Tissue culture studies on immortalized Schwann cells.

1. 用含有SV-40大Tantigen基因的质粒转染成年小鼠背根神经节（DRG）和周围神经培养的雪旺细胞，其中一株被命名为MS1的转染细胞系具有不同的雪旺细胞表型，如S100蛋白、层粘连蛋白、2′，3′-环核苷酸3′-磷酸二酯酶（CNP）和Poprotein。二丁基环腺苷-3′，5′-单磷酸腺苷（dbc AMP）、福斯克林和维甲酸孵育可部分抑制MS1细胞的生长。采用溴脱氧尿苷（BrdU）掺入法和双免疫荧光法检测S100和BudU，研究了胎牛血清（FCS）、血小板源性生长因子（PDGF）、成纤维细胞生长因子（FGF）、转化生长因子- β (tgf - β)和forskolin对成年小鼠雪旺细胞的有丝分裂作用。在含有FOS的培养基中，PDGF-BB、碱性FGF、tgf - β 1和β 2对Schowann细胞均具有丝分裂性。福斯林抑制有丝分裂活性。在无血清培养基中，PDGF-BB和bFGF也具有有丝分裂性，但tgf - β 1和β 2没有。肝素- sepharose色谱法获得的FCS的肝素结合部分与TGF-beta1和beta2协同产生有丝分裂反应。由于免疫吸收和免疫印迹检测均未检测到PDGF-BB、酸性FGF、碱性FGF，因此表明成年小鼠雪旺细胞FCS生物活性中存在未识别的肝素结合分子。我们从成年小鼠雪旺细胞的长期培养中获得了几个瞬时永生化的细胞系。从其中一个细胞系衍生的条件培养基对原始雪旺细胞培养物具有有丝分裂活性。这些培养物可用于体外研究雪旺细胞的自分泌生长机制。我们建立了神经鞘骨髓鞘病小鼠的雪旺细胞培养，该小鼠出现肝脾肿大和伴震颤的共济失调运动。用抗thy1.2处理小鼠DRG和周围神经原代培养物。和兔补体，得到纯化的雪旺细胞培养物。电镜下可见片状包涵体。我们还从长期培养中获得了自发永生化的雪旺细胞菌落。这些雪旺细胞培养物可用于spm小鼠神经系统受累的研究。少

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 … More 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