Ambros I M, Rumpler S, Luegmayr A, Hattinger C M, Strehl S, Kovar H, Gadner H, Ambros P F
Children's Cancer Research Institute CCRI, St. Anna Kinderspital, Vienna, Austria.
Eur J Cancer. 1997 Oct;33(12):2043-9. doi: 10.1016/s0959-8049(97)00204-9.
Human neuroblastoma cell lines frequently exhibit MYCN amplification and many are characterised by the presence of morphologically distinct cell types. The neuronal cells (N-cells) and the so-called flat cells (F-cells) are thought to represent manifestations of different neural crest cell lineages and are considered to be the consequence of neuroblastoma cell pluripotency. In this study, various neuroblastoma cell lines were examined for micronuclei. In F-cells of neuroblastoma cell lines with extrachromosomally amplified MYCN, we observed the frequent occurrence of micronuclei. Using fluorescence in situ hybridisation (FISH) with a MYCN specific probe, we demonstrated that these micronuclei were packed with MYCN hybridisation signals. In addition, in a minor percentage of cells, MYCN signals occurred in clusters, adhered to the nuclear membrane and aggregated in nuclear protrusions. In F-cells, a substantial reduction or lack of amplified MYCN copies was observed. These observations let us conclude that extrachromosomally amplified genes can be actively eliminated from the nucleus resulting in a dramatic loss of amplified sequences in the F-cells. Moreover, reduction or loss of amplified sequences in F-cells was shown to be accompanied by downregulation of MYCN expression, by a decrease in proliferative activity and by upregulation of molecules of the major histocompatibility complex class I (MHC I). Interestingly, F-cells are not restricted to neuroblastoma cell cultures, but also occur in cell lines of other tissue origin. All F-cells share important biological features, interpreted as cell revertance, i.e. loss of the malignant phenotype and properties. This fact, together with the demonstration that neuroblastoma cells do not differentiate into Schwann cells in vivo [1] Ambros et al. NEJM 1996, 334, 1505-1511, do not support the hypothesis that F-cells represent Schwannian/glial differentiation in vitro. We therefore postulate that the elimination of amplified MYCN gene copies in cultivated neuroblastoma cells is in line with the phenomenon of tumour cell revertance.
人类神经母细胞瘤细胞系经常表现出MYCN基因扩增，并且许多细胞系的特征是存在形态学上不同的细胞类型。神经元细胞（N细胞）和所谓的扁平细胞（F细胞）被认为代表不同神经嵴细胞谱系的表现，并且被认为是神经母细胞瘤细胞多能性的结果。在本研究中，检测了各种神经母细胞瘤细胞系中的微核。在具有染色体外扩增的MYCN的神经母细胞瘤细胞系的F细胞中，我们观察到微核频繁出现。使用针对MYCN的特异性探针进行荧光原位杂交（FISH），我们证明这些微核充满了MYCN杂交信号。此外，在一小部分细胞中，MYCN信号成簇出现，附着于核膜并聚集在核突起中。在F细胞中，观察到扩增的MYCN拷贝大量减少或缺失。这些观察结果使我们得出结论，染色体外扩增的基因可以从细胞核中被主动消除，导致F细胞中扩增序列的显著丢失。此外，F细胞中扩增序列的减少或丢失伴随着MYCN表达的下调、增殖活性的降低以及主要组织相容性复合体I类（MHC I）分子的上调。有趣的是，F细胞不仅存在于神经母细胞瘤细胞培养物中，也出现在其他组织来源的细胞系中。所有F细胞都具有重要的生物学特征，被解释为细胞逆转，即恶性表型和特性的丧失。这一事实，连同神经母细胞瘤细胞在体内不会分化为施万细胞的证明[1]安布罗斯等人。《新英格兰医学杂志》1996年，334卷，1505 - 1511页，不支持F细胞代表体外施万细胞/神经胶质细胞分化的假设。因此，我们推测培养的神经母细胞瘤细胞中扩增的MYCN基因拷贝的消除与肿瘤细胞逆转现象一致。
