Bio-Mechanics of Directional Migration of Leukocytes

 描述：先天和获得性免疫反应包括从血液中招募白细胞到感染和炎症部位。在到达该位置后，白细胞清除入侵者，并开始消化和修复受损组织的过程。然而，当身体不能适当地调节白细胞的募集时，炎症就会变成慢性的，导致不可逆转的组织损伤和功能丧失。 类风湿性关节炎、炎症性肠病、1型糖尿病和多发性硬化症都是由白细胞不受控制的募集引起的自身免疫性疾病。虽然许多研究致力于确定与白细胞募集有关的特定生化过程的级联，但对推动白细胞迁移的机械事件，特别是它们如何产生必要的牵引力以穿过血管壁并进一步穿越三维血管外空间的了解要少得多。因此，本研究的主要目的是通过使用新的3D傅立叶牵引力显微镜(3DFTFM)技术以及遗传和药物操作，提供迫切需要的补充信息，将特定的细胞分子过程(即黏附动力学、肌动蛋白周转和肌球蛋白II收缩)与调节白细胞外溢及其随后在血管外组织中定向迁移的细胞力的产生联系起来。为了实现这一目标，我们提出了三个具体目标。我们将首先描述白细胞在平面上爬行所产生的三维牵引力的时空特征(目标1)；然后我们将研究调节跨越血管内皮单分子层和基底膜的迁移的机械过程(目标2)；最后，我们将开发一种新的弹性成像3DTFM来确定细胞外基质的牵引力和非线性材料属性，以阐明调控白细胞在三维环境中趋化迁移的机制的分子机制(目标3)。建议的体外方法克服了测量驱动白细胞外溢和迁移的3-D牵引力的许多现有挑战，并建立在我们由生物学家和工程师组成的多学科团队积累的广泛经验基础上，他们在过去七年里一直在研究阿米巴细胞迁移的机制。这项研究的结果将导致对白细胞运动机制的更全面的了解，并将有可能帮助开发新的方法，这些方法可以针对特定的机械过程来抑制(或减缓)白细胞的运动，并有助于设计治疗炎症性疾病的补充方案。

 DESCRIPTION: The innate and adaptive immune response involves the recruitment of leukocytes from the blood stream to the site of infection and inflammation. Upon reaching the location, leukocytes clear invaders and begin the process of digesting and repairing damaged tissues. However, when the body fails to properly regulate the recruitment of leukocytes, the inflammation can become chronic, resulting in irreversible tissue injury and loss of functionality. Rheumatoid arthritis, inflammatory bowel disease, type-1 diabetes, and multiple sclerosis are all examples of autoimmune diseases caused by the uncontrolled recruitment of leukocytes. While much research has been dedicated to the identification of the cascade of specific biochemical processes involved in the recruitment of leukocytes, much less is known about the mechanical events driving their migration, in particular how they generate the necessary traction forces to cross the vascular wall and further traverse the three- dimensional (3-D) extravascular space. Thus, the main objective of this study is to provide the much needed complementary information connecting specific cell molecular processes (i.e., adhesion dynamics, actin turnover, and myosin II contraction) to the generation of cellular forces that regulate leukocyte extravasation and their subsequent directional migration in 3-D extravascular tissues through the use of novel 3D Fourier Traction Force Microscopy (3DFTFM) techniques and genetic and pharmacological manipulations. To achieve this objective, we propose three Specific Aims. We will first characterize the temporal and spatial generation of 3-D traction forces exerted by leukocytes crawling on flat surfaces (Aim 1); we will then investigate the mechanical processes regulating transmigration across the vascular endothelial monolayer and the basement membrane (Aim 2); and finally, we will develop a novel Elastographic 3DTFM to determine both traction stresses and the non-linear material properties of the Extra Cellular Matrix to elucidate the molecular mechanisms regulating the mechanics of leukocytes' chemotactic migration in 3-D environments (Aim 3). The proposed in vitro approach overcomes a number of existing challenges to measuring the 3-D traction forces driving leukocyte extravasation and migration and builds on the extensive experience accumulated by our multidisciplinary team of biologists and engineers who have been studying the mechanics of amoeboid cell migration for the last seven years. The outcome of this research will result in a far more comprehensive understanding of the mechanics of leukocyte motility than that available to date and will have the potential to aid the development of new approaches that could target specific mechanical processes to inhibit (or slow down) leukocyte motility and help in the design of complementary regimens to treat inflammatory diseases.