| Project/Area Number |
06557014
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| Research Category |
Grant-in-Aid for Developmental Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
Pathological medical chemistry
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| Research Institution | KYOTO UNIVERSITY (1995)
Okazaki National Research Institutes (1994) |
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Principal Investigator |
TSUKITA Shoichiro KYOTO UNIVERSITY,FACULTY OF MEDICINE PROFESSOR, 医学研究科, 教授 (50155347)
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| Co-Investigator(Kenkyū-buntansha) |
YONEMURA Shigenobu KYOTO UNIVERSITY,FACULTY OF MEDICINE LECUTURER, 医学研究科, 講師 (60192811)
ITOH Masahiko KYOUTO UNIVERSITY,FACULTY OF MEDICINE ASSISTANT, 医学研究科, 助手 (70270486)
永渕 昭良 岡崎国立共同研究機構, 生理学研究所, 助手 (80218023)
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| Project Period (FY) |
1994 – 1995
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| Project Status |
Completed (Fiscal Year 1995)
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| Budget Amount *help |
¥12,500,000 (Direct Cost: ¥12,500,000)
Fiscal Year 1995: ¥5,000,000 (Direct Cost: ¥5,000,000)
Fiscal Year 1994: ¥7,500,000 (Direct Cost: ¥7,500,000)
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| Keywords | blood-brain barrier / tight junction / occludin / claudin / knock-out / cell adhesion / phosphrylation / endothelial cell / アンチセンスDNA / 機能阻害抗体 / 上皮細胞 / Z0-1 / 脳血管バリアー |
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| Research Abstract |
To modulate the blood-brain barrier, i.e.tight junctions, we analyzed the functions of occludin. Occludin is the only known integral membrane protein of tight junctions (TJ), and is now believed to be directly involved in the barrier and fence functions of TJ.Occludin-deficient embryonic stem (ES) cells were generated by targeted disruption of both alleles of the occludin gene. When these cells were subjected to suspension culture, they aggregated to form simple, and then cystic embryoid bodies (EBs) with the same time course as EB formation from wild-type ES cells. Immunofluorescence microscopy and ultrathin section electron microscopy revealed that polarized epithelial (visceral endoderm-like) cells were differentiated to delineate EBs not only from wild-type but also from occludin-deficient ES cells. Freeze fracture analyzes indicated no significant differences in number or morphology of TJ strands between wild-type and occludin-deficient epithelial cells. Furthermore, ZO-1, a TJ-associated peripheral membrane protein, was still exclusively concentrated at TJ in occludin-deficient epithelial cells. In good agreement with these morphological observations, TJ in occludin-deficient epithelial cells functioned as a primary barrier to the diffusion of a low molecular mass tracer through the paracellular pathway. These findings indicate that there are as yet unidentified TJ integral membrane protein(s) which can form strand structures, recruit ZO-1, and function as a barrier without occludin.
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