基于自修复胶体凝胶的干细胞力学微环境研究

Cells interactions with the extracellular matrix (ECM) trigger numerous responses that have essential roles in the regulation of their behavior and fate. To develop artificial ECM that can mimic physiological cellular environment can open numerous new avenues for the regeneration of defected tissues/organs caused by trauma or disease. Here we propose to develop a class of self-healing colloidal hydrogels with tunable viscoelasticity which can mimic the mechanical environmental cues of natural ECM. By design and synthesis of nano-sized colloidal particles based on biocompatible polymer gelatin, colloidal networks composed of oppositely charged colloids can be generated, which we proposed to use as 3D matrix for stem cell culture. By fine-tuning the physicochemical properties of the colloids (including size, zeta potential and stiffness), the interparticle electrostatic interactions and the volume fraction, we are able to mediate the elasticity, viscosity as well as the 3D topography of the colloidal network. These changes of mechanical and structural properties of the gels can potentially affect stem cells behavior (including morphology, attachment, spreading, migration and proliferation) and their commitment to different cell linages. Furthermore, we propose to perform in-depth study of the underlying mechanism that how stem cells sense and response to matrix viscoelasticity. In general, this proposal can provide a new platform for the study of stem cell niche, and could provide paradigm shift for the development of the next generation of biomaterials.

构建高度仿生的细胞微环境是体外研究干细胞行为的基础，是组织工程和再生医学领域急需攻克的关键科学命题。本项目围绕构建模拟胞外基质粘弹性的材料平台来研究干细胞粘弹性响应的基础科学问题，提出自修复胶体凝胶作为三维细胞培养基质，基于胶体物理学原理，通过对胶体间作用势能、体积分数和网络分支局部硬度改变，调控胶体凝胶粘弹性（储存模量、损耗模量）；利用胶体凝胶自修复过程的结构粗化现象，调控凝胶网络三维拓扑结构；最终在多场耦合体系中，实现对单个信号的孤立调控，使精确探讨微环境力学信号对干细胞命运的影响成为可能；通过对粘弹性响应相关细胞信号通路研究，试图揭示干细胞粘弹性响应机理。项目涉及了生物材料、胶体物理、干细胞生物学、组织工程学等多学科交叉的前沿课题，将弥补生物材料领域对胶体凝胶系统性、机理性研究的不足，有望产生干细胞力学响应方面的新发现和新理论建立，具有重要的指导作用和实用价值。

细胞不但会感知并响应周围环境的化学信号，也会对物理信号，包括力学信号、微观拓扑结构等，做出识别和响应。尤其是近年来针对细胞外基质的力学信号（硬度、粘弹性参数）的研究成为细胞生物学和生物材料学领域研究的热点。本项目提出基于纳米颗粒可逆自组装构建的颗粒水凝胶网络设计，构建新型的、可用于研究细胞力学响应的培养平台。为此，本项目设计和开发出一系列可控力学信号的新型水凝胶作为干细胞微环境的研究平台，通过生物相容性好、带相反电荷明胶基纳米颗粒的自组装，构建自修复胶体凝胶，实现可三维尺度调控基质粘弹性和三维拓扑结构的参数精准控制。同时，我们为了避免三维加载的细胞受到剪切力的破坏，通过单细胞微凝胶包裹的微流控技术为细胞构建力学屏蔽“铠甲”，从而实现基于凝胶微球保护屏蔽的细胞加载技术。进一步，我们探究了力学和结构信号对细胞命运的影响。我们发现基质力学和结构信号对巨噬细胞为主的免疫细胞具有同样的决定性作用。我们首先发现硬基质相较于软基质更易引起巨噬细胞向趋炎症表型分化；进一步发现加载干细胞的软硬基质可进一步通过干细胞外泌调控炎症反应；同时我们还发现具有粘弹性的胶体凝胶相比于同样基质材料的块体凝胶会引起更弱的免疫反应。上述成果进一步在探究力学和拓扑结构对细胞分化行为的影响研究方面，也带来了对细胞微环境影响细胞命运机理的深入认识。项目执行期间以通讯作者身份发表科研论文15篇、申报发明专利10项，培养博士和硕士研究生6人，顺利完成了项目的指标。