Ligand Anchorage and Materials Stiffness to Modulate Cell Adhesion Signals

细胞外微环境的物理化学核心影响活细胞的命运决定。我们的目标是调节配体锚定到材料表面，以剖析细胞黏附中主要外源信号的高度错综复杂的信号通路。因此，我们想要用一种相互独立的多参数方法来证明配体锚定相对于另外两个重要线索，即材料硬度和配体密度的特殊影响。配体锚定将从强烈的共价偶联调节为细胞黏附配体如纤维连接蛋白与聚丙烯酰胺水凝胶层最上面的非共价分子间逐渐减弱的相互作用，因为它允许调节黏附细胞的感受器作用力。在配体与水凝胶层的偶联过程中，可以调节配体的密度，而水凝胶的硬度可以通过其交联密度来控制。利用这样的材料平台，将通过牵引力细胞仪来评估生物物理细胞响应的大小和时间依赖性。生物物理特征将伴随着细胞内信号的生化量化，如蛋白质表达和磷酸化状态。解剖的生物物理和生化细胞反应之间的相互关系有望为细胞黏附信号和配体锚定的具体影响提供新的见解。这一结果有望为组织工程和再生治疗的细胞培养支架提供有针对性的材料设计。

Physicochemical Coes of the extracellular microenvironment aect fate decisions of living cells. We aim to modulate ligand anchorage to materials surfaces in order to dissect the highly convoluted signalling pathways of major exogenous cues in cell adhesion. Thereby we want to demonstrate the specic impact of ligand anchorage in relation to two other important cues, namely materials stiffness and ligand density in a multiparameter approach independently of each other. Ligand anchorage will be modulated from strong covalent coupling to gradually weaker non-covalent intermolecular interactions of cell adhesion ligands like fibronectin to the topmost surface of the polyacrylamide hydrogel layers as it allows to tune receptor forces of adherent cells. Ligand density is adjusted during ligand coupling to the hydrogel layers, while hydrogel stiffness can be controlled via its cross-linking density. Using such a materials platform the magnitude and time dependence of the biophysical cell response will be assessed in terms of traction force cytometry. The biophysical characterisation will be accompanied by biochemical quantification of intracellular signals, such as protein expression and phosphorylation state. The correlation of both the dissected biophysical and biochemical cell response to each other is expected to provide new insights in cell adhesion signalling and the specific impact of ligand anchorage. The results are envisioned to enable a targeted materials design of cell culture scaffolds for tissue engineering and regenerative therapies.