软骨组织诱导性功能肽水凝胶支架的设计及在关节软骨损伤修复中的应用

Repair of aticular cartilage injury is one of the major clinical problem, but hope arises in a new treatment modality based on the development of tissue engineering technology. In this project, we proposed the use of peptide self-assembly technique to build cartilage-directed tissue engineering scaffold, comprising organically in biocompatibility, structure and functions to achieve the regeneration and reconstruction of damaged cartilage tissue. Two chondrogenic-directed functional peptides, in which one can mediate condensation of MSCs in early signal transduction to induce differentiation into chondrocytes direction and the other can transform TGF-beta1 to induce differentiation and proliferation of chondrocyte, were designed for supramolecular self-assembly into nanofibril hydrogels. The peptide hydrogels can be further proved in physical and chemical properties and self-healing capacity via phenylboronic acid - diol “click chemistry” reaction. In vitro culture of MSCs in hydrogels and in vivo implantation will be extensively studied to explore the chondrogenic differentiation and neocartilage formation. Moreover, the synergistically cartilage-induced differentiation mechanism will be explored to evaluate the activity of the functional peptide hydrogel scaffold, for constructing chondrogenic-directed functional materials in model and design method, and further promoting the effective repair of articular cartilage injury.

关节软骨损伤修复是临床的重大难题之一，组织工程技术的发展为软骨损伤修复提供了新的治疗模式和希望。本项目提出将材料的生物相容性、结构和功能有机地整合起来，利用多肽自组装技术构建软骨组织诱导性功能支架，以期实现对软骨损伤组织的再生和重建。我们拟设计、合成两条共组装的软骨组织诱导性功能肽分子，分别能介导间充质干细胞（MSCs）在早期的凝聚、信号传导进而向软骨方向分化，以及结合转化生长因子-beta1（TGF-beta1）诱导软骨细胞的分化和增殖。通过苯硼酸-二羟基“点击化学”反应改善多肽凝胶的物理、化学性能以及凝胶的自愈合能力。对功能肽凝胶体外诱导MSCs向软骨方向分化，以及植入体内促进新生软骨形成的修复效果进行系统、深入的研究。探索、评价功能肽凝胶支架协同诱导软骨分化的作用机制，建立软骨组织诱导性材料的模型和设计方法，为推进支架材料真正实现关节软骨损伤修复提供新的策略。

本项目设计、合成了一系列自组装及共组装的软骨组织诱导性功能肽分子，构建了基于苯硼酸-邻二羟基“点击化学”交联的共组装功能肽水凝胶，介导MSCs在早期的凝聚以及信号传导，诱导MSCs向软骨方向分化，促进软骨缺损的再生修复。通过固相多肽合成技术FMOC/tBu策略合成了5条功能肽分子，其纯度分别高于90%（DFP2和BFP2）和95%（DFP1，BFP1和CFP），分子量鉴定准确。DFPs和BFPs在近中性pH下可共组装形成基于苯硼酸-邻二羟基“点击化学”交联的水凝胶（C Gels），并能有效负载TGF-beta1，释放周期长达2周。进一步将软骨细胞，骨髓间充质干细胞（MSCs）与功能肽水凝胶共培养，结果表明，共组装功能肽水凝胶组对软骨细胞，骨髓间充质干细胞（MSCs）的增殖有明显促进作用，共组装功能肽水凝胶组中软骨细胞标记物I型胶原、II型胶原和糖胺聚糖（GAG）显著高于对照组，表明其能显著诱导MSCs向软骨分化。兔关节软骨缺损模型的体内修复初步结果表明，该水凝胶体内植入稳定性好，植入2周后显示有少量新生软骨形成，表明该功能肽水凝胶有利于促进新生软骨生成，长期植入的实验组存活状态良好。