Department of Chemistry, Stanford University, Stanford, CA 94305, United States; Department of Materials Science and Engineering, Stanford University, Stanford, CA 94305, United States.
Department of Mechanical Engineering, Stanford University, 452 Escondido Mall, Stanford, CA 94305, United States.
Biomaterials. 2018 Feb;154:213-222. doi: 10.1016/j.biomaterials.2017.11.004. Epub 2017 Nov 6.
The physical and architectural cues of the extracellular matrix (ECM) play a critical role in regulating important cellular functions such as spreading, migration, proliferation, and differentiation. Natural ECM is a complex viscoelastic scaffold composed of various distinct components that are often organized into a fibrillar microstructure. Hydrogels are frequently used as synthetic ECMs for 3D cell culture, but are typically elastic, due to covalent crosslinking, and non-fibrillar. Recent work has revealed the importance of stress relaxation in viscoelastic hydrogels in regulating biological processes such as spreading and differentiation, but these studies all utilize synthetic ECM hydrogels that are non-fibrillar. Key mechanotransduction events, such as focal adhesion formation, have only been observed in fibrillar networks in 3D culture to date. Here we present an interpenetrating network (IPN) hydrogel system based on HA crosslinked with dynamic covalent bonds and collagen I that captures the viscoelasticity and fibrillarity of ECM in tissues. The IPN hydrogels exhibit two distinct processes in stress relaxation, one from collagen and the other from HA crosslinking dynamics. Stress relaxation in the IPN hydrogels can be tuned by modulating HA crosslinker affinity, molecular weight of the HA, or HA concentration. Faster relaxation in the IPN hydrogels promotes cell spreading, fiber remodeling, and focal adhesion (FA) formation - behaviors often inhibited in other hydrogel-based materials in 3D culture. This study presents a new, broadly adaptable materials platform for mimicking key ECM features of viscoelasticity and fibrillarity in hydrogels for 3D cell culture and sheds light on how these mechanical and structural cues regulate cell behavior.
细胞外基质 (ECM) 的物理和结构线索在调节细胞的重要功能方面起着关键作用，如扩散、迁移、增殖和分化。天然 ECM 是一种复杂的粘弹性支架，由各种不同的成分组成，这些成分通常组织成纤维状的微观结构。水凝胶经常被用作 3D 细胞培养的合成 ECM，但由于共价交联，它们通常具有弹性和非纤维状。最近的工作揭示了粘弹性水凝胶中的应力松弛在调节扩散和分化等生物学过程中的重要性，但这些研究都利用了非纤维状的合成 ECM 水凝胶。迄今为止，在 3D 培养中，只有在纤维状网络中才观察到关键的机械转导事件，如焦点黏附的形成。在这里，我们提出了一种基于 HA 与动态共价键交联的互穿网络 (IPN) 水凝胶系统，该系统模拟了组织中 ECM 的粘弹性和纤维状。该 IPN 水凝胶在应力松弛中表现出两个明显的过程，一个来自于胶原蛋白，另一个来自于 HA 交联动力学。通过调节 HA 交联剂亲和力、HA 的分子量或 HA 浓度，可以调节 IPN 水凝胶中的应力松弛。IPN 水凝胶中的更快的松弛促进了细胞扩散、纤维重塑和焦点黏附（FA）的形成——这些行为在 3D 培养中的其他水凝胶基材料中经常受到抑制。这项研究提出了一种新的、广泛适用的材料平台，用于模拟 3D 细胞培养中水凝胶中粘弹性和纤维状的关键 ECM 特征，并阐明了这些机械和结构线索如何调节细胞行为。
