环状RNA circPVT1调控肌腱干细胞衰老与分化的作用及机制研究

Tendon stem/progenitor cells (TSPCs) provide a potent cell source for regeneration and replacement of damaged tendons that are currently poorly responsive to more conventional treatments. However, the molecular mechanisms governing TSPC aging and tenogenic differentiation remain unclear at present. CircRNAs are believed to influence many cellular processes. Circular RNAs characterized by covalently closed loop structures without open terminals, are not sensitive to RNase R and more stable than linear RNA. Therefore, circular RNA has important research significance. Our previous results showed that the expression of circPVT1 is downregulated during aging of TSPCs. CircPVT1 siRNA transfection promoted senescence and inhibited tenogenic differentiation in early stage TSPCs. Moreover, we found that circPVT1 can modulate miR-140-5p activity, and miR-140-5p also can control circPVT1 levels. On this basis, we will further investigate the functions and mechanisms of circPVT1 in senescence and tenogenic differentiation of TSPCs combined with RIP, RNA pull-down, luciferase analysis and RNA FISH assays. Improved understanding of the mechanisms governing TSPCs aging and tenogenic differentiation may provide clues to effective targeted therapy.

肌腱来源的干细胞(Tendon stem/progenitor cells，TSPCs)在肌腱再生修复中显示出巨大的潜力，然而对于其衰老和向肌腱分化的分子机制并未阐明。环状RNA是细胞功能调控网络中重要的一员，其环状结构对RNase不敏感，导致它比线性RNA更为稳定，衰老中环状RNA的研究尚处于起步阶段，亟需找到在衰老中发挥重要功能的环状RNA。我们前期结果显示：环状RNA circPVT1在TSPCs衰老进程中表达显著下调，早期细胞中circPVT1敲除后明显促进TSPCs衰老，抑制了TSPCs成肌腱分化。我们发现circPVT1能与miR-140-5p结合进行相互调控。本研究将通过RNA结合蛋白免疫共沉淀、RNA pull-down、荧光素酶报告基因及原位杂交等技术，深入研究circPVT1调控TSPCs衰老及肌腱分化的作用及分子机制。为临床利用TSPSs治疗肌腱病提供新的思路及依据。

肌腱干/祖细胞(TSPC)衰老可导致年龄依赖性肌腱疾病，并破坏人类肌腱修复和替换能力。TSPC衰老和对不良因素敏感的机制是复杂的。在本研究中，我们分析了circPVT1参与TSPC衰老。circPVT1的表达在体外长时间培养下明显降低。circPVT1的下调促进了TSPC衰老进程，抑制了TSPCs的自我更新、迁移和成肌腱分化。此外，我们发现circPVT1直接靶向microRNA (miR)-199a-5p，从而减弱其对SIRT1表达的负调控。miR-199a-5p抑制或SIRT1过表达减弱了circPVT1敲低的衰老促进作用，这意味着circPVT1部分通过上调miR-199a-5p-SIRT1信号轴来抑制TSPC衰老。我们的研究结果最终解释了circPVT1在TSPC衰老调控中的主要作用；circPVT1是减少肌腱衰老的一种新的潜在治疗靶点。