Quantitative analysis of epithelial cell scatter

项目总结 这项拟议工作的主要目的是发展对上皮细胞的定量理解。 散布，这一过程与癌症发展的晚期转移阶段密切相关。在.期间 转移，上皮组织结构被严重破坏，因为上皮细胞逃离其 原发部位和侵袭周围组织。一种有效的涉及细胞的体外转移模型 散开。在体外，上皮细胞成团生长，让人想起它们的单层，紧密排列 活体内的形态。与转移相关的基因扰动促进细胞从 聚集并散布到周围地区。因此，细胞散射试验被用来鉴定 癌基因(OGs)和肿瘤抑制基因(TSG)可能在肿瘤转移中发挥作用。然而， 目前的方法仅限于定性描述，很难评估如何 一个特定的OG/TSG可能是有效的，以及哪些组合显示出最大的协同效应。回答 这样的问题可以引导我们选择最有效的药物靶点，并可以帮助我们设计 最有效的联合治疗。此外，为了更深入地了解OG/TSG如何 数量上影响群体水平的表型，必须检查细胞水平的过程， 有助于多细胞散射。这些像元级属性包括单个像元的迁移 和多细胞群。此外，迁移中的细胞会发生碰撞。无论这些碰撞是否重新播撒小的种子 或碰撞的细胞是否“反弹分开”(类似于弹性碰撞)会影响范围和 细胞分散的动力学。目前量化这些单元级属性的技术过于繁琐 能够对大量OG/TSG的影响进行系统规模的定量分析。 在拟议的工作中，我们试图解决这些挑战，以便阐明 黏附微环境和OG/TSGs对细胞散射的影响。我们将量化两个细胞- 以及使用集成了自动化的多方面策略的细胞分散的总体水平方面 高通量成像和微图案化。具体目标是： 1.发展细胞水平运动特性的定量自动化测量方法。 2.阐明黏附微环境对多细胞散射的定量影响 基础单元格级属性。 3.阐明转移基因对多细胞的数量贡献和协同作用 散布和到基础单元格级属性。 拟议工作的结果将提供对OG/TSGs和 黏附微环境影响细胞水平的运动特性和多细胞分散，这是一个过程 与转移密切相关。

PROJECT SUMMARY The major aim of the proposed work is to develop a quantitative understanding of epithelial cell scatter, a process that is closely linked to late metastatic stages of cancer development. During metastasis, epithelial tissue structure is significantly disrupted as epithelial cells escape from their primary site and invade surrounding tissue. An effective ex vivo model of metastasis involves cell scatter. Epithelial cells grow in clusters ex vivo, reminiscent of their monolayer, well-packed morphology in vivo. Metastasis-associated genetic perturbations promote cells to "peel away" from clusters and scatter into the surrounding region. Thus, the cell scatter assay has been used to identify oncogenes (OGs) and tumor suppressor genes (TSGs) that may play a role in metastasis. However, current approaches are limited to qualitative characterizations from which it is difficult to assess how potent a particular OG/TSG might be and which combinations exhibit the most synergism. Answers to such questions can guide us to the most potent choice of drug targets and could help us design the most effective combination treatments. Furthermore, to understand more deeply how OG/TSGs quantitatively affect population-level phenotype, it is essential to examine the cell-level processes that contribute to multicellular scatter. These cell-level properties include the migration of individual cells and multicellular groups. In addition, migrating cells will collide. Whether these collisions re-seed small clusters or whether colliding cells "bounce apart" (akin to an elastic collision) will affect the extent and dynamics of cell scatter. Current techniques to quantify these cell-level properties are too cumbersome to permit quantitative, systems-scale analysis of the effects of numerous OG/TSGs. In the proposed work, we seek to address these challenges in order to elucidate the quantitative effects of the adhesive microenvironment and OG/TSGs on cell scatter. We will quantify both the cell- and population-level aspects of cell scatter using a multi-faceted strategy that integrates automated high-throughput imaging and micropatterning. The Specific Aims are: 1. To develop quantitative automated methods for measuring cell-level motility properties. 2. To elucidate the quantitative effect of the adhesive microenvironment on multicellular scatter and the underlying cell-level properties. 3. To elucidate the quantitative contributions and synergisms of metastatic genes to multicellular scatter and to the underlying cell-level properties. Results from the proposed work will provide a deeper quantitative understanding of how OG/TSGs and the adhesive microenvironment affect cell-level motile properties and multicellular scatter, a process closely related to metastasis.