RETRACTED ARTICLE: MicroRNA-15b shuttled by bone marrow mesenchymal stem cell-derived extracellular vesicles binds to WWP1 and promotes osteogenic differentiation

Abstract

Background Osteogenic differentiation is an essential process for bone regeneration involving bone marrow mesenchymal stem cells (BMSCs). BMSC-secreted extracellular vesicles (EVs) enriched with microRNAs (miRs) have vital roles to play in mediating osteogenic differentiation. Therefore, this study aimed to explore the effect of BMSC-derived EVs loaded with miR-15b on osteogenic differentiation. Methods Human BMSCs (hBMSCs) were cultured and treated with plasmids overexpressing or knocking down KLF2, WWP1, and miR-15b to define the role of derived EVs in osteogenic differentiation in vitro. The expression of osteogenic differentiation-related marker was measured by Western blot analysis. The interaction among miR-15b, WWP1, and ubiquitination of KLF2 was investigated by dual-luciferase reporter, immunoprecipitation, and GST pull-down assays. Moreover, EVs from hBMSCs transfected with miR-15b inhibitor (EV-miR-15b inhibitor) were injected into ovariectomized rats to verify the effect of miR-15b on bone loss in vivo. Results WWP1 was downregulated, and KLF2 was upregulated during osteogenic differentiation. After co-culture with EVs, miR-15b expression was elevated and WWP1 expression was reduced in hBMSCs. Upregulation of miR-15b or KLF2 or downregulation of WWP1 or NF-κB increased ALP activity and cell mineralization, as well as osteogenic differentiation-related marker expression in hBMSCs. Mechanistically, miR-15b targeted and inhibited WWP1, thus attenuating KLF2 degradation and inhibiting NF-κB activity. Co-culture of EVs increased the bone volume and trabecular number, but decreased bone loss in ovariectomized rats, which could be reversed after treatment with EV-miR-15b inhibitor. Conclusion Collectively, BMSC-derived EVs loaded with miR-15b promoted osteogenic differentiation by impairing WWP1-mediated KLF2 ubiquitination and inactivating the NF-κB signaling pathway. Graphical abstract

Extracellular Vehicles of Oxygen-Depleted Mesenchymal Stromal Cells: Route to Off-Shelf Cellular Therapeutics?

“…One could speculate that analysis of the BMSC EV composition might shed light on BMSC physiology as well. Indeed, miRNA-15b, that is enriched in BMSC EVs revealed intracellular regulators of osteogenic differentiation confirming that BMSC EVs could provide invaluable information on BMSC-related processes [ 133 ].…”

Research Progress of Exosomes in Bone Diseases: Mechanism, Diagnosis and Therapy

“…Then they found that BMSC-derived exosomal let-7a-5p, let-7c-5p, miR-328a-5p, and miR-31a-5p could activate the phosphorylation of Smad1/5/9 and activate BMP/Smad signaling pathway by targeting Acvr2b to mediate osteogenic differentiation and repair bone defects ( Figure 2B ). Besides, other BMSC-derived exosomal miRNAs, such as miR-122-5p ( Liao et al, 2019 ), miR-15b ( Li Y. et al, 2020 ), miR-206 ( Huang et al, 2021d ), miR-19b ( Huang et al, 2021c ), miR-146a ( Liu L. et al, 2021 ), miR-877 ( Liang et al, 2021 ), miR-218 ( Hassan et al, 2012 ), and miR-128-3p ( Xu et al, 2020 ) also played significant roles in stimulating osteogenesis and angiogenesis. Briefly, BMSCs-derived exosomal miRNAs could participate in various pathways to promote bone formation, suggesting BMSC may be a key cell in maintaining bone metabolism.…”

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“…The study showed that WWP1 was targeted by miR-142-5p and consequently promoted osteoblast activity and matrix mineralization in favor of bone healing [ 114 ]. BMSC-secreted extracellular vesicles loaded with miR-15b, which targeted and repressed WWP1 expression, could promote osteogenic differentiation by attenuating WWP1-mediated KLF2 ubiquitination thus inhibiting the KLF2/NF-κB axis [ 115 ]. MiR-19b enriched in BMSC-derived exosomes targeted and suppressed the expression of WWP1 or Smurf2, thereby reducing KLF5 protein degradation by ubiquitination, promoting osteoblast differentiation, and facilitating fracture healing by modulating the Wnt/β-catenin signaling pathway [ 116 ].…”