外泌体功能化的仿ECM凝胶介导增殖/抗炎修复软骨缺损

Cartilage defects and degeneration highly associated with osteoarthritis are commom in orthopedics. Tissue engineering represents a promising method to solve this problem. In our previous study, dextran-gelatin hydrogels and 3D printed nanocomposite scaffolds based from gelatin nanoparticles were prepared for application in cartilage tissue engineering. Simultaneously, synovium-derived mesenchymal stem cells and induced pluripotent stem cells were also evaluated for application in tissue engineering. Compared with the cell-based therapies above, the cell-free therapy shows lower risks in exogenous cell proliferation, differentiation, mutation or other issues. Therefore, the cell-free therapy for cartilage regeneration will be proposed in this study. We propose to fabricate exosome-functionalized ECM-like hydrogels using three dimensional additive manufacturing technology for cartilage repair, which may facilitate cartilage regeneration and modulate immune reactivity. Subsequently, the cellular processes modulated by iPSC exosomes will be further studied and the mechanism of action underlying the exosome-mediated responses in cartilage repair will be illustrated. The expression of PI3K-Akt pathway related factors in the process of cartilage regeneration will be analyzed to explore the molecular mechanism of chondrocyte proliferation and inhibition of apoptosis. Then the immune reactivity modulated by iPSC exosomes through NF-κB signaling and the inhibition of cartilage extracellular matrix degradation through MAPKs pathway will be investigated to reveal the molecular mechanism of exosome-mediated cartilage dynamic balance. Together, the aim of this study is to confirm the role of exosome-functionalized ECM-like hydrogels in the process of cartilage repair and clarify the biological molecular mechanism of chondrocyte proliferation and immune reactivity.

关节软骨缺损及退变是一种常见的骨科疾病，常伴随骨关节炎的发生。组织工程为软骨修复提供了基础。在之前的研究中，本项目组制备了用于软骨组织工程再生的葡聚糖-明胶水凝胶和明胶纳米微球基3D打印支架。同时，本项目组也培养了滑膜间充质干细胞和诱导多潜能干细胞用于组织修复。与细胞策略相比，无细胞疗法不存在外源性细胞增殖、分化、突变等问题，具有更低的安全风险。因此，本研究提出无细胞的组织再生策略，拟构建外泌体功能化的仿ECM凝胶用于修复软骨缺损，既考虑促进软骨再生，也考虑调控炎症反应。同时，我们希望探究iPSC外泌体在软骨修复过程中的双重作用，一方面调控PI3K/Akt信号通路促进软骨细胞增殖、抑制凋亡，另一方面通过NF-κB信号减少炎症反应，调控MAPKs信号通路抑制细胞外基质降解，恢复和维持软骨动态平衡。最终证实外泌体功能化的仿ECM凝胶用于修复软骨缺损的应用价值，并明确其修复软骨的生物学分子机制。

关节软骨缺损及退变是一种常见的骨科疾病，常伴随骨关节炎的发生。组织工程为软骨修复提供了基础。本项目组制备了用于软骨组织工程再生的葡聚糖-明胶水凝胶和明胶纳米微球基3D打印支架。同时，本项目组培养人成纤维细胞，通过病毒感染重编程诱导为多潜能干细胞（iPSC）。建立的iPSC表达多能性细胞表面标志物、体外可以多向诱导分化，体内向三个胚层诱导分化，可用于组织修复。与细胞策略相比，无细胞疗法不存在外源性细胞增殖、分化、突变等问题，具有更低的安全风险。我们用iPSC外泌体替代诱导多潜能干细胞，发挥等同于干细胞的巨大再生能力来促进软骨修复。构建的外泌体功能化的仿ECM凝胶用于修复软骨缺损，既促进软骨再生，也调控炎症反应。体外实验证实iPSC外泌体通过内吞途径将iPSC内容物水平传递到软骨细胞，包括功能蛋白、信使mRNA、miRNA等，进而调控下游PI3K/Akt和NF-κB，MAPKs信号通路，发挥增殖/抗炎双重作用促进软骨缺损的修复。实验证实外泌体功能化的仿ECM凝胶用于修复软骨缺损的应用价值，并明确其修复软骨的潜在传递机制。

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