Ikram Z, Norton T, Jat P S
Ludwig Institute for Cancer Research, London, United Kingdom.
Proc Natl Acad Sci U S A. 1994 Jul 5;91(14):6448-52. doi: 10.1073/pnas.91.14.6448.
Normal mammalian fibroblasts cultured in vitro undergo a limited number of divisions before entering a senescent phase in which they can be maintained for long periods but cannot be induced to divide. In rodent fibroblasts senescence can be prevented by expression of simian virus 40 large tumor antigen (T antigen). Cells expressing T antigen can proliferate indefinitely; however, such cells are absolutely dependent upon continued expression of T antigen for maintenance of growth; inactivation of T antigen results in a rapid and irreversible entry into a postmitotic state. To determine when, after the initial expression of T antigen, fibroblasts become dependent upon it for continued growth, we serially cultivated embryonic fibroblasts prepared from H-2Kb-tsA58 transgenic mice. We show that these fibroblasts become dependent upon T antigen for maintenance of proliferation only when their normal mitotic life-span has elapsed and that the biological clock that limits the mitotic potential continues to function normally, even in cells expressing this immortalizing gene. Our results suggest that random accumulation of cellular damage is unlikely to be the factor that limits fibroblast division but support the hypothesis that senescence is regulated via a genetic program.
体外培养的正常哺乳动物成纤维细胞在进入衰老期之前会经历有限次数的分裂，在衰老期它们可以长时间维持，但不能被诱导分裂。在啮齿动物成纤维细胞中，猿猴病毒40大T抗原（T抗原）的表达可以阻止衰老。表达T抗原的细胞可以无限增殖；然而，此类细胞的生长维持绝对依赖于T抗原的持续表达；T抗原失活会导致细胞迅速且不可逆地进入有丝分裂后状态。为了确定在T抗原最初表达后，成纤维细胞何时开始依赖于它来持续生长，我们连续培养了从H-2Kb-tsA58转基因小鼠制备的胚胎成纤维细胞。我们发现，这些成纤维细胞只有在其正常有丝分裂寿命结束后才开始依赖T抗原维持增殖，并且限制有丝分裂潜能的生物钟即使在表达这种永生化基因的细胞中仍能正常发挥作用。我们的结果表明，细胞损伤的随机积累不太可能是限制成纤维细胞分裂的因素，但支持衰老通过遗传程序调控的假说。
