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Mesenchymal stem cell-derived exosomes have altered microRNA profiles and induce osteogenic differentiation depending on the stage of differentiation.

ABSTRACT: Human mesenchymal stem cell (hMSC)-derived exosomes have shown regenerative effects, but their role in osteogenesis and the underlying mechanism are yet to be determined. In this study, we examined the time-course secretion of exosomes by hMSCs during the entire process of osteogenic differentiation. Exosomes derived from hMSCs in various stages of osteogenic differentiation committed homotypic cells to differentiate towards osteogenic lineage, but only exosomes from late stages of osteogenic differentiation induced extracellular matrix mineralisation. Exosomes from expansion and early and late stages of osteogenic differentiation were internalised by a subpopulation of hMSCs. MicroRNA profiling revealed a set of differentially expressed exosomal microRNAs from the late stage of osteogenic differentiation, which were osteogenesis related. Target prediction demonstrated that these microRNAs enriched pathways involved in regulation of osteogenic differentiation and general mechanisms how exosomes exert their functions, such as "Wnt signalling pathway" and "endocytosis". Taken together, the results show that MSCs secrete exosomes with different biological properties depending on differentiation stage of their parent cells. The exosomal cargo transferred from MSCs in the late stage of differentiation induces osteogenic differentiation and mineralisation. Moreover, it is suggested that the regulatory effect on osteogenesis by exosomes is at least partly exerted by exosomal microRNA.

SUBMITTER: Wang X

PROVIDER: S-EPMC5814093 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The need for an autologous cell source for bone tissue engineering and medical applications has led researchers to explore multipotent mesenchymal stromal cells (MSC), which show stem cell plasticity, in various human tissues. However, MSC with different tissue origins vary in their biological properties and their capability for osteogenic differentiation. Furthermore, MSC-based therapies require large-scale ex vivo expansion, accompanied by cell type-specific replicative senescence, which affects osteogenic differentiation. To elucidate cell type-specific differences in the osteogenic differentiation potential and replicative senescence, we analysed the impact of BMP and TGF-β signaling in adipose-derived stromal cells (ASC), fibroblasts (FB), and dental pulp stromal cells (DSC). We used inhibitors of BMP and TGF-β signaling, such as SB431542, dorsomorphin and/or a supplemental addition of BMP-2. The expression of high-affinity binding receptors for BMP-2 and calcium deposition with alizarin red S were evaluated to assess osteogenic differentiation potential. Our study demonstrated that TGF-β signaling inhibits osteogenic differentiation of ASC, DSC and FB in the early cell culture passages. Moreover, DSC had the best osteogenic differentiation potential and an activation of BMP signaling with BMP-2 could further enhance this capacity. This phenomenon is likely due to an increased expression of activin receptor-like kinase-3 and -6. However, in DSC with replicative senescence (in cell culture passage 10), osteogenic differentiation sharply decreased, and the simultaneous use of BMP-2 and SB431542 did not result in further improvement of this process. In comparison, ASC retain a similar osteogenic differentiation potential regardless of whether they were in the early (cell culture passage 3) or later (cell culture passage 10) stages. Our study elucidated that ASC, DSC, and FB vary functionally in their osteogenic differentiation, depending on their tissue origin and replicative senescence. Therefore, our study provides important insights for cell-based therapies to optimize prospective bone tissue engineering strategies.

Project description:BackgroundPeriodontal tissue regeneration is the ultimate goal of periodontitis treatment. Exosomes are nanoscale vesicles secreted by cells that participate in and regulate the physiological activities between cells. However, the relationship between inflammatory macrophage-derived exosomes and osteoblast differentiation in periodontitis has not been thoroughly reported. Here, we attempt to explore the role of inflammatory macrophage-derived exosomes in crosstalk with osteoblasts.MethodsPorphyromonas gingivalis lipopolysaccharide was used to stimulate macrophages and inflate their inflammatory cellular state. Exosomes were extracted from inflammatory macrophages using supercentrifugation, and their characteristics were detected by transmission electron microscopy, particle size analysis, and Western blotting. Exosome uptake bybone marrow mesenchymal stem cells (BMSCs) was observed by fluorescence microscopy. The effects of exosomes on the BMSC inflammatory response and on osteogenic differentiation were detected by quantitative polymerase chain reaction and Western blot analysis. Alkaline phosphatase activity was tested for verification.ResultsWe successfully extracted and identified inflammatory macrophage-derived exosomes and observed that BMSCs successfully took up exosomes. Inflammatory macrophage-derived exosomes upregulated the expression levels of the inflammatory factors interleukin-6 and tumour necrosis factor-alpha in BMSCs and mediated inflammatory stimulation. Additionally, they inhibited the transcription levels of the osteogenic genes alkaline phosphatase, type I collagen, and Runt-related transcription factor 2 as well as the alkaline phosphatase activity, while the use of the exosome inhibitor GW4869 attenuated this effect.ConclusionOur study shows that macrophages in periodontitis can mediate inflammatory stimulation and inhibit the osteogenic differentiation of bone marrow mesenchymal stem cells through the exosome pathway. Interference with exosome secretion is likely to be a promising method for bone tissue regeneration in inflammatory states.

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Mesenchymal stem cell-derived exosomes have altered microRNA profiles and induce osteogenic differentiation depending on the stage of differentiation.

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