Multifunctional Synthetic Microgels as Scaffolds to Control Protein-Cell Interactions and Extracellular Matrix Heterogeneity on the Nanoscale

该项目旨在将干细胞封装成合成的多功能微凝胶，该微凝胶可以模拟细胞外基质（ECM），以精确控制并从根本上了解细胞-基质，细胞-配体和细胞-细胞相互作用。作为实现这一目的的材料平台，我建议制备和使用带有环辛基和叠氮基团的多臂聚乙二醇（peg）。这些前体聚合物可以在活细胞存在的情况下通过细胞相容性菌株促进的叠氮-炔环加成交联从而包裹住它们。通过将这种交联化学与基于液滴的微流体相结合，可以制备单细胞负载微凝胶，并有效控制其单分散性、粒径和聚合物网络拓扑结构。后一个方面可以通过使用不同数量（四臂和八臂）和臂长的多臂聚乙二醇来解决，从而在纳米尺度上控制异质性和网络尺寸。那些对网络形成不是至关重要的聚合物臂可以用细胞粘附配体、生长因子或进一步的官能团进行修饰。为此，分别选择了四臂和八臂聚乙二醇；因此，配体的呈现也可以在纳米尺度上进行控制。这种模块化的材料工具包允许研究细胞外基质环境对干细胞分化、扩散和增殖的影响，具有前所未有的一致性。此外，微凝胶颗粒的使用允许通过组装单细胞负载微凝胶到更大的3D结构来研究细胞-细胞相互作用。通过这种方法，可以构建包含不同细胞类型的高功能组织。在此基础上，我设想建立体外造血模型，探索间充质干细胞和造血干细胞以及血管细胞和成骨细胞对造血干细胞生态位的影响。

The proposed project aims at the encapsulation of stem cells into synthetic multifunctional microgels that can mimic the extracellular matrix (ECM) to precisely control and fundamentally understand cell-matrix, cell-ligand, and cell-cell interactions. As a material platform for this purpose, I propose to prepare and use multi-arm poly(ethylene glycols) (PEGs) that bear cyclooctyne as well as azide groups. These precursor polymers can be crosslinked by the cytocompatible strain-promoted azide-alkyne cycloaddition in the presence of living cells thereby encapsulating them. By combining this crosslinking chemistry with droplet-based microfluidics, single-cell-laden microgels can be prepared with effective control of their monodispersity, particle size, and the polymer network topology. The latter aspect can be addressed by using multi-arm PEGs of a different number (four and eight) and length of arms, thereby controlling the heterogeneity as well as the network mesh size on the nanoscale. Those of the polymer arms that are not crucially needed for the network formation can be modified with cell adhesive ligands, growth factors or further functional groups. For this purpose, tetra- and octa-arm PEGs are chosen, respectively; as a result the ligand presentation can be controlled on the nanoscale, too. This modular material toolkit allows for studying the influence of extracellular matrix environments on stem cell differentiation, spreading, and proliferation with unprecedented consistency. Moreover, the use of microgel particles allows for the investigation of cell-cell interactions by assembling single-cell-laden microgels to larger 3D constructs. By this means, highly functional tissues that contain different cell types can be build up. Based hereon, I envisage to develop an in-vitro model for hematopoiesis to explore the influence of mesenchymal and hematopoietic stem cells as well as vascular cells and osteoblasts on the hematopoietic stem cell niche.