Nakamura T, Takeshita I, Fukui M
Department of Neurosurgery, Neurological Institute, Faculty of Medicine, Kyushu University, Fukuoka, Japan.
J Neurooncol. 1991 Dec;11(3):215-24. doi: 10.1007/BF00165529.
We have electrophoretically obtained platelet-derived growth factor (PDGF)-related protein from human glioma (glioma derived PDGF-related protein: GD-PDGF) and produced rabbit antiserum against the monomer of GD-PDGF. By methods of immunoaffinity chromatography and Western blotting, we analyzed GD-PDGF in cultured human glioma cells and conditioned medium. The intracellular GD-PDGF was only detected at 17 kd molecular weight by the purified rabbit antibody. When the intracellular 17 kd monomer was purified by the IgG-coupled immunoaffinity chromatography, the eluted protein was not detected at 17 kd but at 52 kd. The 52 kd GD-PDGF was spontaneously and immediately converted to 56 kd, which was partly degraded to 32 and 35 kd within 24 hours. On the other hand, in the conditioned media of glioma cell lines GD-PDGF presents mainly as 56 kd. The assembled forms of GD-PDGF exhibited a powerful activity to induce membrane ruffle formation and reorganization of actin filaments in cultured glial cells and glioma cells. These results indicated that GD-PDGF is intracellularly stored as 17 kd monomer and exists extracellularly as assembled forms, which may act as an autocrine and paracrine effect on the surrounding cells.
我们通过电泳从人胶质瘤中获得了血小板衍生生长因子(PDGF)相关蛋白(胶质瘤衍生的PDGF相关蛋白:GD-PDGF),并制备了针对GD-PDGF单体的兔抗血清。通过免疫亲和色谱法和蛋白质印迹法,我们分析了培养的人胶质瘤细胞和条件培养基中的GD-PDGF。纯化的兔抗体仅在17kd分子量处检测到细胞内GD-PDGF。当通过IgG偶联的免疫亲和色谱法纯化细胞内17kd单体时,洗脱的蛋白在17kd处未检测到,而是在52kd处检测到。52kd的GD-PDGF会自发且立即转化为56kd,在24小时内部分降解为32kd和35kd。另一方面,在胶质瘤细胞系的条件培养基中,GD-PDGF主要以56kd的形式存在。GD-PDGF的组装形式在培养的神经胶质细胞和胶质瘤细胞中表现出强大的诱导膜皱襞形成和肌动蛋白丝重组的活性。这些结果表明,GD-PDGF在细胞内以17kd单体形式储存,在细胞外以组装形式存在,可能对周围细胞起自分泌和旁分泌作用。