Probing Contact Guidance on Exquisitely Tunable ECM Surfaces

描述（由申请人提供）：长期以来，人们一直认识到肿瘤微环境中细胞外基质（ECM）的变化可以驱动肿瘤的进展。随着最近成像技术的进步，ECM成分如胶原蛋白组织的变化已经被描述。周向组织的胶原纤维被重组成径向排列的纤维，导致定向迁移或接触引导远离肿瘤。这种胶原组织特征已被提出作为潜在入侵的诊断指标，将接触指导与入侵联系起来。不幸的是，虽然在二维环境中有大量的接触制导信息，但对三维环境中的接触制导知之甚少。此外，胶原纤维网络可以在调节细胞感知和响应接触引导线索的能力方面表现出很大的特性变化，有效地改变了胶原组织和入侵之间的关系。我的长期目标是通过使用各种工程方法了解肿瘤、基质和免疫细胞如何整合多种线索在肿瘤微环境中进行定向迁移。本研究的目的是了解胶原I基质的组织和组成在指导接触指导中的作用。这项建议的具体目标包括：(1)评估拓扑和约束在解释二维和三维环境下接触引导差异方面的相对贡献；(2)验证纤维结构（如纤维和交联密度以及对齐程度）的变化调节癌细胞接触引导的假设；(3)验证促进粘附或去粘附的I型胶原结合蛋白调节癌细胞接触引导的假设。胶原纤维在云母上的外延生长以及凝胶中胶原纤维的磁性排列将用于设计具有特定接触引导特性的环境。细胞迁移将在2D， 3D和混合环境中使用活细胞显微镜进行评估。了解不同性质的胶原纤维网络如何调节接触引导，将进一步完善诊断活检图像的预后能力。胶原纤维密度的量化，活检的机械性能的测量，交联密度的代理，以及额外的胶原结合伙伴的染色将深入了解接触引导远离单个肿瘤的效率。未来的工作将着眼于在小鼠肿瘤模型的体内和活组织检查中进行这些测量，并将这些生物物理和组成特性与肿瘤进展或预后联系起来。

DESCRIPTION (provided by applicant): It has been long appreciated that changes in the extracellular matrix (ECM) in the tumor microenvironment can drive tumor progression. With the recent advance in imaging techniques changes in the organization of ECM components like collagen have been described. Circumferentially organized collagen fibers are reorganized into radially aligned fibers, leading to directed migration or contact guidance away from the tumor. This collagen organizational signature has been proposed as a diagnostic indicator of potential invasion, linking contact guidance to invasion. Unfortunately, while there is an abundance of information on contact guidance in 2D environments, much less is known about contact guidance in 3D environments. In addition, collagen fiber networks can exhibit a large variation in properties separate from alignment that regulate the ability of cells to sense and respond to contact guidance cues, effectively altering the relationship between collagen organization and invasion. My long-term goal is to understand how tumor, stromal and immune cells integrate multiple cues for directional migration in the tumor microenvironment by using various engineering approaches. The objective of this research is to understand the role of the organization and composition of collagen I matrix in directing contact guidance. The specific aims of this proposal include: (1) Assess the relative contributions of topology and confinement in explaining the differences between contact guidance in 2D and 3D environments, (2) Test the hypothesis that changes in fiber structure such as fiber and crosslinking density as well as degree of alignment regulate the contact guidance of cancer cells and (3) Test the hypothesis that collagen I binding proteins that promote adhesion or de-adhesion regulate the contact guidance of cancer cells. Epitaxial growth of collagen fibers on mica as well as magnetic alignment of collagen fibers in gels will be used to engineer environments with specify contact guidance characteristics. Cell migration will be assessed using live cell microscopy in 2D, 3D and hybrid environments. Understanding how different properties of collagen fiber networks regulate contact guidance will further refine the prognostic ability of diagnostic biopsy images. Quantification of collagen fiber density, measurement of mechanical properties of the biopsy, a proxy for crosslinking density, and staining of additional collagen binding partners will give insight into the efficiency of contact guidance away from an individual tumor. Future work will be geared towards making these measurements in vivo and in biopsies from mouse tumor models and correlating these biophysical and compositional properties to tumor progression or prognosis.