Probing Contact Guidance on Exquisitely Tunable ECM Surfaces

描述(由申请人提供)：长期以来，人们一直认为肿瘤微环境中细胞外基质(ECM)的变化可以推动肿瘤的进展。随着成像技术的最新进展，细胞外基质成分如胶原的组织结构的变化已被描述。周向排列的胶原纤维被重组为放射状排列的纤维，导致定向迁移或接触引导远离肿瘤。这种胶原蛋白的组织特征被认为是潜在侵袭性的诊断指标，将接触指导与侵袭性联系起来。不幸的是，虽然有大量关于2D环境中的接触指导的信息，但关于3D环境中的接触指导的信息要少得多。此外，胶原纤维网络的性质可以表现出很大的变化，不同于调节细胞感知和响应接触指导信号的能力的排列，有效地改变了胶原组织和侵袭之间的关系。我的长期目标是了解肿瘤、间质和免疫细胞如何通过使用各种工程方法在肿瘤微环境中整合多个定向迁移的线索。本研究的目的是了解I型胶原基质的组织和组成在指导接触指导中的作用。该建议的具体目的包括：(1)评估拓扑和限制在解释2D和3D环境中接触指导之间的差异方面的相对贡献；(2)检验纤维结构(如纤维和交联度)的变化以及排列程度调节癌细胞接触指导的假设；以及(3)测试促进粘连或去粘连的I型胶原结合蛋白调节癌细胞接触指导的假说。胶原蛋白纤维在云母上的外延生长以及胶原蛋白纤维在凝胶中的磁性排列将用于设计具有特定接触引导特性的环境。细胞迁移将使用活细胞显微镜在2D、3D和混合环境中进行评估。了解胶原纤维网络的不同特性如何调节接触指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、活检组织机械性能的测量、交联物密度的替代以及额外的胶原结合伙伴的染色将使我们深入了解接触指导远离单个肿瘤的效率。未来的工作将是在体内和小鼠肿瘤模型的活检中进行这些测量，并将这些生物物理和成分特性与肿瘤进展或预后相关联。

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.