Cell Structure and Function
Online ISSN : 1347-3700
Print ISSN : 0386-7196
ISSN-L : 0386-7196
Volume 19, Issue 1
Displaying 1-6 of 6 articles from this issue
  • Renu Wadhwa, Sunil C. Kaul, Youji Mitsui
    1994 Volume 19 Issue 1 Pages 1-10
    Published: 1994
    Released on J-STAGE: March 27, 2006
    JOURNAL FREE ACCESS
    The roots of cellular mortality-limited capacity of normal cells to divide, and immortalization-unabated proliferation of cancerous cells remain undefined so far. Out of a variety of experimental strategies employed, the cell fusion approach has been proven to be significantly informative. The present article reviews some of the more important recent results and describes the use of natural and conditional aging systems obtained by the fusion of mortal and immortal mouse fibroblasts to identify putative senescence-determining and/or senescence-escaping genes. The strategy has led to the isolation of a novel 66-kDa protein, mortalin- a unique member of the mouse heat shock protein 70 (hsp 70) family. The intracellular distributions of mortalin, i.e., cytosolic and perinuclear, distinguish the mortal phenotype from the immortal one, respectively. Consistently, the cytosolic mortalin is seen to have a senescence-inducing function in contrast to the perinuclear mortalin which has no detectable effect on cellular phenotype. It is suggested that mortalin can be exploited to unravel some aspects of cellular mortality and immortality and also for the early detection of cancerous cells.
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  • Norio Kawai, Ichiro Ichihara
    1994 Volume 19 Issue 1 Pages 11-19
    Published: 1994
    Released on J-STAGE: March 27, 2006
    JOURNAL FREE ACCESS
    To study ultrastructurally the mechanisms of lysosome reactions to cell membrane-derived intracellular membranes we developed a cell free system using small inside-out and rightside-out cell membrane vesicles (IOVs and ROVs)as a target of the reactions. The IOVs were generated from rat erythrocyte ghosts in a low ionic strength alkaline solution in the absence of divalent cations after erythrocytes were reacted with wheat germ agglutinin-coated colloidal gold [WGA (CG)], while ROVs were from ghosts homogenized in a buffer with MgSO4 and bovine serum albumin-coated CG [BSA (CG)]. WGA (CG) s bound to the cell surface were rearranged on the membrane and distributed irregularly on the inner surface of generated small IOVs. A coat structure derived from the ghost's submembranous coat was almost depleted from their outer surface. By contrast, BSA (CG)-binding to the membranes was negligible in the process of ROV formation. When isolated rat liver lysosomes were incubated with these WGA (CG)-binding small IOVs at 37°C, CG particles were found in several lysosomes under electron microscopy. Some lysosomes adhered to the IOVs, and their limiting membranes were found to collapse and disappear partially at the adhering region, suggesting their fusion. This reaction seems to occur even in cytosol-free solution. By contrast, the lysosomes indicated very low reaction to BSA (CG)-containing ROV, and to WGA (CG) or BSA (CG) alone. Therefore, it is suggested that isolated liver lysosomes react, at least to fuse, in a cytosol-independent fashion, with surface coat-depleted IOVs derived from WGA (CG)-bound and then -rearranged erythrocyte membranes.
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  • Keiji Sugasawa, Junko Deguchi, Toyokazu Okami, Akitsugu Yamamoto, Koic ...
    1994 Volume 19 Issue 1 Pages 21-28
    Published: 1994
    Released on J-STAGE: March 27, 2006
    JOURNAL FREE ACCESS
    The retinal pigment epithelium (RPE) is unique in that Na, K-ATPase is predominantly localized on its apical surface. We studied the distributions of Na, K-ATPase and glucose transporter GLUT1, insulin and transferrin receptors in developing rat RPE cells immunocytochemically. Na, K-ATPase, first detected in 17-day-old embryonic eyes, was already distributed predominantly on the apical surface. This reversed distribution of Na, K-ATPase was maintained throughout their life. Insulin receptor and transferrin receptor were distributed exclusively on the basolateral surface. By quantitative immunogold electron microscopic technique we found that glucose transporter GLUT1 is distributed almost equal in amount on both the apical and basolateral surfaces of RPE cells, thus presumably constructing an efficient pathway for glucose transport from the choriocapillaries to the neural retina through the blood-retinal barrier. These results suggest that in the RPE cells the intrinsic basolateral plasma membrane proteins are sorted out at least in three different ways.
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  • Eimei Sato, Maki Inoue, Yuji Takahashi, Yutaka Toyoda
    1994 Volume 19 Issue 1 Pages 29-36
    Published: 1994
    Released on J-STAGE: March 27, 2006
    JOURNAL FREE ACCESS
    The in vitro effects of derivatives of cyclic adenosine 3', 5'-monophosphate (cAMP) and glycosaminoglycans (GAGs) on the spontaneous induction of fragmentation of cultured porcine oocytes were examined. Oocytes cultured for 72h or longer undergo spontaneous fragmentation, and the percent of fragmented oocytes increased thereafter. The fragmented oocytes consisted of several "blastomeres" showing uneven distribution of DNA among the "blastomeres". Cytoplasmic bodies were also identified on the surface of fragmented oocytes and in the space among fragmented "blastomeres". Dibutyryl cyclic adenosine 3', 5'-monophosphate (dbcAMP) at concentrations exceeding 50μM markedly increased the induction of fragmentation. 8-bromoadenosine 3', 5'-cyclic monophosphate, butyrate, cAMP and GAGs isolated from porcine follicular fluid (pFF) did not stimulate the induction of fragmentation. pFF-GAGs added to the suspending medium at concentrations of 10 μg/ml or greater prevented the occurrence of dbcAMP-stimulated fragmentation of isolated porcine oocytes in a dose-dependent manner. Preparations of hyaluronic acid and chondroitin sulfate from a commercial source prevented the occurrence of fragmentation stimulated by dbcAMP. The present findings suggest that cAMP may involve the induction of fragmentation of porcine oocytes and GAGs prevent the activation of cAMP-dependent fragmentation.
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  • Hidetaka Hashi, Mika Hatai, Fusao Kimizuka, Ikunoshin Kato, Yoshihito ...
    1994 Volume 19 Issue 1 Pages 37-47
    Published: 1994
    Released on J-STAGE: March 27, 2006
    JOURNAL FREE ACCESS
    We constructed a fusion protein of the cell-binding domain of human fibronectin and human basic fibroblast growth factor, and prepared a polypeptide with both cell-adhesive activity and growth factor activity. A human gene fragment coding for basic fibroblast growth factor was amplified by the polymerase chain reaction, and introduced into the expression vector pTF7520, which encodes the cell-binding domain of human fibronectin. The resulting plasmid encoded a fusion protein in which basic fibroblast growth factor was added covalently to the C-terminal end of the fibronectin fragment. The fusion protein was expressed in Escherichia coli JM109 cells and purified from the extract by heparin affinity chromatography. The purified fusion protein had cell-adhesive activity toward BALB/c 3T3 cells, and stimulated their DNA synthesis in serum-depleted cultures. The fusion protein gave maximum mitogenic activity at the concentration of 10nM. The fusion protein adsorbed to culture dishes, or added to collagen gels, stimulated the growth of human umbilical-vein endothelial cells. The fusion protein stimulated the angiogenesis in chorioallantoic membranes of developing chick embryos.
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  • Yasuki Ogawa, Noriko Ohno, Kayoko Kameoka, Sonomi Yabe, Tetsuo Sudo
    1994 Volume 19 Issue 1 Pages 49-56
    Published: 1994
    Released on J-STAGE: March 27, 2006
    JOURNAL FREE ACCESS
    Bioassay and northern blot analyses revealed that, among several functional murine macro-phage (Mφ) clones, lipopolysaccharide (LPS) stimulation generated in distinct induction levels of granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), and macrophage colony-stimulating factor (M-CSF). When compared with these induction profiles, the Mφ clones could be classified into two types ; G type (G-CSF+++GM-CSF-M-CSF+) and GM type (G-CSF±GM-CSF+++M-CSF+) of Mφ clones. Unlike G-CSF and GM-CSF that were inducible factors, M-CSF mRNA was constitutively expressed without stimulation and was differentially controlled between the G and GM types ; LPS induction decreased M-CSF mRNA in the former, but increased it in the latter. Further northern blot analysis revealed that interferon-γ (IFN-γ) suppressed constitutive expression of M-CSF mRNA, and that costimulation with both LPS and IFN-γ reduced expression of G-CSF and M-CSF mRNA in the G type of Mφ clone, but induced higher expression of GM-CSF and M-CSF mRNAs in the GM type of Mφ clone compared with LPS alone. However, in either case, IFN-γ completely inhibited LPS-induced production of active CSF of the Mφ clones, which was observed even in IFN-γ pretreatment, and also abrogated autoactivation of GM-CSF. Our present results suggested that murine Mφ clones had differentially regulated expressions of CSFs and that IFN-γ had a regulatory function of inhibiting CSF production of murine Mφs.
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