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網膜グリア細胞の形質制御による糖尿病網膜症の予防と治療法の基礎研究

Research Project

Project/Area Number 14657448
Research Category

Grant-in-Aid for Exploratory Research

Allocation TypeSingle-year Grants
Research Field Ophthalmology
Research InstitutionSapporo Medical University

Principal Investigator

澤田 典均  札幌医科大学, 医学部, 教授 (30154149)

Project Period (FY) 2002 – 2003
Project Status Completed (Fiscal Year 2003)
Budget Amount *help
¥3,200,000 (Direct Cost: ¥3,200,000)
Fiscal Year 2003: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2002: ¥1,800,000 (Direct Cost: ¥1,800,000)
Keywords血液脳関門 / 血液網膜関門 / タイト結合 / MAPK / PKA / 糖尿病網膜症 / グリア細胞 / 血管透過性 / VEGF / GDNF / AGE
Research Abstract

血液網膜関門(BRB)や血液脳関門(BBB)の本体は、血管内皮細胞のタイト結合である。そこでこれらの細胞のタイト結合に発現しているclaudin-1とclaudin-5に着目して、テトラサイクリン依存性転写因子(tet-on system)を導入した血管内皮細胞株(RLE)を用いて、claudin-1とclaudin-5の発現調節によるタイト結合機能制御機構を検討した。
1.野生型またはMAPキナーゼ(MAPK)の標的部位と想定されるThr203をAlaに置換したclaudin-1遺伝子を導入し、それらの発現を随時誘導できるRLE細胞株を樹立した。この血管内皮細胞にclaudin-1の発現を誘導すると、野生型、変異型claudinの両者とも、機能的なタイト結合を再構成した。さらにThr203をAlaに置換したclaudin-1を発現誘導しMAPK阻害剤を作用させた実験から、MAPKがclaudin-1タンパクを標的分子としてタイト結合のバリア機能を増強すること、claudin-1配列内のThr^<203>がTriton-X100非可溶性claudin-1の量やタイト結合のバリア機能に関与していることが明らかになった。
2.野生型またはPKAの標的部位と想定されるThr207をAlaに置換したclaudin-5遺伝子を導入し、それらの発現を随時誘導できるRLE細胞株を樹立した。この細胞株にこれらのclaudin-5を発現させて解析したところ、野生型、変異型claudinの両者とも内皮細胞のタイト結合を再構成すること、claudin-5のThr207がPKAによるリン酸化部位であること、そのリン酸化が内皮細胞のバリア機能の制御に重要な役割を果たすことが明らかになった。

Report

(2 results)
  • 2003 Annual Research Report
  • 2002 Annual Research Report
  • Research Products

    (18 results)

All Other

All Publications (18 results)

  • [Publications] Sawada N et al.: "Tight junctions and human diseases."Med Electron Microsc.. 36. 147-156 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Kojima T et al.: "Regulation of the blood-biliary barrier : interaction between gap and tight junctions in hepatocytes."Med Electron Microsc.. 36. 157-164 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] 澤田典均 他: ""すきま"の細胞生物学 細胞間接着装置タイト結合とヒト疾患"札幌医学雑誌. 72. 1-7 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Chiba H et al.: "Hepatocyte nuclear factor (HNF)-4α triggers formation of functional tight junctions and establishment of polarized epithelial morpgology in F9 embryonal carcinoma cells."Exp Cell Res.. 286. 288-297 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Ishizaki M et al.: "Cyclic AMP induces phosphorylation of claudin-5-immunoprecipitates and expression of claudin-5 gene in blood-brain-barrier endothelial cells via protein kinase A-dependent and -independent pathways."Exp Cell Res.. 290. 275-288 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Kojima T et al.: "Role of the p38 MAP-kinase signaling pathway for cx32 and claudin-1 in the rat liver."Cell Communication and Adhesion.. 10. 1-7 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Tobioka H et al.: "Occludin expression decreases with the progression of human endome trialcarcinoma."Human Pathol.. 35. 159-164 (2004)

    • Related Report
      2003 Annual Research Report
  • [Publications] Tokunaga Y et al.: "Expression of occludin in human rectal carcinoid tumours as a possible marker for glandular differentiation."Histopathology.. 44. 247-250 (2004)

    • Related Report
      2003 Annual Research Report
  • [Publications] 小島隆 他: "ギャップ結合とタイト結合-形態学的関係から機能的関係へ-"顕微鏡学会誌. 印刷中.

    • Related Report
      2003 Annual Research Report
  • [Publications] Kojima T et al.: "Inhibition of MAP-kinase activity moderates changes in expression and function of Cx32 but not claudin-1 during DNA synthesis in primary cultures of rat hepatocytes."Med Electron Microsc.. in press.

    • Related Report
      2003 Annual Research Report
  • [Publications] Fujibe M et al.: "Thr203 of claudin-1, a putative phosphorylation site for MAP kinase, is required to promote the barrier function of tight junctions."Exp Cell Res.. in press.

    • Related Report
      2003 Annual Research Report
  • [Publications] Takakuwa Y et al.: "Bile canalicular barrier function and expression of tight junctional molecules in rat hepatocytes during common bile duct ligation"Cell Tissue Res.. 307・2. 181-189 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Tobioka H et al.: "Polarised distribution of carcinoembryonic antigen is associated with a tight junction in human colorectal adenocarcinomas"J Pathol.. 198・2. 207-212 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Kojima T et al.: "Cx32 formation and/or cx32-mediated intercellular communication induce expression and function of tight junctions in hepatocytic cell line"Exp Cell Res.. 276・1. 40-51 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Osanai M et al.: "Hepatocyte Nuclear Factor (HNF)-4α induces expression of endothelial Fas ligand (FasL) to prevent cancer cell transmigration : A novel defense mechanism of encothelium against cancer metastasis"Jpn J Cancer Res.. 93・5. 532-541 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Kamimura Y et al.: "Barrier function of microvessels and roles of glial cell line-derived neurotrophic factor in the rat testis"Med Electron Microsc.. 35・3. 139-145 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Chiba H et al.: "Hepatocyte nuclear factor (HNF)-4α triggers formation of functional tight junctions and establishment of polarized epithelial morpgology in F9 embryonal carcinoma cells"Exp Cell Res.. (in press).

    • Related Report
      2002 Annual Research Report
  • [Publications] 澤田典均: "病気と細胞内小器官"森道夫編著・文光堂. 236 (2002)

    • Related Report
      2002 Annual Research Report

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Published: 2002-04-01   Modified: 2016-04-21  

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