图案化凝胶微反应器设计调控MSCs/软骨细胞的交互诱导与软骨再生

The synergistic induction between mesenchymal stem cells (MSCs) and chondrocytes is an effective mode for of cartilage regeneration, which can promote chondrogenic differentiation and cell proliferation. The synergy is highly correlated with cell density and ratio, as well as the cell spatial organization and contact mode within the micro-scope, while conventional tissue engineering technology is insufficient to construct these elements. Microgels fabricated by photomask-based stereolithography makes it available to precisely regulate the spatial organization and density of cells on the microscale. On the basis of this, we first synthesized photocrosslinkable alginate with bioactive peptide modification, and then, micropatterned hydrogels with spatially distributed MSCs and chondrocytes were generated. The alginate hydrogel here was constructed to mimic cartilage ECM to enhance the cell function expression. And the patterned microgel was designed to control the contact mode, spacing distribution and cell ratio between MSCs and chondrocytes, thus regulate the intercellular induction of the co-culture model. The design of microgel based co-culture model, which was similar to micro-reactor, easily established different cellular environment and cellular interactions, and accordingly directed cartilage regeneration via synergistic induction between MSCs and chondrocytes. Besides, this kind of hydrogel based micro-reactor can also be used as a micromodule to realize cartilage tissue construction from microcosmic to macroscopic by bottom-up assembling or 3D tissue printing.

MSCs与软骨细胞的交互诱导可促进细胞软骨分化和增殖，是软骨再生构建的有效模式。两种细胞的交互作用与细胞密度、比例及微区范围内的细胞间距、接触模式等高度相关，而常规组织工程技术难以实现这些因素的构建和调控。基于光掩膜设计的光交联微凝胶制造可精确调控细胞密度和微米尺度范围内的细胞空间分布。为此，本研究通过多肽改性等方法获得生物活性光交联海藻酸钠，制备装载MSCs和软骨细胞的图案化微凝胶，其中海藻酸凝胶模拟软骨细胞外基质促进细胞功能表达，图案化设计调控MSCs与软骨细胞的接触方式、间距、分布及细胞比例等，进而调控两种细胞间的相互诱导模式。这种微凝胶设计方便地构建了微空间中细胞环境及细胞间的不同作用模式，类似建立了MSCs与软骨细胞的微反应器，可有效调控MSCs/软骨细胞交互诱导的软骨再生。该微反应器还可作为微模块单元，进一步通过组装或3D组织打印技术实现从微观到宏观的软骨组织构建。

基于凝胶功能化分区结构设计，本项目探索了MSCs与软骨细胞交互诱导作用调控软骨分化与组织再生。针对MSCs成软骨分化的机理特点，重点围绕适宜于MSCs细胞功能表达的软骨诱导微环境构建，以及基于光交联技术的凝胶功能化分区结构精细调控展开了研究工作。以具有天然生物活性的胶原水凝胶为基础，通过甲基丙烯酰胺化改性赋予胶原共价交联的特性，显著提高了其力学性能，且光交联特性便于凝胶功能化分区结构的构建和调控。胶原生物活性的保留和力学性能的提升使得凝胶具备了抵抗细胞肌动蛋白介导的基质收缩能力，确保了MSCs细胞功能的正常表达，促进了MSCs成软骨分化、软骨基质分泌以及关节软骨修复。进一步通过光交联技术构建了凝胶功能化分区结构，设计优化了MSCs与软骨细胞交互诱导的凝胶微反应器。具有优良生物活性的光交联胶原水凝胶体系除可用于本研究的凝胶微反应器构建外，还可广泛用于细胞载体和组织工程，为再生医学提供新型功能化载体材料。同时，基于光交联的凝胶功能化分区微反应器，实现了微米尺度范围内交互作用模型的设计，具有重要的科学意义和广泛应用前景。