Bone mesenchymal stem cells (BMSCs)-derived exosomal microRNA-21-5p regulates Kruppel-like factor 3 (KLF3) to promote osteoblast proliferation <i>in vitro</i>

Bone mesenchymal stem cells (BMSCs)-derived exosomes (Exos) play important roles in osteoporosis, while the regulation of microRNA (miR)-21-5p remains unclear. The BMSCs-derived exosomes were isolated from femoral bone marrow of trauma patients, which were then used to stimulate human osteoblasts (hFOB1.19 cells). The miR-21-5p mimic or inhibitor was transfected into BMSCs to overexpress or knockdown miR-21-5p. The functions of miR-21-5p in osteoporosis were assessed by cell counting kit-8 (CCK-8) assay, alkaline phosphatase (ALP) staining and alizarin red staining assays. We found that BMSCs-derived exosomes could enhance proliferation, osteoblastic differentiation and ALP activity of hFOB1.19 cells. BMSCs-derived exosomes with upregulated miR-21-5p could further enhance these protective impacts compared with that in BMSCs-derived exosomes, while BMSCs-derived exosomes with downregulated miR-21-5p reduced these cell phenotypes. MiR-21-5p could directly bind to the 3’-untranslated region (UTR) of Kruppel-like factor 3 (KLF3), and knockdown of KLF3 obviously attenuated these inhibitory effects of BMSCs-derived exosomes with downregulated miR-21-5p on osteoblastic differentiation and ALP activity of hFOB1.19 cells. In summary, BMSCs-derived exosomal miR-21-5p improved osteoporosis through regulating KLF3, providing a potential therapeutic strategy for osteoporosis.骨髓间充质干细胞（Bone mesenchymal stem cells, BMSCs）分泌的外泌体（exosomes, Exos）在骨质疏松症中发挥关键调控作用，而微小RNA（microRNA, miR）-21-5p的调控机制尚未阐明。本研究从创伤患者股骨骨髓中分离获取BMSCs源性外泌体，并用其刺激人成骨细胞（hFOB1.19细胞）；通过向BMSCs中转染miR-21-5p模拟物或抑制剂，分别实现miR-21-5p的过表达与敲低。采用细胞计数试剂盒-8（cell counting kit-8, CCK-8）检测、碱性磷酸酶（alkaline phosphatase, ALP）染色及茜素红染色实验，对miR-21-5p在骨质疏松症中的生物学功能进行评价。结果显示，BMSCs源性外泌体可增强hFOB1.19细胞的增殖能力、成骨分化水平及ALP活性；过表达miR-21-5p的BMSCs源性外泌体可进一步强化上述保护性作用，而敲低miR-21-5p的BMSCs源性外泌体则会削弱此类细胞表型。机制研究表明，miR-21-5p可直接结合Kruppel样转录因子3（Kruppel-like factor 3, KLF3）的3’非编码区（3’-UTR），且敲低KLF3可显著逆转敲低miR-21-5p的BMSCs源性外泌体对hFOB1.19细胞成骨分化及ALP活性的抑制效应。综上，BMSCs源性外泌体携带的miR-21-5p通过调控KLF3改善骨质疏松症，为该病的临床治疗提供了潜在策略。