Regulation of Overall Cell Numbers During Epithelial Tissue Homeostasis and Pathogenesis

项目摘要/摘要 细胞周转对上皮组织的形态和功能至关重要。细胞周转的速度在 衰老，可以在损伤和修复过程中加速，并在致癌过程中被早熟刺激；但 在活组织中引导它的机制还不是很清楚。我们最近发现，消除不合适的 通过从上皮中挤出细胞来刺激附近干细胞的增殖，以取代丢失的细胞并保持 组织中的细胞总数。这一耐人寻味的发现表明，挤压在细胞丢失之间提供了关键联系 和增殖，从而控制细胞周转的速度。因此，确定 底层挤压可以提供对内生过程的新见解，这些过程可以被利用来促进 细胞替换或防止不想要的新细胞的增加。我们的长期目标是定义细胞 以及上皮组织中细胞周转率背后的分子机制。使用正在开发的 斑马鱼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他们的 肌动球蛋白以搏动性收缩的形式表现的机械性能，由浓缩的 生物活性脂鞘氨醇-1-磷酸(S1P)。我们还询问了细胞丢失诱导的信号事件 以及细胞反应，包括炎症细胞招募和表皮细胞增殖，这推动了 营业额。我们发现了表皮生长因子受体配体EpiGen的显著上调表达 (EPGN)在诱导细胞挤出时，提示EPGN的一过性增加可能有助于维持上皮细胞 细胞丢失时的形态和功能。与这一想法一致，我们发现用重组人 EPGN(HrEPGN)在受到损伤刺激后抑制上皮细胞挤出，这反过来又减少了 代偿性干细胞增殖。这些数据导致了一种假设，即EPGN调节挤出来决定 上皮组织中细胞周转率。研究细胞周转和测试的一个巨大挑战 在生物体中，这一假说涉及到可视化和扰乱挤压之间复杂的相互作用 细胞，周围的干细胞，取代丢失的细胞和免疫细胞，以感知和响应中断 在正直方面。因此，我们创造了工具来单独操作不同的细胞和分子成分 或组合在发育中的斑马鱼活体上皮组织中，并分析其在 先天免疫系统的存在。我们在未来五年的工作中将利用这一新方法，并将 将重点放在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差距 细胞周转率。首先，我们将确定物理力量局部变化的调节机制 这是通过挤压去除有缺陷的电池所需的。第二，我们将确定细胞丢失是如何促进 周围干细胞表观遗传和转录状态的变化，以刺激增殖和 替换丢失的信元。第三，我们将确定先天免疫系统在促进细胞更新中的作用 和维持上皮组织的动态平衡。

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.