Department of Chemical and Biomolecular Engineering, University of Pennsylvania, Philadelphia, PA, USA.
Department of Pathology and Laboratory Medicine, Children's Hospital of Philadelphia, Philadelphia, PA, USA.
Ann Biomed Eng. 2018 Sep;46(9):1348-1361. doi: 10.1007/s10439-018-2041-7. Epub 2018 May 22.
Dendritic cell (DC) migration is required for efficient presentation of antigen to T cells and the initiation of an adaptive immune response. In spite of its importance, many aspects of DC migration have not been characterized. DCs encounter a variety of environments with different stiffness and geometry, but the effect of these parameters on DC migration has not yet been determined. We addressed this question by comparing DC motility on standard migration surfaces (polydimethylsiloxane (PDMS)-coated coverslips) and micropost array detectors (mPADs). These two surfaces differ in both stiffness and geometry. We found that DC migration was affected by substrate type, with significant increases in speed and significant decreases in persistence time on mPADs made of PDMS as compared to spin-coated PDMS coverslips. To determine whether the geometry or compliance of the post arrays was responsible for these changes in DC migration, we quantified DC motility on mPADs of identical geometry but different stiffness. Migration was indistinguishable on these mPADs, suggesting that DCs are responsive to geometry of ligand presentation and not stiffness. Further, by micropatterning ligands on flat PDMS surfaces in similar geometries to the mPAD arrays, we determined that DCs respond to the geometry of printed ligand. Finally, we used a variety of small molecule inhibitors to identify pathways involved in geometry sensing. We saw a significant role for myosin contractility and αβ integrin engagement. We also noted significant reorganization of the actin cytoskeleton into dynamic actin rings when DCs were motile on posts. From these experiments, we conclude that DCs are insensitive to substrate compliance in the range tested but respond to changes in geometry via a mechanism that involves integrin function, myosin contractility, and remodeling of the actin cytoskeleton. As a possible explanation, we postulate a consistent role for filopodial extension and contraction as the driver of DC motility.
树突状细胞 (DC) 的迁移对于有效呈递抗原给 T 细胞和启动适应性免疫反应至关重要。尽管其重要性不言而喻，但 DC 迁移的许多方面尚未得到充分描述。DC 会遇到具有不同硬度和几何形状的各种环境，但这些参数对 DC 迁移的影响尚未确定。我们通过比较标准迁移表面（聚二甲基硅氧烷 (PDMS) 涂层盖玻片）和微柱阵列检测器 (mPAD) 上的 DC 迁移来解决这个问题。这两种表面在硬度和几何形状上都有所不同。我们发现，DC 迁移受到基底类型的影响，与涂有 PDMS 的旋转涂覆 PDMS 盖玻片相比，mPAD 上的速度显著增加，持久性时间显著降低。为了确定柱阵列的几何形状或顺应性是否是导致 DC 迁移变化的原因，我们在具有相同几何形状但不同硬度的 mPAD 上量化了 DC 迁移。在这些 mPAD 上，迁移是无法区分的，这表明 DC 对配体呈递的几何形状有反应，而不是对硬度有反应。此外，通过在具有类似 mPAD 阵列的几何形状的平面 PDMS 表面上微图案化配体，我们确定了 DC 对印刷配体的几何形状有反应。最后，我们使用各种小分子抑制剂来鉴定参与几何形状感知的途径。我们发现肌球蛋白收缩性和αβ整合素结合在其中起着重要作用。当 DC 在柱上运动时，我们还注意到肌动蛋白细胞骨架的显著重组成动态肌动蛋白环。从这些实验中，我们得出结论，DC 在测试范围内对基底顺应性不敏感，但通过一种涉及整合素功能、肌球蛋白收缩性和肌动蛋白细胞骨架重塑的机制对几何形状的变化做出反应。作为一种可能的解释，我们假设丝状伪足的延伸和收缩作为 DC 迁移的驱动力发挥着一致的作用。
