利用3D生物打印技术构建类软骨多级结构功能化仿生支架在软骨再生中的应用

Articular cartilage injury, which is the most common disease in sports medicine, is very hard to be cured. As the speeding development especially in rencent decades, tissue engineering (TE) technology has been a most promising way for cartilage regeneration, since cartilage can hardly regenerate by itself. Articular cartilage is a highly anisotropic tissue whose composition and organization varies greatly with depth. Traditional methods in material science can hardly re-construct the complexity of cartilage precisely and effectively. And the emerging and developing of the 3D bio-printing technology gave us a powful tool to solve this problem. In the present study, based on the previous results, we first planned to synthesize a brand new bio-ink using hyaluronic acid (HA) and silk fibroin mixed hydrogel, and then make modification on this hydrogel with mesenchymal stem cells (MSCs) affinity peptide, in order to let this hydrogel possess the function of recruiting endogenous MSCs. After this, we will use this bio-ink to print a HA-SF multi-layer funtional hydrogel biomimetic scaffold via 3D bio-printing technology for the cartilage regeneration. In the printing process, we will also add BMP-2 and TGF-β3 in the different layer for the chondrogenesis and the interface healing of subchondral bone and cartilage. Finally, we will construct a biomimetic functional multi-layer scaffold for the cartilage regeneration, which can simulate the normal cartilage structure, recruit endogenous MSCs, and provide a promising microenviroment for the cartilage regeneration. In this study, we will also investigate the detail mechanism of the MSCs affinity peptide and the microenviroment changing during the whole regeneration process.

关节软骨损伤是运动医学领域的难治性伤病。组织工程技术的出现和不断发展，为像关节软骨这样自我修复能力差的组织再生带来了希望。关节软骨具有多级分层的复杂结构，传统的材料学方法难以精确且高效地模拟和构建其结构，而近些年3D生物打印技术迅速发展为此提供了一个强有力的工具。本研究首先在先前研究的基础上，合成透明质酸（HA）-蚕丝蛋白（SF）生物墨水，并利用间充质干细胞（MSCs）亲和多肽对其进行修饰，使其具有募集内源性MSCs的能力。随后采用3D生物打印技术，以这种功能化的生物墨水为原料，构建一个模拟正常关节软骨结构的HA-SF多级分层功能化水凝胶仿生支架，并在打印过程中分层加入BMP-2和TGF-β3来诱导成软骨以及与软骨下骨的界面愈合，最终获得一个模拟正常软骨结构、能够募集内源性MSCs、且能够为软骨再生提供一个良好微环境的功能化水凝胶支架。并对亲和多肽作用机制和再生过程中微环境的改变进行阐明。

关节软骨损伤是运动医学领域的难治性伤病。正常的关节软骨具有多级分层的复杂结构，传统的材料学方法难以精确且高效地模拟和构建其复杂的结构。本研究利用3D打印技术逐层打印和构建一个模拟正常关节软骨结构的丝素蛋白-透明质酸多级分层功能化水凝胶仿生支架，并利用间充质干细胞亲和多肽（E7）对其进行修饰，使其具 有募集内源性间充质干细胞的能力，最终获得一个模拟正常软骨结构、能够募集内源性间充质干细胞、且能够为软骨再生提供一个良好的微环境的功能化水凝胶支架。并对亲和多肽作用机制以及再生过程中微环境的改变进行阐明。.本项目最终通过构建葡苷聚糖微球这种本身不具有细胞黏附能力的材料作为E7多肽载体，验证了E7对于间充质干细胞的亲和能力，并且发现E7通过LncRNA H19/miR675表观调控轴促进BMSC向软骨细胞分化，初步阐明了E7多肽对于间充质干细胞粘附、增殖以及分化的作用机制。随后，通过3D打印技术成功构建了基于天然材料丝素蛋白和透明质酸的水凝胶生物仿生支架用于关节软骨修复。研究结果表明，所构建的支架具有类软骨的多层多孔结构以及良好的生物力学强度，并且通过利用干细胞亲和多肽E7对水凝胶支架进行修饰，提升了募集自体干细胞的能力，为软骨再生提供了一个有利于细胞迁移、增殖以及分化的微环境。最终通过动物实验证明了该支架具有优越的关节软骨修复能力，将来具有非常良好的临床应用前景。