性を知ることができる。本研究では、細胞の固定前後にRNA分解が起こらない酸性条件を特定した。その条件下ではたらく酸性プロテアーゼにより細胞の分散を試みたが細胞の損傷を抑えて機能する最適条件は今後の研究に委ねられる。

Report

(5 results)
  • 2019 Annual Research Report   Final Research Report ( PDF )
  • 2018 Research-status Report
  • 2017 Research-status Report
  • 2016 Research-status Report
  • Research Products

    (2 results)

All 2016

All Presentation (2 results) (of which Int'l Joint Research: 1 results)

  • [Presentation] Histamine as a topical regulatory factor in the lactating mammary gland2016

    • Author(s)
      Manabu Matsuda
    • Organizer
      The 22nd International Congress of Zoology
    • Place of Presentation
      沖縄コンベンションセンター(沖縄県宜野湾市)
    • Year and Date
      2016-11-14
    • Related Report
      2016 Research-status Report
    • Int'l Joint Research
  • [Presentation] 標識抗体によるタンパク質の検出における疎水性ドメインの影響2016

    • Author(s)
      松田 学
    • Organizer
      第47回乳腺・泌乳研究会
    • Place of Presentation
      東京大学(東京都文京区)
    • Year and Date
      2016-11-05
    • Related Report
      2016 Research-status Report

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Published: 2016-04-21   Modified: 2021-02-19  

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