Department of Orthopaedics, The Fifth Affiliated Hospital of Sun Yat-Sen University, No. 52, Meihua East Road, Xiangzhou District, Zhuhai, 519000, Guangdong, China.
Department of General Surgery, The Second Affiliated Hospital of Nanchang University, Nanchang, 330000, Jiangxi Province, China.
Cell Commun Signal. 2022 May 30;20(1):75. doi: 10.1186/s12964-022-00881-9.
The transforming growth factor-beta (TGF-β) signaling pathway is an important pathway associated with the pathogenesis of osteoarthritis (OA). This study was to investigate the involvement of circRNAs in the TGF-β signaling pathway.
Cell Counting Kit-8 (CCK-8) assay and 5-ethynyl-2'-deoxyuridine (EdU) assay were used to detect the proliferation of primary mouse chondrocytes (PMCs). RNA-sequencing together with bioinformatics analysis were used to systematically clarify TGF-β1 induced alternations of circRNAs in PMCs. The regulatory and functional role of circPhf21a was examined in PMCs. Downstream targets of circPhf21a were explored by RNA-sequencing after overexpression of circPhf21a and verified by RT-qPCR in PMCs. Finally, the role and mechanism of circPhf21a in OA were explored in mouse models.
We found that TGF-β1 promoted the proliferation of PMCs. Meanwhile, RT-qPCR and western blotting indicated that TGF-β1 promoted extracellular matrix (ECM) anabolism. RNA-sequencing revealed that a total of 36 circRNAs were differentially expressed between PMCs treated with and without TGF-β1. Of these, circPhf21a was significantly decreased by TGF-β1. Furthermore, circPhf21a knockdown promoted the proliferation and ECM synthesis of PMCs, whereas overexpression of circPhf21a showed the opposite effects. Mechanically, the expression profiles of the mRNAs revealed that Vegfa may be the target of circPhf21a. Additionally, we found that circPhf21a was significantly upregulated in the mouse OA model, and inhibition of circPhf21a significantly relieved the progression of OA.
Our results found that TGF-β1 promoted the proliferation and ECM synthesis of PMCs via the circPhf21a-Vegfa axis, which may provide novel therapeutic targets for OA treatment. Video abstract.
转化生长因子-β（TGF-β）信号通路是与骨关节炎（OA）发病机制相关的重要通路。本研究旨在探讨 circRNAs 在 TGF-β信号通路中的作用。
使用细胞计数试剂盒-8（CCK-8）检测和 5-乙炔基-2'-脱氧尿苷（EdU）检测来检测原代小鼠软骨细胞（PMCs）的增殖。RNA 测序结合生物信息学分析系统地阐明了 TGF-β1 诱导 PMCs 中 circRNAs 的变化。在 PMCs 中检测 circPhf21a 的调节和功能作用。过表达 circPhf21a 后通过 RNA 测序探索 circPhf21a 的下游靶标，并在 PMCs 中通过 RT-qPCR 进行验证。最后，在小鼠模型中探索 circPhf21a 在 OA 中的作用和机制。
我们发现 TGF-β1 促进了 PMCs 的增殖。同时，RT-qPCR 和 Western blot 表明 TGF-β1 促进细胞外基质（ECM）合成。RNA 测序显示，TGF-β1 处理的 PMCs 中有 36 个 circRNAs 差异表达。其中，circPhf21a 显著受 TGF-β1 下调。此外，circPhf21a 敲低促进 PMCs 的增殖和 ECM 合成，而过表达 circPhf21a 则表现出相反的效果。从机制上讲，mRNA 的表达谱表明 Vegfa 可能是 circPhf21a 的靶标。此外，我们发现 circPhf21a 在小鼠 OA 模型中显著上调，抑制 circPhf21a 显著缓解 OA 的进展。
我们的结果发现，TGF-β1 通过 circPhf21a-Vegfa 轴促进 PMCs 的增殖和 ECM 合成，这可能为 OA 的治疗提供新的治疗靶点。
