Quantitative analysis of epithelial cell scatter

项目摘要 这项工作的主要目的是发展对上皮细胞的定量了解， 扩散，这是一个与癌症发展的晚期转移阶段密切相关的过程。期间 转移时，上皮组织结构被显著破坏，因为上皮细胞从它们的 原发部位并侵入周围组织。转移的有效离体模型涉及细胞 散开上皮细胞在体外成簇生长，使人联想到它们的单层，包装良好， 体内形态学转移相关的遗传扰动促进细胞从肿瘤细胞“剥离”。 聚集并分散到周围区域。因此，细胞散射测定已用于鉴定 癌基因（OG）和肿瘤抑制基因（TSG），可能在转移中发挥作用。然而，在这方面， 目前的方法仅限于定性描述，难以评估如何 特定OG/TSG可能是有效的，以及哪种组合表现出最大的协同作用。的答案 这些问题可以指导我们选择最有效的药物靶点，并帮助我们设计 最有效的联合治疗。此外，为了更深入地了解OG/TSG如何 定量地影响群体水平的表型，有必要检查细胞水平的过程， 有助于多细胞分散。这些细胞水平的特性包括单个细胞的迁移 和多细胞群体。此外，迁移的细胞会发生碰撞。这些碰撞是否会重新产生小的 群集或碰撞细胞是否“弹开”（类似于弹性碰撞）将影响程度， 细胞分散的动力学。目前的技术来量化这些细胞级的属性是太繁琐 允许对许多OG/TSG的影响进行定量的系统规模分析。 在拟议的工作中，我们寻求解决这些挑战，以阐明定量的 粘附微环境和OG/TSGs对细胞散射的影响。我们将量化细胞- 和人口水平方面的细胞分散使用多方面的战略， 高通量成像和微图案化。具体目标是： 1.建立细胞水平运动特性的自动化定量测定方法。 2.为了阐明粘附微环境对多细胞散射的定量影响， 底层单元格级属性。 3.为了阐明转移基因对多细胞肺癌的定量贡献和协同作用， 分散和底层单元格级属性。 从拟议的工作结果将提供一个更深入的定量了解如何OG/TSG和 粘附微环境影响细胞水平运动特性和多细胞分散， 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.