Kino K, Fushimi K, Gao C, Shima T, Mihara K, Namba M
Department of Cell Biology, Okayama University Medical School, Japan.
In Vitro Cell Dev Biol Anim. 1997 Sep;33(8):628-32. doi: 10.1007/s11626-997-0113-3.
The study of in vitro cell transformation is valuable for understanding the multistep carcinogenesis of human cells. The difficulty in inducing neoplastic transformation of human cells by treatment with chemical or physical agents alone is due to the difficulty in immortalizing normal human cells. Thus, the immortalization step is critical for in vitro neoplastic transformation of human cells. We transfected a mutant p53 gene (mp53: codon 273Arg-His) into normal human fibroblasts and obtained two G418-resistant mp53-containing clones. These clones showed an extended life span but ultimately senesced. However, when they were treated with either 4-nitroquinoline 1-oxide or X-rays, they were immortalized. The immortalized cells showed both numerical and structural chromosome abnormalities, but they were not tumorigenic. The expression of mutant but not wild type p53 was detected in the immortalized cells by RT-PCR. Expression of p21, which is located downstream of p53, was remarkably reduced in the immortalized cells, resulting in increased cdk2 and cdc2 kinase activity. However, there was no significant difference between the normal and immortalized human cells in expression of another tumor suppressor gene, p16. These findings indicate that the p53-p21 cascade may play an important role in the immortalization of human cells.
体外细胞转化研究对于理解人类细胞的多步骤致癌过程具有重要价值。仅通过化学或物理试剂处理诱导人类细胞发生肿瘤转化存在困难，这是由于使正常人细胞永生化存在困难。因此，永生化步骤对于人类细胞的体外肿瘤转化至关重要。我们将突变型p53基因（mp53：密码子273Arg-His）转染到正常人成纤维细胞中，获得了两个对G418有抗性的含mp53克隆。这些克隆显示出延长的寿命，但最终衰老。然而，当用4-硝基喹啉-1-氧化物或X射线处理它们时，它们实现了永生化。永生化细胞显示出数量和结构染色体异常，但它们不具有致瘤性。通过RT-PCR在永生化细胞中检测到突变型而非野生型p53的表达。位于p53下游的p21的表达在永生化细胞中显著降低，导致cdk2和cdc2激酶活性增加。然而，另一个肿瘤抑制基因p16在正常人和永生化人细胞中的表达没有显著差异。这些发现表明，p53-p21级联可能在人类细胞的永生化中起重要作用。
