Structure and Function of a Ganglioside found Exclusively on the Surface of Neuroblastoma cells

我们用完整的神经母细胞瘤GOTO细胞和黑色素瘤G361细胞免疫小鼠，获得了两种有趣的单克隆抗体（mAb）， mAb I-13和mAb AG23，它们分别识别细胞表面抗原。我试图鉴定两种单克隆抗体的抗原，并利用这些单克隆抗体研究GOTO细胞的分化。通过荧光活化细胞分选（FACS）分析各种细胞，mAb I-13可以很好地识别神经母细胞瘤细胞GOTO和IMR-32，但不能识别其他细胞。另一方面，mAb AG23识别除GOTO外的所有测试细胞。结果表明，mAb I-13的抗原为鞘脂，AG23的抗原为糖蛋白，经CD44.5-Bromo-2' -脱氧尿苷（BrdU）处理的GOTo细胞分化为雪旺细胞后，在细胞表面表达CD44。表达的CD44种为造血形式（CD44H），编码长细胞质尾。经血清缺失分化为神经细胞的GOTO细胞不表达CD44。另一方面，GOTO细胞分化为雪旺细胞和神经细胞后，对mAb I-13的反应有所降低。抗原脂质只存在于膜微域，称为脂筏。brdu处理的GOTO细胞中的CD44也会分裂成脂筏，尽管在黑色素瘤G361细胞的质膜部分和脂筏部分中都发现了CD44。我认为brdu处理的GOTO细胞中的CD44可能在细胞分化为雪旺细胞中发挥重要作用。

We immunized mice with intact cells of neuroblastoma GOTO cells and melanoma G361 cells and obtained two interesting monoclonal antibodies(mAbs), mAb I-13 and mAb AG23, which respectively recognized cell surface antigens. I tried to identify the antigens for two mAbs, and I investigated the differentiation of GOTO cells using these mAbs.By fluorescence-activated cell sorter(FACS)analysis of various cells, the mAb I-13 recognized neuroblastoma cells GOTO and IMR-32 very well, but did not recognize other cells tested. On the other hand, the mAb AG23 recognized all cells tested except GOTO.It turned out that the antigen for mAb I-13 was a sphingolipid and that for AG23 was a glycoprotein, CD44.5-Bromo-2' -deoxyuridine(BrdU)-treated GOTo cells, which are differentiated int Schwann cells, expressed CD44 on the cell surface. The CD44 species expressed was the hemopoietic form(CD44H)encoding long cytoplasmic tail. GOTO cells differentiated into neural cells by serum depletion did not express CD44. On the other hand, GOTO cells became a little less responsive with the mAb I-13 after the differentiation into Schwann as well as neural cells.The antigen lipid was found exclusively in membrane microdomain, called lipid rafts. CD44 in BrdU-treated GOTO cells also partitioned itself into lipid rafts, although it was found in both plasma membrane fractions and lipid rafts fractions in the case of melanoma G361 cells. I think that CD44 in BrdU-treated GOTO cells might play an important role in cells differentiated into Schwann cells.