背景:骨细胞是骨骼中关键的机械感觉细胞,机械刺激的骨细胞产生可以诱导成骨的外泌体。microRNAs(miRNAs)是外泌体的重要组成部分,骨细胞中的一些miRNAs调节成骨分化;以前的研究表明,机械应变的骨细胞中一些差异表达的miRNAs可能会影响成骨细胞的分化。因此,筛选和选择调节机械刺激的骨细胞外泌体成骨分化的miRNA是重要的。
结果:每天0.5Hz1h时2500με的机械拉伸应变,持续3天,MLO-Y4骨细胞的前列腺素E2(PGE2)和胰岛素样生长因子-1(IGF-1)水平和一氧化氮合酶(NOS)活性升高,并促进MC3T3-E1成骨细胞的成骨分化。14种miRNAs仅在MLO-Y4骨细胞中差异表达,这些骨细胞受到机械拉伸应变的刺激,被筛选,并鉴定了与成骨相关的miRNAs。四个差异表达的miRNA(miR-1930-3p,miR-3110-5p,miR-3090-3p,和miR-3058-3p)仅在机械应变的骨细胞中发现,和四个miRNA,仅在机械应变的成骨细胞中差异表达的八个靶向mRNA,也被确认了。此外,机械应变的骨细胞来源的外泌体促进MC3T3-E1细胞的成骨分化,外泌体被成骨细胞内化,以及在机械应变的骨细胞中上调的miR-3110-5p和miR-3058-3p,都在外泌体中增加,通过逆转录定量聚合酶链反应(RT-qPCR)验证。
结论:在骨细胞中,在0.5Hz时2500με的机械拉伸应变诱导了14种差异表达的miRNA,这些miRNA可能在骨细胞的外泌体中并参与成骨。机械应变的骨细胞来源的外泌体包含增加的miR-3110-5p和miR-3058-3p(14个miRNA中的两个),促进成骨细胞分化。
BACKGROUND: Osteocytes are critical mechanosensory cells in bone, and mechanically stimulated osteocytes produce exosomes that can induce osteogenesis. MicroRNAs (miRNAs) are important constituents of exosomes, and some miRNAs in osteocytes regulate osteogenic differentiation; previous studies have indicated that some differentially expressed miRNAs in mechanically strained osteocytes likely influence osteoblastic differentiation. Therefore, screening and selection of miRNAs that regulate osteogenic differentiation in exosomes of mechanically stimulated osteocytes are important.
RESULTS: A mechanical tensile strain of 2500 με at 0.5 Hz 1 h per day for 3 days, elevated prostaglandin E2 (PGE2) and insulin-like growth factor-1 (IGF-1) levels and nitric oxide synthase (NOS) activity of MLO-Y4 osteocytes, and promoted osteogenic differentiation of MC3T3-E1 osteoblasts. Fourteen miRNAs differentially expressed only in MLO-Y4 osteocytes which were stimulated with mechanical tensile strain, were screened, and the miRNAs related to osteogenesis were identified. Four differentially expressed miRNAs (miR-1930-3p, miR-3110-5p, miR-3090-3p, and miR-3058-3p) were found only in mechanically strained osteocytes, and the four miRNAs, eight targeted mRNAs which were differentially expressed only in mechanically strained osteoblasts, were also identified. In addition, the mechanically strained
osteocyte-derived exosomes promoted the osteoblastic differentiation of MC3T3-E1 cells in vitro, the exosomes were internalized by osteoblasts, and the up-regulated miR-3110-5p and miR-3058-3p in mechanically strained osteocytes, were both increased in the exosomes, which was verified via reverse transcription quantitative polymerase chain reaction (RT-qPCR).
CONCLUSIONS: In osteocytes, a mechanical tensile strain of 2500 με at 0.5 Hz induced the fourteen differentially expressed miRNAs which probably were in exosomes of osteocytes and involved in osteogenesis. The mechanically strained
osteocyte-derived exosomes which contained increased miR-3110-5p and miR-3058-3p (two of the 14 miRNAs), promoted osteoblastic differentiation.