Regulation of Overall Cell Numbers During Epithelial Tissue Homeostasis and Pathogenesis

项目概要/摘要 细胞更新对于上皮组织的形成和功能是必不可少的。细胞更新的速度在 衰老，在损伤和修复过程中可以加速，并且在致癌过程中受到早熟刺激;但是， 在活组织中引导它的机制还不清楚。我们最近发现， 通过从上皮细胞挤出的细胞刺激附近干细胞的增殖，以取代丢失的细胞并维持 组织中的细胞总数。这一有趣的发现表明，挤压提供了细胞损失之间的关键联系 和增殖，从而控制细胞更新的速率。因此，确定 潜在的挤压可能会提供新的见解内源性过程，可以利用促进 细胞替换或防止不需要的新细胞的添加。我们的长期目标是定义细胞 以及上皮组织中细胞更新速率的分子机制。使用显影 为了研究活的上皮组织中的细胞挤出，我们发现注定要挤出的细胞改变了它们的 机械性能的形式脉动肌动球蛋白收缩，这是由富集的控制， 生物活性脂质鞘氨醇-1-磷酸（S1P）。我们还询问了细胞损失诱导的信号事件 和细胞反应，包括炎症细胞募集和表皮细胞增殖， 周转我们发现表皮生长因子受体配体epigen的表达显著上调， （EPGN）诱导细胞挤出后，提示EPGN的短暂增加可能有助于维持上皮细胞的生长。 在细胞丧失期间形成和功能。与这一想法一致，我们发现用重组人 EPGN（hrEPGN）抑制接受损伤刺激后的上皮细胞挤出，这反过来又减少了 代偿性干细胞增殖这些数据导致了一个假设，即EPGN调节挤出， 上皮组织中细胞更新的速率。研究细胞周转和测试的一个巨大挑战是 在活的有机体中的这一假设涉及可视化和扰动挤压之间的复杂相互作用， 细胞，周围的干细胞，取代失去的细胞和免疫细胞，以感知和响应中断 在完整性。因此，我们创造了工具来单独操纵不同的细胞和分子成分 或组合在发育中的斑马鱼的活上皮组织中进行，并分析在发育中的斑马鱼的上皮组织中的周转变化。 先天免疫系统的存在。我们在未来五年的工作中将采用这种新方法， 重点关注我们正在进行的研究中出现的三个基本领域，并解决我们知识中的关键空白 细胞周转的过程。首先，我们将确定调节物理力局部变化的机制 这是通过挤压去除缺陷细胞所需的。其次，我们将确定细胞损失如何促进 改变周围干细胞的表观遗传和转录状态以刺激增殖， 替换丢失的细胞。第三，我们将确定先天免疫系统在促进细胞更新中的作用 和维持上皮组织的稳态。

PROJECT SUMMARY/ABSTRACT Cellular turnover is essential for the form and function of epithelial tissues. The rate of cell turnover slows during aging, can be accelerated during injury and repair, and is precociously stimulated during carcinogenesis; but the mechanisms guiding it in living tissues is not well understood. We recently discovered that elimination of unfit cells by extrusion from epithelia stimulates proliferation of nearby stem cells to replace the lost cells and maintain overall cell numbers in the tissue. This intriguing finding suggests extrusion provides a key link between cell loss and proliferation, and thereby controls the rate of cell turnover. Thus, identification of the mechanisms that underlie extrusion may provide new insights into endogenous processes that can be leveraged to promote cellular replacement or prevent the unwanted addition of new cells. Our long-term goal is to define the cellular and molecular mechanisms underlying the rate of cellular turnover in epithelial tissues. Using the developing zebrafish to study cell extrusion in a living epithelial tissue, we have found that cells fated to extrude alter their mechanical properties in the form of pulsatile actomyosin contractions that are controlled by enrichment of the bioactive lipid sphingosine-1-phosphate (S1P). We have also interrogated the cell loss-induced signaling events and cellular responses, including inflammatory cell recruitment and epidermal cell proliferation, that drive turnover. We identified a significant upregulated expression of the epidermal growth factor receptor ligand epigen (EPGN) upon induced cell extrusion, suggesting that transient increases in EPGN may aid in sustaining epithelial form and function during cell loss. Consistent with this idea, we found that treatment with recombinant human EPGN (hrEPGN) suppressed epithelial cell extrusion after receiving damage stimuli, which in turn reduced the compensatory stem cell proliferation. These data led to the hypothesis that EPGN regulates extrusion to dictate the rate of cellular turnover in epithelial tissues. One formidable challenge to studying cellular turnover and testing this hypothesis in a living organism involves visualizing and perturbing the complex interplay between extruding cells, the surrounding stem cells that replace the lost cells and immune cells to sense and respond to disruptions in integrity. Therefore, we created tools to manipulate different cellular and molecular components individually or in combination in living epithelial tissues of developing zebrafish and analyze changes to turnover in the presence of an innate immune system. Our work over the next five years we utilize this new approach and will focus on three essential areas that emerged from our ongoing studies and address key gaps in our knowledge of cellular turnover. First, we will determine the mechanisms regulating the localized changes in physical forces that are required to remove defective cells by extrusion. Second, we will determine how cell loss promotes changes in the epigenetic and transcriptional states in surrounding stem cells to stimulate proliferation and replace the lost cells. Third, we will determine the role of the innate immune system in promoting cell turnover and maintenance of epithelial tissue homeostasis.