Understanding How Invadosomes Breach Basement Membrane In Vivo

了解体内侵袭体如何突破基底膜

• 批准号: 8413036
• 负责人: David R Sherwood
• 金额: $ 28.05万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-02-01 至 2015-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8413036
• 关键词: 4D Imaging; Actins; Basement membrane; Behavior; Biological Process; Caenorhabditis elegans; Cancer cell line; Cell Culture Techniques; Cell membrane; Cells; Cues; Development; Developmental Process; Disease; Disseminated Malignant Neoplasm; Endothelial Cells; Epithelial Cells; Extracellular Matrix; F-Actin; Fibroblasts; Genes; Genetic; Goals; Guanosine Triphosphate Phosphohydrolases; Health; Human; Immunologic Surveillance; In Vitro; Infiltration; Integrins; Invaded; Lead; Leukocytes; Life; Malignant Neoplasms; Mediating; Membrane; Mission; Modeling; Molecular Analysis; Molecular Target; Mutagenesis; Neoplasm Metastasis; Optics; Pathway interactions; Penetration; Physiological Processes; Process; Regulation; Research; Role; Signal Transduction; Site; Specific qualifier value; Stereotyping; Structure; Subcellular structure; Therapeutic; Tissues; Visual; Work; cancer cell; cell type; cellular imaging; clinically relevant; cofilin; genetic analysis; human disease; in vivo; in vivo Model; macrophage; netrin receptor; novel; tumor progression; vasodilator-stimulated phosphoprotein

DESCRIPTION (provided by applicant): Basement membrane is a thin, dense, sheet-like extracellular matrix that encircles most tissues and provides structural support for epithelial and endothelial cells. Understanding how specialized invasive cells cross basement membrane is of profound importance to human health: Basement membrane transmigration underlies the dispersal of cells in many developmental processes, is required in leukocytes for immune surveillance and inappropriate manifestation of cell-invasive behavior underpins the development of metastatic cancers. F-actin-rich, cell membrane associated structures, termed invadosomes, were identified over three decades ago in normally invasive cell types and metastatic cancer cell lines. Invadosomes are thought to mediate the ability of cells to invade through basement membrane barriers. Owing to the difficulty of examining cell-invasive behavior in vivo, invadosomes have only been studied in vitro, where cell culture conditions do not recapitulate native environmental signals or matrix conditions. As a result, the relevance, regulation and potential functions of invadosomes in vivo remain one of the most critical gaps in our understanding of cell- invasive behavior. Anchor cell invasion in C. elegans is a simple, highly stereotyped in vivo model of cell invasion through basement membrane that uniquely combines single-cell visual and genetic analysis. Through the development of live-cell imaging approaches, our group has recently identified dynamic invadosome structures within the anchor cell that breach the basement membrane. We find that when one invadosome penetrates the BM, new invadosome formation ceases, and a single invasive protrusion matures from the infiltrated invadosome. Using optical highlighting of basement membrane components, we have found that this protrusion pushes the basement membrane aside to clear a path for invasion. The goal of this application is to uncover the mechanistic details of how these invadosomes are used to breach basement membrane. Our proposed research will combine live-cell imaging with genetic and molecular analysis to determine: (1) how key pathways that regulate anchor cell invasion coordinately regulate the formation, dynamics and function of invadosomes prior to and during BM penetration, (2) the role of netrin signaling in selecting a single invadosome for basement membrane invasion, (3) the function of the actin regulator Ena/VASP in promoting basement membrane gap expansion at the anchor cell-basement membrane contact points after invadosome penetration. Together, this work will identify new mechanisms underlying cell invasion and elucidate how they function together to breach basement membrane. This project is relevant to NIH's mission because it will lead to specific therapeutic strategies to control invasive behavior in diseases such as metastatic cancer.

描述(申请人提供)：基底膜是一种薄的、致密的、片状的细胞外基质，包围着大多数组织，为上皮细胞和内皮细胞提供结构支持。了解特殊的侵袭细胞如何穿过基底膜对人类健康具有深远的意义：在许多发育过程中，基底膜迁移是细胞扩散的基础，是免疫监视所必需的白细胞，细胞侵袭行为的不适当表现是转移性癌症发展的基础。富含肌动蛋白的细胞膜相关结构，称为侵袭体，在三十多年前在正常侵袭性细胞类型和转移性癌细胞系中被发现。侵袭体被认为通过基底膜屏障调节细胞的侵袭能力。由于很难在体内检测细胞侵袭行为，侵袭体只在体外进行研究，在体外细胞培养条件不能概括自然环境信号或基质条件。因此，体内侵袭体的相关性、调节和潜在功能仍然是我们理解细胞侵袭行为最关键的空白之一。线虫锚定细胞入侵是一种简单的、高度定型的通过基底膜进行细胞入侵的体内模型，它独特地结合了单细胞视觉和遗传分析。通过开发活细胞成像方法，我们团队最近发现锚定细胞内的动态侵袭体结构可以突破基底膜。我们发现，当一个侵袭体穿透BM时，新的侵袭体形成停止，单个侵袭性突起从浸润性侵袭体成熟。通过光学突出基底膜成分，我们发现这种突起将基底膜推到一边，为侵袭扫清了一条道路。这项应用的目标是揭示这些侵入体是如何被用来突破基底膜的机械细节。我们建议的研究将结合活细胞成像与遗传和分子分析来确定：(1)调节锚定细胞入侵的关键途径如何在BM穿透之前和期间协调调节侵袭体的形成、动态和功能；(2)netrin信号在选择单个侵袭体进行基底膜侵袭中的作用；(3)肌动蛋白调节因子Ena/Vasp在促进侵袭体穿透锚定细胞-基底膜接触点基底膜间隙扩大中的作用。总之，这项工作将确定细胞入侵的新机制，并阐明它们如何共同作用以突破基底膜。这个项目与NIH的任务相关，因为它将导致控制转移性癌症等疾病的侵袭行为的具体治疗策略。