MOLECULAR AND CYTOGENETIC STUDIES OF HUMAN CELL AGING

正常人体细胞在体外表现出严格的限制 分裂能力与肿瘤来源的和病毒来源的相比， 致癌物或辐射转化的细胞可以分裂 无限期的(不朽的)。我们还不了解其中的机制 这限制了正常人类细胞的分裂潜力或 产生永生细胞的变化。然而，从单元格 杂交研究我们发现，不朽的表型导致 从隐性变化中控制正常细胞的生长。我们有 利用这一事实分离出24个不同的永生细胞 分成四个实施组，无限期划分。 这表明至少有四种途径可以长生不老。 至少有四个基因或一组基因参与了 正常的细胞生长控制。我们还发现，这是正常的 衰老的人类细胞表达一种DNA合成的蛋白质抑制物 含有大量抑制DNA合成的mRNAs， 这两种物质都不存在于年轻的增殖细胞中。我们的 目前的假设是，这种蛋白质的表达是最终- 导致细胞衰老的遗传程序的时间点。 现在我们已经将永生细胞系分配到特定的组 我们可以继续确定发生了哪些更改 对于互补组内的细胞系来说是共同的，可能是 与不朽有关。 细胞遗传学分析结合限制性片段的使用 长度多态可以用于识别 例如，与某些特定基因变化相关的特定基因变化 肿瘤类型、囊性纤维化和杜氏肌营养不良。 我们计划采取类似的方法来确定染色体的变化 在一个互补的不朽细胞系中是常见的 一群人。然后我们就可以确定这些染色体是否 通过杂交或微细胞融合研究而永垂不朽。 我们还将分析不同细胞系中的 实施小组，以确定他们是否已成为 因为蛋白质抑制物表达的变化而永生 在衰老细胞中产生的DNA合成。结果是 从这些研究中应该可以进一步洞察在 在基因和分子水平上参与 使人类细胞永生，从而使基因或基因集永生 与细胞衰老有关的基因。

Normal human cells in vitro exhibit a stringent limitation of division capacity in contrast to tumor-derived and virus-, carcinogen- or irradiation-transformed cells that can divide indefinitely (immortal). We do not yet understand the mechanisms that limit the division potential of normal human cells or the changes occurring to yield immortal cells. However, from cell hybrid studies we have found that the immortal phenotype results from recessive changes in normal cell growth control. We have exploited this fact to separate twenty four different immortal cell lines into four implementation groups for indefinite division. This indicates that there are at least four paths to immortality and that at least four genes or sets of genes are involved in normal cell growth control. We have also found that normal senescent human cells express a protein inhibitor of DNA synthesis and contain a high abundance of mRNAs that inhibit DNA synthesis, neither of which is present in young proliferating cells. Our current hypothesis is that expression of this protein is the end- point of a genetic program that results in cellular senescence. Now that we have assigned immortal cell lines to specific groups we can proceed to determine what changes have occurred that are common to cell lines within a complementation group that could be involved in immortalization. Cytogenetic analysis combined with the use of restriction fragment length polymorphisms have allowed for the identification of specific genetic changes associated with, for example, certain types of tumors, cystic fibrosis and Duchenne muscular dystrophy. We plan to take a similar approach to identify chromosomal changes that are common to immortal cell lines within a complementation group. We can then determine whether these chromosomes are involved in immortalization by hybrid or microcell fusion studies. We will also analyze cell lines within the different implementation groups to determine whether they have become immortal because of changes in expression of the protein inhihitor of DNA synthesis that is produced in senescent cells. The results from these studies should provide further insight into changes at the genetic and molecular level that are involved in immortalization of human cells and thereby to the genes or sets of genes involved in cellular senescence.