Mechanisms of mesenchymal chemotaxis

描述（由申请人提供）：细胞在各种情况下在称为趋化性的过程中向可溶性化学线索迁移。尽管近世纪的研究，趋化性的机械基础仍然没有完全理解。生长因子的空间梯度指导组织中间充质细胞的运动以协调和加速生理学上重要的过程，例如伤口愈合，并且间充质趋化性已经涉及病理状况，例如心血管和纤维化疾病。然而，绝大多数趋化性研究都集中在白细胞和其他快速移动的变形虫细胞上。间充质趋化性一直难以研究，因为它需要维持稳定的梯度许多小时。传统的方法，如transwell测定提供很少或没有动态信息，并从实际的方向感测运动的效率差的区别效果。为了克服这些技术限制，我们最近建立了一种微流控趋化性测定，允许在稳定的线性梯度中直接观察间充质细胞数小时，允许单细胞跟踪和高分辨率活细胞成像方法，如TIRF显微镜。我们的初步数据表明，生长因子受体PDGF-R通过PLC > PKC >肌球蛋白II途径控制间充质趋化性，并需要信号事件和细胞骨架组织的协调。我们建议通过以下方式阐明间充质趋化性的机制：1）剖析间充质细胞中趋化性信号传导的时空性质; 2）理解这种细胞类型中趋化性过程中细胞骨架的动态组织; 3）描绘信号传导和细胞骨架事件的协调，这些事件导致复杂的趋化性行为，例如在3D环境中重新定向到新的线索和趋化性。这些研究将直接有助于我们理解肿瘤转移、纤维化和心血管疾病等疾病状态的生理基础，以及我们对伤口愈合等生理过程的理解。

DESCRIPTION (provided by applicant): Cells in a variety of contexts migrate towards soluble chemical cues in a process known as chemotaxis. Despite nearly a century of study, the mechanistic underpinnings of chemotaxis remain incompletely understood. Spatial gradients of growth factors direct the movements of mesenchymal cells in tissues to coordinate and accelerate physiologically important processes such as wound healing, and mesenchymal chemotaxis has been implicated in pathological conditions such as cardiovascular and fibrotic diseases. Yet, the vast majority of chemotaxis studies have focused on leukocytes and other fast-moving, amoeboid cells. Mesenchymal chemotaxis has been prohibitively difficult to study, because it requires maintenance of stable gradients for many hours. Traditional methods such as transwell assays provide little or no dynamic information and poorly discriminate effects on the efficiency of motility from actual directional sensing. To overcome these technical limitations we recently established a microfluidic chemotaxis assay that allows direct observation of mesenchymal cells in stable, linear gradients over many hours, allowing both single-cell tracking and high-resolution live-cell imaging approaches such as TIRF microscopy. Our preliminary data indicate that the growth factor receptor, PDGF-R controls mesenchymal chemotaxis by a PLC > PKC > Myosin II pathway and requires the coordination of signaling events and cytoskeletal organization. We propose to elucidate the mechanisms of mesenchymal chemotaxis by 1) Dissecting the spatio- temporal nature of chemotactic signaling in mesenchymal cells 2) Understanding the dynamic organization of the cytoskeleton during chemotaxis in this cell type and 3) Delineating the coordination of signaling and cytoskeletal events that lead to complex chemotactic behaviors such as re-orientation to new cues and chemotaxis in 3D environments. These studies will directly contribute to our understanding the physiological basis of disease states such tumor metastasis, fibrosis and cardiovascular disease, as well as our understanding of physiological processes such as wound healing.