生物活性水凝胶复合LIFR高表达MSC亚群重建关节软骨的作用及机制研究

Cartilage tissue can not heal itself once it is damaged. Although transplantation of mesenchymal stem cells (MSC) has been developed and applied gradually, its effect on cartilage repair is still limited. Our previous studies shown that cell proliferation and chondrogenic differentation potential were greatly enhanced in the MSC after chondrogenic inductive “differentiation-dedifferentiation” treatment (M-MSC), which then restored articular cartilage. Our pilot data of RNA-sequencing demonstrated that leukemia inhibitory factor receptor (LIFR) was significantly up-regulated in the M-MSC. And the stemness was remarkably enhanced in the LIFR highly expressed MSC sub-population after cell sorting in the M-MSC. We further found that the leukemia inhibitory factor (LIF) loaded bioactive hydrogel could significantly enhance the cell viability when combined with the MSC sub-population. However, the underlying mechanism and their effect on cartilage repair need to be clarified. In this study, molecular biological mechanism analysis, dose-response relationship and safety evaluation, hydrogel activity test, and cartilage defect model validation will be performed through the methods including genomic methylation sequencing, immunoblotting, biomechanical test, and gait analysis. We will elucidate the underlying mechanism of LIF binding to its receptor LIFR on the regulation of MSC stemness and evaluate the effect of bioactive hydrogel combined with LIFR highly expressed MSC sub-population on the reconstruction of articular cartilage. The results will help to offer scientific evidence for the cartilage regeneration by bioactive hydrogel combined with MSC sub-population.

软骨组织一旦受损难以自行修复。尽管间充质干细胞（MSC）移植技术逐步得到开发和应用，但对软骨修复作用有限。前期研究证实，经成软骨诱导“分化-去分化”处理的MSC（M-MSC）的增殖和成软骨分化能力显著增强，可重建关节软骨。预实验通过表达谱测序发现M-MSC中白细胞抑制因子受体（LIFR）表达增高，从M-MSC进一步筛选出的LIFR高表达的细胞亚群具有更强的干性；缓释白细胞抑制因子（LIF）的活性水凝胶复合该细胞亚群后能显著增强细胞活力，但具体机制及其对软骨修复作用有待阐明。我们拟通过分子生物学机制分析、量效关系和安全性评估、水凝胶活性测试、软骨缺损模型验证，采用全基因组甲基化测序、免疫印迹、生物力学、步态分析等方法，阐明LIF结合LIFR对MSC干性的调控机制，评价活性水凝胶复合高表达LIFR的MSC亚群重建关节软骨的效果。研究结果将为生物活性水凝胶复合MSC亚群实现软骨再生提供科学依据。

软骨组织一旦受损难以自行修复。尽管间充质干细胞(MSC)移植技术逐步得到开发和应用，但对软骨修复作用有限。前期研究证实，经成软骨诱导“分化-去分化”处理的MSC(M-MSC)的增殖和成软骨分化能力显著增强，可重建关节软骨。我们进一步实验通过表达谱测序发现M-MSC中白细胞抑制因子受体(LIFR)表达增高，从M-MSC进一步筛选出的LIFR高表达的细胞亚群具有更强的干性;缓释白细胞抑制因子(LIF)的活性水凝胶复合该细胞亚群后能显著增强细胞活力，但具体机制及其对软骨修复作用有待阐明。我们通过分子生物学机制分析、量效关系和安全性评估、水凝胶活性测试、软骨缺损模型验证，采用免疫印迹、骨组织影像学、组织形态学等方法，初步阐明LIF结合LIFR对MSC干性的调控机制，评价活性水凝胶复合高表达LIFR的MSC亚群重建关节软骨的效果。研究结果将为生物活性水凝胶复合MSC亚群实现软骨再生提供科学依据。