| Co-Investigator(Kenkyū-buntansha) |
FUKUDA Takahiro Jikei Univ.Sch.Med., Neuropathology, Res.Assist., 神経病理, 助手 (60228913)
TANAKA Junichi Jikei Univ.Sch.Med., Neuropathology, Professor, 神経病理, 教授 (60079712)
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| Budget Amount *help |
¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1993: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1992: ¥1,300,000 (Direct Cost: ¥1,300,000)
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| Research Abstract |
1. Cultured Schwann cells derived from adult mouse dorsal rootganglia (DRG) and peripheral nerves were transfected with a plasmid containing SV-40 large Tantigen gene One of transfected cell lines, designated MS1, had distinct Schwann cell phenotypes such as S100 protein, laminin, 2', 3'-cyclic nucleotide 3'-phosphodiesterase (CNP) and Poprotein, as shown by imuunofluorescence. The growth of MS1 cells was partially suppressed by incubation with dibutyryl cyclic adenosine-3', 5'-monophosphate (dbc AMP), forskolin and retinoic acid.
2. Mitogenic effects of fetal calf serum (FCS), platelet-derived growth factor (PDGF), fibroblast growth factor (FGF), transforming growth factor-beta (TGF-beta) and forskolin to adult mouse Schwann cells were examined by bromodeoxy uridine (BrdU) incorporation and double immunofluorescence for S100 and BudU.PDGF-BB, basic FGF, TGF-beta 1 and beta 2 were all mitogenic for Schowann cells in media containing FOS.Forsk olin suppressed the mitogenic activity of th
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ese factors. In serum-free media, PDGF-BB and bFGF were also mitogenic, but TGF-beta1 and beta2 were not. Heparin-binding fractions of FCS obtained by heparin-Sepharose chromatography synergized with TGF-beta1 and beta2 to produce a mitogenic response. Since PDGF-BB, acidic FGF, basic FGF were not detected in these fractions by immunoabsorption and immunoblot assays, the presence of unidentified heparin-binding molecules in FCS bioactive for adult mouse Schwann cells is suggested.
3. We obtained several of spotaneously immortalized celllines from long-term cultures of adult mouse Schwann cells. Conditioned media derived from one of the celllines had a mitogenic activity to original Schwann cell cultures. These cultures can be useful in studies on autocrine growth mechanism of Schwann cells in vitro.
4. We established Schwann cell cultures of sphingomyelinosis (spm) mouse, which develops hepatosplenomegaly and ataxic movements with tremors. Primary cultures of DRG and peripheral nerves derived from affected mice were treated with anti-Thy1.2.and rabbit complement, which resulted purified Schwann cell cultures. Electron-microscopically, characteristic lamellar inclusions were demonstrated in these cells. We also obtained spontaneously immortalized Schwann cell colonies from long-term cultures. These Schwann cell cultures can be useful in studies on the nervous system involvement of spm mouse. Less
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