Sox9-regulated miRNA-574-3p inhibits chondrogenic differentiation of mesenchymal stem cells

PLoS One. 2013 Apr 23;8(4):e62582. doi: 10.1371/journal.pone.0062582. Print 2013.

Abstract

The aim of this study was to identify new microRNAs (miRNAs) that are modulated during the differentiation of mesenchymal stem cells (MSCs) toward chondrocytes. Using large scale miRNA arrays, we compared the expression of miRNAs in MSCs (day 0) and at early time points (day 0.5 and 3) after chondrogenesis induction. Transfection of premiRNA or antagomiRNA was performed on MSCs before chondrogenesis induction and expression of miRNAs and chondrocyte markers was evaluated at different time points during differentiation by RT-qPCR. Among miRNAs that were modulated during chondrogenesis, we identified miR-574-3p as an early up-regulated miRNA. We found that miR-574-3p up-regulation is mediated via direct binding of Sox9 to its promoter region and demonstrated by reporter assay that retinoid X receptor (RXR)α is one gene specifically targeted by the miRNA. In vitro transfection of MSCs with premiR-574-3p resulted in the inhibition of chondrogenesis demonstrating its role during the commitment of MSCs towards chondrocytes. In vivo, however, both up- and down-regulation of miR-574-3p expression inhibited differentiation toward cartilage and bone in a model of heterotopic ossification. In conclusion, we demonstrated that Sox9-dependent up-regulation of miR-574-3p results in RXRα down-regulation. Manipulating miR-574-3p levels both in vitro and in vivo inhibited chondrogenesis suggesting that miR-574-3p might be required for chondrocyte lineage maintenance but also that of MSC multipotency.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Differentiation / genetics*
  • Chondrocytes / cytology*
  • Chondrocytes / metabolism
  • Chondrogenesis / genetics*
  • Gene Expression Profiling
  • Gene Expression Regulation
  • Humans
  • Mesenchymal Stem Cells / cytology*
  • Mesenchymal Stem Cells / metabolism*
  • Mice
  • MicroRNAs / genetics*
  • Retinoid X Receptor alpha / genetics
  • SOX9 Transcription Factor / genetics
  • SOX9 Transcription Factor / metabolism*

Substances

  • MIRN574 microRNA, human
  • MicroRNAs
  • Retinoid X Receptor alpha
  • SOX9 Transcription Factor

Grant support

Work in the laboratory Inserm U844 was supported by the Inserm Institute, the University of Montpellier I and funding from the Fondation pour la Recherche Médicale (FRM). D. Philipot was granted by “Région Languedoc-Roussillon and University of Montpellier I”. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.