Genes Responsible for Senescence and Differentiation

描述：（由申请人提供）我们研究的长期目标是 了解人体细胞衰老和分化的机制 上皮细胞作为实现我们目标的第一步，我们已经建立了一个 细胞衰老和分化的体外模型 传代培养的正常人口腔角质形成细胞（NHOK）。NHOK最多经历了 通过不同的复制阶段，22+3个群体倍增（PD）： 指数、衰老、衰老和终末分化。 利用我们的体外模型，我们研究了NHOK的衰老模式， 与其它人体细胞如人二倍体成纤维细胞相比， （HDF）。与HDF相反，指数复制的NHOK显示端粒酶 活性，并在没有端粒缩短的情况下进入衰老阶段。 此外，与霍夫相反，其衰老程序由p53触发， p21 WAF 1/CIP 1，衰老的NHOK表现出这些蛋白的水平降低。 这些发现表明NHOK在端粒中进入衰老 长度无关的方式，并通过与HDF不同的机制。我们 还发现继代培养的NHOK具有衰老的特征， 以顺序不同的方式分化，这表明， 在体外传代过程中，这些机制引起这些过程。因此我们 提出了一个问题：什么基因构成了复制的分子途径？ 衰老和分化的NHOK？这个问题将在 本提案的具体目标如下：（1）调查 在指数复制中的基因表达谱和动力学， 衰老，衰老和终末分化的NHOK在连续 传代培养，（2）鉴定与衰老相关的基因和/或 NHOK在体外的分化，和（3）确定是否异位 所鉴定的基因的表达可以触发衰老， 在指数复制的NHOK和永生化的 人口腔角质形成细胞。 有了这些具体目标，我们希望解决以下问题： 基因参与衰老和/或分化在NHOK？是同一类型 是否存在参与衰老和分化途径的基因？可以 我们用特定的基因诱导细胞衰老和/或分化， 我们的具体目标？预计，这一结果 这一建议将发展成为新的假设，我们可以进一步研究 细胞复制、衰老和分化的分子机制 人类上皮细胞。

DESCRIPTION: (provided by applicant) The long-term goal of our research is to understand the mechanism of cellular senescence and differentiation in human epithelial cells. As the first step toward our goal, we have established an in vitro model of cellular senescence and differentiation with serially subcultured normal human oral keratinocytes (NHOK). NHOK underwent a maximum of 22+3 population doublings (PDs) through distinct phases of replication: exponential, senescing, senescent and terminal differentiation. Using our in vitro model, we have investigated the mode of senescence of NHOK in comparison to other human somatic cells, such as human diploid fibroblasts (HDF). In contrast to HDF, exponentially replicating NHOK showed telomerase activity and entered the senescent phase in the absence of telomere shortening. Also, in contrast to HOF, whose senescence program is triggered by p53 and p21WAF1/CIP1, senescent NHOK exhibited decreased levels of these proteins. These findings indicate that NHOK enter senescence in a telomere length-independent manner and through a distinct mechanism from that of HDF. We also found that subcultured NHOK exhibited the features of senescence and differentiation in a sequentially distinct manner, suggesting that separate mechanisms elicit these processes during in vitro passage. Accordingly, we raise a question: What genes constitute the molecular pathways of replicative senescence and differentiation in NHOK? This question will be addressed in the present proposal with the following specific aims: (1) To investigate the profile and the kinetics of gene expression in exponentially replicating, senescing, and senescent and terminally differentiating NHOK during serial subculture, (2) to identify the gene(s) associated with senescence and/or differentiation of NHOK in vitro, and (3) to determine whether the ectopic expression of the identified genes can trigger the senescence and differentiation programs in exponentially replicating NHOK and in immortalized human oral keratinocytes. With these specific aims, we expect to address the following questions: Which genes are involved in senescence and/or differentiation in NHOK? Are same type of gene(s) involved in both senescence and differentiation pathways or not? Can we induce senescence and/or differentiation of cells with specific genes identified by our specific aims? It is expected that the outcome of this proposal will develop into novel hypotheses with which we can further study the molecular mechanisms of cellular replication, senescence, and differentiation of human epithelial cells.