Table_1_Long Non-coding RNAs LOC100126784 and POM121L9P Derived From Bone Marrow Mesenchymal Stem Cells Enhance Osteogenic Differentiation via the miR-503-5p/SORBS1 Axis.DOCX

Long non-coding RNAs (lncRNAs) play pivotal roles in mesenchymal stem cell differentiation. However, the mechanisms by which non-coding RNA (ncRNA) networks regulate osteogenic differentiation remain unclear. Therefore, our aim was to identify RNA-associated gene and transcript expression profiles during osteogenesis in bone marrow mesenchymal stem cells (BMSCs). Using transcriptome sequencing for differentially expressed ncRNAs and mRNAs between days 0 and 21 of osteogenic differentiation of BMSCs, we found that the microRNA (miRNA) miR-503-5p was significantly downregulated. However, the putative miR-503-5p target, sorbin and SH3 domain containing 1 (SORBS1), was significantly upregulated in osteogenesis. Moreover, through lncRNA-miRNA-mRNA interaction analyses and loss- and gain-of-function experiments, we discovered that the lncRNAs LOC100126784 and POM121L9P were abundant in the cytoplasm and enhanced BMSC osteogenesis by promoting SORBS1 expression. In contrast, miR-503-5p reversed this effect. Ago2 RNA-binding protein immunoprecipitation and dual-luciferase reporter assays further validated the direct binding of miR-503-5p to LOC100126784 and POM121L9P. Furthermore, SORBS1 knockdown suppressed early osteogenic differentiation in BMSCs, and co-transfection with SORBS1 small interfering RNAs counteracted the BMSCs’ osteogenic capacity promoted by LOC100126784- and POM121L9P-overexpressing lentivirus plasmids. Thus, the present study demonstrated that the lncRNAs LOC100126784 and POM121L9P facilitate the osteogenic differentiation of BMSCs via the miR-503-5p/SORBS1 axis, providing potential therapeutic targets for treating osteoporosis and bone defects.

长链非编码RNA（long non-coding RNAs，lncRNAs）在间充质干细胞分化中发挥关键作用。然而，非编码RNA（non-coding RNA，ncRNA）网络调控成骨分化的具体机制仍不明确。因此，本研究旨在鉴定骨髓间充质干细胞（bone marrow mesenchymal stem cells，BMSCs）成骨过程中与RNA相关的基因及转录本表达谱。通过对骨髓间充质干细胞成骨分化第0天和第21天的样本开展转录组测序，筛选差异表达的ncRNA与信使RNA（mRNA），我们观察到微小RNA（microRNA，miRNA）miR-503-5p的表达显著下调。而其预测的miR-503-5p靶基因索宾与SH3结构域包含蛋白1（sorbin and SH3 domain containing 1，SORBS1）在成骨过程中表达显著上调。此外，通过长链非编码RNA-微小RNA-信使RNA相互作用分析及功能缺失与功能获得实验，我们发现长链非编码RNA LOC100126784与POM121L9P在细胞质中富集，并通过促进SORBS1的表达增强骨髓间充质干细胞的成骨分化能力；而miR-503-5p则可逆转这一效应。Ago2 RNA结合蛋白免疫沉淀实验与双荧光素酶报告基因实验进一步验证了miR-503-5p可直接结合LOC100126784与POM121L9P。此外，敲低SORBS1可抑制骨髓间充质干细胞的早期成骨分化，且共转染SORBS1小干扰RNA（small interfering RNAs，siRNAs）可抵消过表达LOC100126784与POM121L9P的慢病毒质粒所促进的骨髓间充质干细胞成骨能力。综上，本研究证实，长链非编码RNA LOC100126784与POM121L9P可通过miR-503-5p/SORBS1轴促进骨髓间充质干细胞的成骨分化，为骨质疏松症与骨缺损的治疗提供了潜在靶点。