Age-related Responses of Epidermal Stem Cells to Environ

持续更新的组织(如表皮)的动态平衡由体细胞干细胞维持。这些细胞是未分化的、自我更新的细胞，也会产生子代瞬时扩增(TA)细胞，这些细胞构成了增殖群体的大多数。TA细胞在离开增殖室并走向终末分化之前经历了有限数量的细胞分裂，而人们一直认为干细胞在有机体的整个生命周期中持续存在。通过一系列的氚胸苷标记实验，我们之前发现来自成年小鼠皮肤的干细胞不像其他基底细胞那样分裂频繁，但它们在体内确实以稳定的速度分裂。我们还表明，它们在整个生命过程中都会在体内继续增殖，并且在体外具有很高的增殖潜力。因此，表皮干细胞可能并不遵循海弗利克的理论，可能会“永生”。去年，我们开发了一种分选方法，可以从表皮中提取出可存活的干细胞群体。我们证明了这些干细胞具有再生表皮并在再生组织中持续表达重组基因的能力。因此，它们是表皮的干细胞。最近，我们确定这些表皮干细胞也具有显著的参与其他组织形成的能力，这种可塑性类似于胚胎干细胞和其他一些体细胞干细胞。我们的初步数据显示，从新生小鼠皮肤中分离的表皮干细胞可以整合到各种组织中，并在注射到囊胚后改变其表型。因此，这些干细胞的命运似乎直接取决于它们所处的环境。我们还确定，这些细胞的细胞周期曲线似乎是干细胞的固有属性，与细胞的年龄无关。这些数据让我们不禁要问，随着生物体的衰老，表皮干细胞会发生什么变化？在这一应用中，我们建议测试干细胞的年龄是否影响其对环境的反应或决定其命运路径的外部因素。我们提出了以下具体目标：1)比较老年和年轻小鼠表皮干细胞的数量、表型变化和注入囊胚后的寿命，并确定注入老年和年轻干细胞的囊胚小鼠的寿命是否存在差异；以及2)比较老年和年轻人类表皮干细胞的体外生长潜力，以及它们在体外和注入囊胚后对外部环境信号的反应是否不同。

Homeostasis of continuously renewing tissues, such as the epidermis, is maintained by somatic stem cells. These are undifferentiated, self-renewing cells, which also produce daughter transient amplifying (TA) cells that make up the majority of the proliferative population. TA cells undergo a finite number of cell divisions before leaving the proliferative compartment and moving toward terminal differentiation, whereas it has been assumed that the stem cells persist throughout the lifetime of the organism. Through a series of labeling experiments with tritiated thymidine, we previously showed that stem cells from adult mouse skin did not divide as often as the other basal cells, but they did divide at a steady rate in vivo. We also showed that they continued to proliferate in vivo throughout life, and that they have a high proliferative potential in vitro. Thus, it may be that epidermal stem cells do not follow the Hayflick theory and possibly "live forever." Last year, we developed a sorting method that yields a viable population of stem cells from the epidermis. We showed that these stem cells have the capacity to regenerate the epidermis and to continuously express a recombinant gene in the regenerated tissue. Thus, they are the stem cells for the epidermis. Very recently, we determined that these epidermal stem cells also have the remarkable ability to participate in the formation of the other tissues, a plasticity similar to that of embryonic stem cells and a few other somatic stem cells. Our preliminary data show that the epidermal stem cells isolated from neonatal mouse skin incorporate into a variety of tissues and alter their phenotype after injection into blastocysts. Thus, the fate determination of these stem cells appears to be in direct response to their environment. We also determined that the cell cycle profile of these cells appears to be an intrinsic property of the stem cell and not related to the age of the cell. These data have led us to ask what happens to epidermal stem cells as the organism ages? In this application, we propose to test whether the age of the stem cells has an effect on its response to the environment or extrinsic factors in determining its fate pathway. We propose the following specific aims: 1) to compare old vs. young murine epidermal stem cells by determining their number, altered phenotype and longevity after injection into blastocysts, and to determine whether there is a difference in the life span of mice derived from blastocysts injected with old vs. young stem cells; and 2) to compare old vs. young adult human epidermal stem cells by determining if there is a difference in their in vitro growth potential and if they respond differently to extrinsic environmental signals in vitro and after injection into blastocysts.