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Deer Antler Reserve Mesenchyme Cells Modified with MiR-145 Promote Chondrogenesis in Cartilage Regeneration

ABSTRACT: Deer antler-derived reserve mesenchyme cells (RMCs) are a promising source of cells for cartilage regeneration therapy due to their chondrogenic differentiation potential. However, the regulatory mechanism has not yet been elucidated. microRNAs (miRNAs) have been shown to regulate the differentiation of various mesenchymal stem cells (MSCs) and play an important role in the post-transcriptional regulation of chondrogenesis and hypertrophic differentiation. In this study, we demonstrated that the RMCs showed typical MSCs differentiation potentials. During chondrogenic differentiation, we obtained the expression profile of miRNAs, among which miR-145 was the most prominent candidate as key microRNA involved in the balance of chondral and endochondral differentiation. Knockdown of miR-145 promoted chondrogenesis and inhibits hypertrophy differentiation in RMCs. Mechanically, by online databases prediction combined with dual-luciferase reporter assay, SOX9 was suggested as a target of miR-145. Further validation experiments confirmed that knockdown of miR-145 contributed to the balance between endochondral versus chondral differentiation of RMCs by targeting SOX9. Additionally, RMCs transfected with the miR-145 knockdown mediated of lentiviral vector successfully promoted cartilage regeneration in vivo. In summary, our study suggested that the reciprocal negative feedback between SOX9 and miR-145 was essential for balancing between endochondral versus chondral differentiation of RMCs. Our study suggested that modification of RMCs using miRNAs transduction might be an effective treatment for cartilage defects.

ORGANISM(S): Cervus nippon

PROVIDER: GSE277770 | GEO | 2025/01/15

REPOSITORIES: GEO

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Project description:Human mesenchymal stem cells (MSC) display a high potential for the development of novel treatment strategies for cartilage repair. However, the pathways involved in their differentiation to functional and non hypertrophic chondrocytes remain largely unknown, despite the work on embryologic development and the identification of key growth factors including members of the TGFbeta, Hh, Wnt and FGF families. In this study, we asked if we could identify specific biological networks independently from the growth factor used (TGFbeta-3 or BMP-2). To address this question, we used DNA microarrays and performed large-scale expression profiling of MSC at different time points during their chondral differentiation. By comparing these data with those obtained during their differentiation into osteoblasts and adipocytes, we identified 318 genes specific for chondrogenesis. We distributed the selected genes in 5 classes according to their kinetic of expression and used the Ingenuity software in order to identify new biological networks. We could reconstruct 3 phases for chondral differentiation, characterized by functional pathways. The first phase corresponds to cell attachment and apoptosis prevention with the up-regulation of √é¬±5 integrins, BCL6, NFIL3, RGS2 and down-regulation of CTGF and CYR61. The second phase is characterized by a proliferation/differentiation step with the continuous expression of MAF, PGF, HGMA1 or NOTCH3, CHI3L1, WNT5A, LEPR. Finally, the last step of differentiation/hypertrophy is characterized by expression of DKK1, APOD/E, SERPINF1 and TIMP4. These data propose new pathways to understand the complexity of MSC differentiation to chondrocytes and new potential targets for cell therapy applied to cartilage repair. Experiment Overall Design: To identify genes involved in TGFbeta03/BMP2-induced chondrogenesis, MSC were isolated from human bone marrow aspirates by adherence to culture dishes and cultivated to passage 3. After induction of chondrogenic differentiation with BMP2 or TGFbeta-3, gene expression was determined by microarray hybridization after 1, 3, 7 and 21 days and compared to the undifferentiated MSC.