Collaborative Research: Mechanobiology of Fiber Geometry-RhoGTPase Crosstalk at the Leading Edge of Cells Crawling on Fibers

细胞在由纤维蛋白组成的复杂的三维环境中运动。细胞选择移动方向的第一步是通过拖拽纤维来感知环境。这就开始了所谓的“细胞收缩”。在正常或疾病组织中，如何在随机分布或以特定模式分布的不同直径的纤维上实现收缩，大多是未知的。了解细胞和接触的纤维之间的第一次感觉相互作用对于细胞如何侵入纤维组织非常重要，例如当癌细胞离开肿瘤时，或者在纤维上的细胞定向向伤口部位迁移时。我们对细胞迁移的大部分了解来自于在2D平板上进行的经典研究，或者最近使用复杂的凝胶。这两种方法都不允许研究细胞与纤维的相互作用。这项研究将解决量化细胞-纤维相互作用的需要，以了解伤口愈合和疾病过程中的细胞迁移。在这项合作研究中，通过结合纳米纤维制造、细胞信号生物传感器和细胞力学的最新技术，我们将能够看到细胞内部，并确定帮助细胞沿纤维迁移的决策机制。PI将与当地社区大学的教职员工合作，在实验室对他们进行培训，以便他们能够开发教育材料。这将为社区学院的学生创造研究机会。这个项目将定义细胞与纤维相互作用时的机械生物学状态。具体地说，当细胞形成突起并在纤维上迁移时，我们将揭示RhoGTPase信号(RhoA、rac1和CDC42)。这些分子在细胞特定区域的相互作用、定位和总和定义了迁移模式，这在2和3D中已被证明是不同的。为了确定细胞中这些蛋白质在纤维上的活性图谱，我们将设计不同直径(纳米-微米)的纤维网络，这些纤维网络以排列和随机的配置分布，表示亲和抗侵袭条件。通过这样做，我们将能够精确地定位RhoGTP酶的空间和时间激活图，因为细胞拖拽并对纤维施加力。机械生物力量化-生物传感器激活将在细胞迁移到不断变化的纤维环境的可塑性方面发展新的知识，就像在体内遇到的那样。将跨越广泛空间(纳米-微米)和时间(秒-小时)尺度的基于黏附受体的信号与迁移和力调节联系起来，将在入侵驱动的细胞迁移方面提供新的知识，从而在药物发现和开发方面开辟新的方向。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

Cells move in a complex three-dimensional environment composed of fibrous proteins. A first step for a cell in choosing a direction to move is to sense its environment by tugging at the fibers. This starts what is called 'cell contractility.' How contractility is achieved on fibers of varying diameters distributed randomly or in specific patterns, in normal or diseased tissues, is mostly unknown. Understanding the first sensory interactions between a cell and a contacted fiber is important to how cells invade a fibrous tissue, such as when cancer cells leave a tumor, or in directed migration of cells on fibers towards a wound site. Most of what we know about cell migration stems from classical studies conducted on 2D flat substrates or, more recently, using complex gels. Neither approach allows the studying of cell-fiber interactions. This research will address the need to quantify cell-fiber interactions to understand cell migration during wound healing and disease. In this collaborative research, by combining state-of-art technologies in nanofiber manufacturing, cell signaling biosensors, and cell mechanics, we will be able to see inside the cell and determine the decision mechanisms that help cells migrate along fibers. The PI will work with faculty at a local community college to train them in the laboratory so that they can develop educational materials. This will create research opportunities for community college students.This project will define the mechanobiological state of a cell as it interacts with fibers. Specifically, we will reveal RhoGTPase signaling (RhoA, Rac1, and CdC42) as cells form protrusions and migrate on fibers. The interplay, localization, and summation of these molecules at specific regions of the cell define the mode of migration, which has been shown to be different in 2 and 3D. To determine the activity maps of these proteins in cells on fibers, we will design fiber networks of varying diameters (nanometers-microns) distributed in aligned and random configurations signifying pro- and anti-invasive conditions. In doing so, we will be able to pinpoint spatial and temporal activation maps of RhoGTPase's as cells tug and exert forces on fibers. The mechanobiological force quantitation-biosensor activation will develop new knowledge in the plasticity of cell migration to changing fibrous environments, as would be encountered in vivo. Linking adhesion receptor-based signaling spanning a wide range of spatial (nanometers-microns) and temporal (seconds-hours) scales with migration and force modulation will provide new knowledge in invasion-driven cell migration, thus opening new directions in drug discovery and development.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.