The Ohio State University, Department of Biomedical Engineering, 270 Bevis Hall 1080 Carmack Rd., Columbus, OH 43210, USA.
The Ohio State University, Department of Material Science and Engineering, 177 Watts Hall 2041 N. College Rd., Columbus, OH 43210, USA.
Acta Biomater. 2018 Apr 1;70:110-119. doi: 10.1016/j.actbio.2018.01.031. Epub 2018 Feb 2.
A cell's insoluble microenvironment has increasingly been shown to exert influence on its function. In particular, matrix stiffness and adhesiveness strongly impact behaviors such as cell spreading and differentiation, but materials that allow for independent control of these parameters within a fibrous, stromal-like microenvironment are very limited. In the current work, we devise a self-assembling peptide (SAP) system that facilitates user-friendly control of matrix stiffness and RGD (Arg-Gly-Asp) concentration within a hydrogel possessing a microarchitecture similar to stromal extracellular matrix. In this system, the RGD-modified SAP sequence KFE-RGD and the scrambled sequence KFE-RDG can be directly swapped for one another to change RGD concentration at a given matrix stiffness and total peptide concentration. Stiffness is controlled by altering total peptide concentration, and the unmodified base peptide KFE-8 can be included to further increase this stiffness range due to its higher modulus. With this tunable system, we demonstrate that human mesenchymal stem cell morphology and differentiation are influenced by both gel stiffness and the presence of functional cell binding sites in 3D culture. Specifically, cells 24 hours after encapsulation were only able to spread out in stiffer matrices containing KFE-RGD. Upon addition of soluble adipogenic factors, soft gels facilitated the greatest adipogenesis as determined by the presence of lipid vacuoles and PPARγ-2 expression, while increasing KFE-RGD concentration at a given stiffness had a negative effect on adipogenesis. This three-component hydrogel system thus allows for systematic investigation of matrix stiffness and RGD concentration on cell behavior within a fibrous, three-dimensional matrix.
Physical cues from a cell's surrounding environment-such as the density of cell binding sites and the stiffness of the surrounding material-are increasingly being recognized as key regulators of cell function. Currently, most synthetic biomaterials used to independently tune these parameters lack the fibrous structure characteristic of stromal extracellular matrix, which can be important to cells naturally residing within stromal tissues. In this manuscript, we describe a 3D hydrogel encapsulation system that provides user-friendly control over matrix stiffness and binding site concentration within the context of a stromal-like microarchitecture. Binding site concentration and gel stiffness both influenced cell spreading and differentiation, highlighting the utility of this system to study the independent effects of these material properties on cell function.
细胞的不可溶微环境对其功能的影响越来越受到重视。特别是，基质硬度和黏附性强烈影响细胞的铺展和分化等行为，但在纤维状基质样微环境中能够独立控制这些参数的材料非常有限。在目前的工作中，我们设计了一种自组装肽（SAP）系统，该系统可方便地控制具有类似基质细胞外基质微观结构的水凝胶中的基质硬度和 RGD（精氨酸-甘氨酸-天冬氨酸）浓度。在该系统中，RGD 修饰的 SAP 序列 KFE-RGD 和无义序列 KFE-RDG 可以相互替换，以在给定的基质硬度和总肽浓度下改变 RGD 浓度。硬度通过改变总肽浓度来控制，并且可以包含未修饰的基本肽 KFE-8 来进一步增加此硬度范围，因为其模量更高。通过此可调谐系统，我们证明了人骨髓间充质干细胞的形态和分化受三维培养中凝胶硬度和功能性细胞结合位点的影响。具体来说，在封装 24 小时后，只有在包含 KFE-RGD 的较硬基质中细胞才能展开。加入可溶性成脂因子后，较软的凝胶最有利于成脂分化，这可通过脂质空泡的存在和 PPARγ-2 表达来确定，而在给定硬度下增加 KFE-RGD 浓度则对成脂分化有负面影响。因此，这种三组分水凝胶系统允许在纤维状三维基质中系统地研究基质硬度和 RGD 浓度对细胞行为的影响。
细胞周围环境中的物理线索-例如细胞结合位点的密度和周围材料的硬度-越来越被认为是细胞功能的关键调节剂。目前，大多数用于独立调节这些参数的合成生物材料缺乏基质细胞外基质的纤维结构，这对天然存在于基质组织中的细胞可能很重要。在本文中，我们描述了一种 3D 水凝胶封装系统，该系统可在类似于基质的微观结构背景下方便地控制基质硬度和结合位点浓度。结合位点浓度和凝胶硬度都影响细胞的铺展和分化，突出了该系统用于研究这些材料特性对细胞功能的独立影响的实用性。
