Role of p53, sdil, cdk, mdm2 genes for immortalization of human fibroblasts.

永生化是人细胞体外肿瘤转化的重要步骤。这项研究解决了突变p53是否有助于人类细胞的永生化过程的问题。将p53突变基因（mp53；密码子275Arg-His）导入正常人成纤维细胞（OUMS-24），获得抗g418克隆OUMS-24/p6。该克隆表达mp53，寿命延长，但在第79个种群加倍水平（PDL）上衰老。当这些细胞用2gy的x射线或4-硝基喹啉氧化物处理时，它们就变得不朽了。虽然永生化细胞表现出染色体异常，但它们不具有致瘤性。另一方面，没有引入mp53的正常OUMS-24细胞没有通过同样的处理而永生。位于p53下游的p21/cip1/waf1/sdi1的表达在永生化细胞系中显著降低，导致cdk2和cyclin A激酶活性增加。这些发现表明p53-p21级联可能在人类细胞的永生化中发挥一定作用。另一方面，Rb、p16、cdk4、cdk6、cyclin A、cyclinD等蛋白在正常和永生化人成纤维细胞中的表达无显著差异。综上所述，我们的研究结果表明，53基因的突变不足以实现人类细胞的永生，除了p53之外，可能还有一些未知的基因参与了人类细胞的永生过程。

Immortalization is an essential step for in vitro neoplastic transformation of human cells. This study address the question of whether mutant p53 contributes to the immortalization process of human cells.The mutant p53 gene (mp53 ; codon 275Arg-His) was introduced into normal human fibroblasts (OUMS-24), and a G418-resistant clone, OUMS-24/p6, was obtained. This clone expressed mp53 and showed an extended life span, but it senesced at 79th population-doubling level (PDL). When these cells were treated with 2 Gy of x-rays or 4-nitroquinoline 1 oxide, they became immortalized. Although the immortalized cells showed chromosome abnormalities, they were not tumorigenic. On the other hand, normal OUMS-24 cells into which mp53 had not been introduced were not immortalized by the same treatment.Expression of p21/cip1/waf1/sdi1, which is located downstream of p53, was remarkably reduced in the immortalized cell lines, resulting in an increase in cdk2 and cyclin A kinase activity. These findings indicate that the p53-p21 cascade may play some role in the immortalization of human cells. On the other hand, there was no significant different expression of proteins such as Rb, p16, cdk4, cdk6, cyclin A and cyclinD between the normal and immortalized human fibroblasts.Taken togehter, our results indicate that mutation of the 53 gene alone was not sufficient for immortalization and some unknown genes other than p53 may be involved in the immortalization process of human cells.