基于间充质干细胞源的外泌体miRNAs介导的TGF-β调控机制及其软骨修复作用研究

The technology of stem cell has a promising prospect in cartilage repairing. It is reported that exosomes derived from mesenchymal stem cells (MSC-Exos) may take a part of, even the whole, biological effects during the treatment. However, the specific effect and mechanism of MSC-Exos in cartilage repairing are unclear. Previous research showed that: 1. MiRNAs, such as miR-21, are enriched in exosomes and perform a variety of physiological effects. These miRNAs can regulate some key factors of TGF-beta signaling pathway, such as SMAD7. 2. TGF-beta signaling pathway plays an essential role in regulating proliferation and phenotype of chondrocytes. 3. It is reported that integrin-extracellular matrix mediates the distribution differences of exosomes among organs. Based on research findings and theoretical clues, we suppose that exosomes play the key role in cartilage repairing mediated by mesenchymal stem cells. Exo-miRNAs, represented by miR-21, achieve cartilage repair through the regulation of TGF-beta signaling pathway. We also infer that integrin-extracellular matrix mediates MSC-Exos engrafting in cartilage. This project intends to reveale the role of exosomes in the regulation of chondrocyte proliferation and phenotype in vivo and vitro, and to elucidate the effects and mechanisms of Exo-miRNAs in regulation of TGF beta signaling pathway. We expect to explore the molecular mechanism of MSC-Exos adhering to cartilage and to develop a new method for cartilage repairing.

干细胞技术在软骨修复中具有良好的应用前景。研究提示间充质干细胞来源的外泌体（MSC-Exos）可能承担了治疗中部分甚至全部生物效应，但其具体作用及机制尚不明确。研究显示：1.以miR-21为代表的miRNAs在外泌体中富集并发挥多种生理作用，此类miRNAs可以调控TGF-β通路下SMAD7等关键因子的表达；2.TGF-β通路在调控软骨细胞增殖和表型中发挥了重要作用；3.整合素-细胞外基质介导了外泌体在器官间的分布差异。由此推测：外泌体介导了MSC的软骨修复作用，并通过以miR-21为代表的miRNAs调控TGF-β通路促进软骨修复，而整合素-细胞外基质介导了MSC-Exos在软骨组织中的定植。本研究拟通过体内外实验揭示外泌体在调控软骨细胞增殖和表型中的作用，阐明Exo-miRNAs对TGF-β通路的调控作用与机制，并探究MSC-Exos向软骨组织定植的分子机制，从而开拓软骨修复的新方法。

骨关节炎（OA）是最常见的退行性关节疾病，涉及关节软骨的丧失、关节骨赘的形成、软骨下骨的重塑和滑膜炎。干细胞技术在软骨修复中具有良好的应用前景。研究提示间充质干细胞来源的外泌体（MSC-Exos）可能承担了治疗中部分甚至全部生物效应，但其具体作用及机制尚不明确。本课题通过以下几个方面进行深入研究：1）MSC-Exos在治疗软骨损伤中的作用；2）MSC-Exos miRNAs在治疗软骨损伤中的作用；3）MSC-Exos的全面蛋白组学分析及其在软骨损伤修复中的作用；4）MSC-Exos介导软骨修复作用的潜在机制；5）MSC-Exos蛋白及miRNA表达的精确调控促进软骨损伤修复的具体实践。通过研究，我们得出以下结果：1. MSC-Exos能够与软骨细胞高度融合，且呈浓度、时间梯度依赖性；2. 脂肪来源MSC-Exos在体外表现出更好的缓解炎症、减轻软骨细胞外基质降解和抑制凋亡的作用；3. 脂肪来源MSC-Exos在体内能够更大程度的改善关节软骨破坏，减轻滑膜炎症，改善胫骨软骨下骨的重塑，并减少软骨细胞的凋亡；4. 生物信息学分析揭示了MSC-Exos在细胞外基质受体中的共同功能。骨髓来源MSC-Exos具有更强的再生能力，脂肪来源MSC-Exos在免疫调节中发挥重要作用，而脐带来源MSC-Exos在组织损伤修复中的作用更为突出；5. MSC-Exos可能是通过降低软骨细胞中RIPK1的表达促进软骨损伤修复，从而缓解OA。总之，本研究通过体内外实验揭示外泌体在调控软骨细胞增殖和表型中的作用，并通过转录组及蛋白质组学对人类MSC来源的外泌体进行系统、全面的分析，从而开拓了软骨修复的新方法。