| Project/Area Number |
03454117
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
General anatomy (including Histology/Embryology)
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| Research Institution | KYUSHU UNIVERSITY |
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Principal Investigator |
SHIBATA Yosaburo KYUSHU UNIVERSITY, FACULTY OF MEDICINE, PROFESSOR, 医学部, 教授 (90037482)
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| Co-Investigator(Kenkyū-buntansha) |
IIDA Hiroshi KYUSHU UNIVERSITY, FACULTY OF MEDICINE, ASSISTANT PROFESSOR, 医学部, 講師 (70150399)
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| Project Period (FY) |
1991 – 1992
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| Project Status |
Completed (Fiscal Year 1992)
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| Budget Amount *help |
¥5,700,000 (Direct Cost: ¥5,700,000)
Fiscal Year 1992: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 1991: ¥4,600,000 (Direct Cost: ¥4,600,000)
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| Keywords | Gap Junction / Connexin / Hepatocyte / Lens Fiber / Deep-Etch Replica / Immunolabelling / Membrane Proteins / Labelled Replica / 合成ペプチド / 免疫蛍光法 / 免疫ブロット / 肝小葉 / 心筋介在板 |
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| Research Abstract |
Molecular architectures of Liver Gap Junctions: The localizations of gap junction proteins, Cx32 and Cx26 were examined in rat and guinea pig livers by using site-specific antibodies against synthetic oligopeptides. Double labeling immunofluorescence microscopy revealed that in guinea pig liver both proteins spread throughout the liver lobules and seemed to localize together within the same gap junctional plaque. Quick-freeze, deep-etch immunoelectron microscopy showed that immunolabeling of isolated guinea pig liver gap junctional plaques with either Cx32 or Cx26 antiserum yielded complete and dense antibody decoration of the cytoplasmic surface of the plaques. We did not observe any focal or patchy clusters of the labeling in any plaques examined. Double labeling immuno electron microscopy confirmed that both Cx32 and Cx26 are colocalized in the same gap junction plaques. These results suggest that in the hepatocytes expressing both Cx32 and Cx26, both types of gap junction proteins are not segregated but intermix randomly within the same plaques.
Cytoplasmic Surface Structures of Bovine lens Fiber Gap Junctions: Bovine lens fibers studied by quick freeze-deep etch replica method revealed particulate substructures on cytoplasmic surfaces in situ. In isolated membranes, thin section electron microscopy showed that 16-17 nm pentalamellar junctions were included in this specimen. Deep-etch replica showed that gap junctional plaques also had particles on their cytoplasmic surfaces clearly distinguished from the surrounding particle-free non-gap junctional surfaces. Treatment of isolated lens membranes with endoproteinase glu-C induced cleavage of MP70 to MP38 in SDS-PAGE and loss of their antigenicity to the anti-MP70 mono-clonal antibody, whereas the particulate patterns persisted still on the cytoplasmic surfaces of gap junctional plaques, suggesting the coexistance of additional proteins other than MP70 in the lens fiber gap junction structures.
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