Abstract
Recent evidence has demonstrated that mesenchymal stem cells (MSCs) can release a large number of functionally specific microRNA (miRNA) microvesicles that play a role in promoting osteogenic differentiation, but the specific mechanism is not yet clear. Under such context, this study aims to elucidate the mechanism of bone marrow mesenchymal stem cell-derived exosomes (BMSC-Exo) promoting fracture healing in mice. We isolated and identified the BMSC-Exo. Bioinformatics analysis predicted high expression of miRNA in exosomes and verified the transfer of miR-25 in exosomes by immunofluorescence. Targeting relationship between miR-25 and Smad ubiquitination regulatory factor-1 (SMURF1) was predicted and verified by dual-luciferase reporter gene assay. Immunoprecipitation and protein stability assays were used to detect Runt-related transcription factor 2 (Runx2) ubiquitination and the effect of SMURF1 on Runx2 ubiquitination, respectively. The effect of miR-25 in BMSC-Exo on fracture healing in mice was assessed using X-ray imaging. alkaline phosphatase, alizarin red staining, EdU, CCK-8, and Transwell were used to evaluate the effects of exosomes transferred miR-25 on osteogenic differentiation, proliferation, and migration of osteoblasts. Bioinformatics analysis predicted that miR-25 expression in exosomes increased significantly. Moreover, the targeted regulation of SMURF1 by miR-25 was verified. SMURF1 inhibited Runx2 protein expression by promoting ubiquitination degradation of Runx2. Notably, miR-25 secreted by BMSC-Exo can accelerate osteogenic differentiation, proliferation, and migration of osteoblasts through SMURF1/Runx2 axis. Our results demonstrate that miR-25 in BMSC-Exo regulates the ubiquitination degradation of Runx2 by SMURF1 to promote fracture healing in mice.
Highlights
Fracture non-union occurs in 10–20% of fractures and has a significant impact on quality of life and total cost of care
We evaluated the ability of mesenchymal stem cells (MSCs) to induce differentiation in vitro, and the results showed that bone marrow mesenchymal stem cells (BMSCs) have osteogenic and adipogenic abilities (Figures 1B,C)
In order to further investigate whether osteoblasts can uptake BMSCs-Exo, exosomes were labeled with a fluorescent dye (PKH67) and added with MC3T3-E1C cells for incubation for 12 h
Summary
Fracture non-union occurs in 10–20% of fractures and has a significant impact on quality of life and total cost of care. It is essential to investigate the underlying mechanism of osteoblast differentiation induced by miRNAs derived from MSC-secreted exosomes so that we may have deeper understanding of bone loss diseases and it would contribute to the development of MSC-based therapeutic strategies for bone diseases. Raghuvaran Narayanan et al [10] illustrated that exosomes of MSCs can cause significant upregulation of various genes, including growth factors such as bone morphogenetic protein 9 (BMP9) and transforming growth factor-β, transcription factors, and extracellular matrix molecules, testifying that MSC-derived exosomes have the potential to induce osteogenic differentiation. The signaling pathways and molecular mechanisms of exosomes promoting fracture healing needs further study
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
