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Bone marrow mesenchymal stem cell-derived exosomal miR-206 promotes osteoblast proliferation and differentiation in osteoarthritis by reducing Elf3.

ABSTRACT: MicroRNAs (miRNAs) serve as gene silencers involved in essential cell functions. The role of miR-206 and E74-like factor 3 (Elf3) has been identified in osteoarthritis (OA), while the effect of exosomal miR-206 from bone marrow mesenchymal stem cells (BMSCs) in OA remains largely unknown. Thus, we aim to explore the role of exosomal miR-206 from BMSCs in OA with the involvement of Elf3. BMSCs and BMSC-derived exosomes (BMSC-exos) were obtained and identified. OA mouse models were constructed by anterior cruciate ligament transection and then treated with BMSC-exos or BMSC-exos containing miR-206 mimic/inhibitor. The expression of miR-206, Elf3, inflammatory factors, osteocalcin (OCN) and bone morphogenetic protein 2 (BMP2) in mouse femoral tissues was assessed. The pathological changes in mouse femur tissues were observed. The mouse osteoblasts were identified and treated with untransfected or transfected BMSC-exos, and then, the expression of miR-206, Elf3, OCN and BMP2 was determined. The alkaline phosphatase (ALP) activity, calcium deposition level, OCN secretion, proliferation, apoptosis and cell cycle arrest in osteoblasts were measured. MiR-206 was down-regulated while Elf3 was up-regulated in OA animal and cellular models. Exosomal miR-206 ameliorated inflammation and increased expression of OCN and BMP2 in mouse femoral tissues. Moreover, exosomal miR-206 promoted ALP activity, calcium deposition level, OCN secretion and proliferation and inhibited apoptosis in OA osteoblasts. Overexpressed Elf3 reversed miR-206 up-regulation-induced effects on OA osteoblasts. BMSC-derived exosomal miR-206 promotes proliferation and differentiation of osteoblasts in OA by reducing Elf3. Our research may provide novel targets for OA treatment.

SUBMITTER: Huang Y

PROVIDER: S-EPMC8358849 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundEmerging evidence suggests that microRNAs (miRs) are associated with the progression of osteoarthritis (OA). In this study, the role of exosomal miR-136-5p derived from mesenchymal stem cells (MSCs) in OA progression is investigated and the potential therapeutic mechanism explored.MethodsBone marrow mesenchymal stem cells (BMMSCs) and their exosomes were isolated from patients and identified. The endocytosis of chondrocytes and the effects of exosome miR-136-5p on cartilage degradation were observed and examined by immunofluorescence and cartilage staining. Then, the targeting relationship between miR-136-5p and E74-like factor 3 (ELF3) was analyzed by dual-luciferase report assay. Based on gain- or loss-of-function experiments, the effects of exosomes and exosomal miR-136-5p on chondrocyte migration were examined by EdU and Transwell assay. Finally, a mouse model of post-traumatic OA was developed to evaluate effects of miR-136-5p on chondrocyte degeneration in vivo.ResultsIn the clinical samples of traumatic OA cartilage tissues, we detected increased ELF3 expression, and reduced miR-136-5p expression was determined. The BMMSC-derived exosomes showed an enriched level of miR-136-5p, which could be internalized by chondrocytes. The migration of chondrocyte was promoted by miR-136-5p, while collagen II, aggrecan, and SOX9 expression was increased and MMP-13 expression was reduced. miR-136-5p was verified to target ELF3 and could downregulate its expression. Moreover, the expression of ELF3 was reduced in chondrocytes after internalization of exosomes. In the mouse model of post-traumatic OA, exosomal miR-136-5p was found to reduce the degeneration of cartilage extracellular matrix.ConclusionThese data provide evidence that BMMSC-derived exosomal miR-136-5p could promote chondrocyte migration in vitro and inhibit cartilage degeneration in vivo, thereby inhibiting OA pathology, which highlighted the transfer of exosomal miR-136-5p as a promising therapeutic strategy for patients with OA.

Project description:The authors' previous study demonstrated that miR?128 may exert an inhibitory effect on the osteogenic differentiation of bone marrow?derived mesenchymal stem cells (BM?MSCs), but its downstream mechanisms remain to be elucidated. The aim of the present study was to investigate the microRNA (miRNA/miR) and mRNA profiles of differentiated and undifferentiated BM?MSCs and explore new downstream targets for miR?128. The sequencing datasets of GSE107279 (miRNA) and GSE112318 (mRNA) were downloaded from the Gene Expression Omnibus database. The differentially expressed miRNAs (DEMs) and genes (DEGs) were identified using the DESeq2 method. The target genes of DEMs were predicted by the miRwalk 2.0 database. The hub target genes of miR?128 were screened by constructing the protein?protein interaction (PPI) network and module analysis. The expression levels of miR?128 and crucial target genes were validated by reverse transcription?quantitative (RT?q) PCR before or after transfection of miR?128 mimics to BM?MSCs. The miRNA expression profile analysis identified miR?128 as one of the significantly downregulated DEMs (total 338) in differentiated BM?MSCs compared with the undifferentiated control. A total of 103 predicted target genes of miR?128?3p were overlapped with upregulated DEGs. By calculating the topological properties of each protein in the PPI network, 6 upregulated genes (KIT, NTRK2, YWHAB, GAB1, AXIN1 and RUNX1; fold change was the highest for NTRK2) were considered to be hub genes. Of these, 4 were enriched in module 4 (RUNX1, KIT, GAB1 and AXIN1; RUNX1 was particularly crucial as it can interact with the others), while one was enriched in module 7 (YWHAB). The expression levels of miR?128 and these 6 target genes during the osteogenic differentiation were experimentally confirmed by RT?qPCR. In addition, the expression levels of these 6 genes were significantly reversed after transfection of miR?128?3p mimics into rat BM?MSCs compared with the miR?control group. These findings indicated that miR?128?3p may inhibit the osteoblast differentiation of BM?MSCs by downregulation of these 6 genes, particularly RUNX1, YWHAB and NTRK2.

Project description:Skeletal muscle growth and maintenance depend on two tightly regulated processes, myogenesis and muscle regeneration. Both processes involve a series of crucial regulatory molecules including muscle-specific microRNAs, known as myomiRs. We recently showed that four myomiRs, miR-1, miR-133a, miR-133b, and miR-206, are encapsulated within muscle-derived exosomes and participate in local skeletal muscle communication. Although these four myomiRs have been extensively studied for their function in muscles, no information exists regarding their endogenous and exosomal levels across age. Here we aimed to identify any age-related changes in the endogenous and muscle-derived exosomal myomiR levels during acute skeletal muscle growth. The four endogenous and muscle-derived myomiRs were investigated in five skeletal muscles (extensor digitorum longus, soleus, tibialis anterior, gastrocnemius, and quadriceps) of 2-week-1-year-old wild-type male mice. The expression of miR-1, miR-133a, and miR-133b was found to increase rapidly until adolescence in all skeletal muscles, whereas during adulthood it remained relatively stable. By contrast, endogenous miR-206 levels were observed to decrease with age in all muscles, except for soleus. Differential expression of the four myomiRs is also inversely reflected on the production of two protein targets; serum response factor and connexin 43. Muscle-derived exosomal miR-1, miR-133a, and miR-133b levels were found to increase until the early adolescence, before reaching a plateau phase. Soleus was found to be the only skeletal muscle to release exosomes enriched in miR-206. In this study, we showed for the first time an in-depth longitudinal analysis of the endogenous and exosomal levels of the four myomiRs during skeletal muscle development. We showed that the endogenous expression and extracellular secretion of the four myomiRs are associated to the function and size of skeletal muscles as the mice age. Overall, our findings provide new insights for the myomiRs' significant role in the first year of life in mice.

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Bone marrow mesenchymal stem cell-derived exosomal miR-206 promotes osteoblast proliferation and differentiation in osteoarthritis by reducing Elf3.

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