Controllable Rigidity Surfaces for T Cell Mechanobiology

 描述：机械力在指导T细胞活化和随后的功能中发挥着越来越多的作用。我们最近发现，T细胞是敏感的刚性的基础基板提出活化配体的CD 3和CD 28。具体而言，原代小鼠CD 4 + T细胞表现出随着材料刚性增加而在呈递针对CD 3和CD 28的抗体的表面上增加的活化。虽然在其他细胞系统中建立，但主要是在整合素-ECM和钙粘蛋白-钙粘蛋白相互作用的背景下，T细胞通过CD 3/TCR和CD 28的机械传感尚未得到很好的理解。解决这一知识差距将提供一个新的系统，在此基础上可以测试其他系统中开发的机械传感概念，并提供对T细胞生理学和免疫治疗先进工具的新见解。 这项研究旨在解决T细胞-底物相互作用像其他细胞一样发生在多个阶段的问题。在每个阶段控制底物刚性的能力将揭示每个阶段在机械传感中的作用，将随后的研究集中在负责这种能力的分子途径中。这些知识可以为复制或改进软基质的益处提供新的目标，但使用化学或生物制剂。为了实现这一目标，我们提出了一个磁致动系统，提供按需，可逆的，可重复的控制机械刚度呈现给贴壁细胞。我们首先专注于开发和验证这个新系统，这是一个适应的弹性体支柱阵列技术用于牵引力显微镜。然后，该系统用于独立评估早期细胞伸展和持续收缩期间的T细胞机械感知。在一个互补的方向，我们还将评估动态的CD 3/TCR信号作为基板刚性的函数，在T细胞mechanosensing的发展模型的核心概念。 这项研究的成功完成将是T细胞机械生物学新兴跨学科领域的一个关键进展。此外，期望所提出的系统将立即和直接地应用于其它蜂窝系统。

 DESCRIPTION: Mechanical forces play increasingly recognized roles in directing T cell activation and subsequent function. We recently discovered that T cells are sensitive to the rigidity of an underlying substrate presenting activating ligands to CD3 and CD28. Specifically, primary mouse CD4+ T cells exhibited increased activation on surfaces presenting antibodies to CD3 and CD28 as material rigidity increased. While established in other cell systems, predominantly in the context of integrin-ECM and cadherin-cadherin interactions, mechanosensing by T cells through the CD3/TCR and CD28 is not well understood. Addressing this gap in knowledge would provide a new system upon which the mechanosensing concepts developed in other systems could be tested, as well as provide new insight into T cell physiology and advanced tools for immunotherapy. The proposed study seeks to address the issue that T cell-substrate interaction, like other cells, occurs in multiple stages. The ability t control the rigidity of the substrate during each stage would reveal the role each one has in mechanosensing, focusing subsequent studies into the molecular pathways responsible for this ability. Such knowledge could provide new targets for replicating or improving upon the benefits of soft substrates, but using chemical or biological agents. Towards this goal, we propose a magnetically actuated system that provides on-demand, reversible, and repeatable control over the mechanical stiffness presented to an adherent cell. We first focus on development and validation of this new system, which is an adaptation of the elastomer pillar array technology used for traction force microscopy. This system is then used to independently assess T cell mechanosensing during early cell spreading and sustained contraction. In a complementary direction, we will also assess dynamics of CD3/TCR signaling as a function of substrate rigidity, a core concept in a developing model of T cell mechanosensing. Successful completion of the proposed study will be a key advance in the emerging interdisciplinary field of T cell mechanobiology. In addition, it is expected that the proposed system will be immediately and directly applicable towards other cellular systems.