Molecular and Cellular Biochemistry

, Volume 387, Issue 1–2, pp 227–239 | Cite as

MicroRNA expression signature for Satb2-induced osteogenic differentiation in bone marrow stromal cells

  • Yiming Gong
  • Fei Xu
  • Ling Zhang
  • Yanyan Qian
  • Jake Chen
  • Huijun Huang
  • Youcheng YuEmail author


Satb2 acts as a potent transcription factor to promote osteoblast differentiation and bone regeneration. Recently, microRNAs (miRNA) have been identified as critical regulators of osteogenic differentiation. This study aimed to identify specific miRNAs and their regulatory roles in the process of Satb2-induced osteogenic differentiation. We studied the differentially expressed miRNAs by Satb2 overexpression in murine bone marrow stromal cells using miRNA microarray. Ten down-regulated miRNAs including miR-27a, miR-125a-5p, and miR-466f-3p, and 18 up-regulated miRNAs including miR-17, miR-20a and miR-210 were found to be differentially expressed and their expression were verified by quantitative real time PCR. The differentially expressed miRNAs were further subjected to gene ontology and KEGG analysis. The highly enriched GOs and KEGG pathway showed target genes of these miRNAs were significantly involved in multiple biological processes (mesenchymal cell differentiation, bone formation, and skeletal development), and several osteogenic pathways (TGF-β/BMP, MAPK, and Wnt signaling pathway). Finally, miR-27a was selected for target verification and function analysis. BMP2, BMPR1A, and Smad9, members of the TGF-β/BMP superfamily, which were predicted to be target genes of miR-27a, were confirmed to be significantly up-regulated in Satb2-overexpressing cells by quantitative real time PCR. Overexpression of miR-27a significantly inhibited osteogenesis and repressed BMP2, BMPR1A, and Smad9 expression. In this study, we identified that a number of differentially regulated miRNAs, whose target genes involved in the TGF-β/BMP signaling pathway, play an important role in the early stage of Satb2-induced osteogenic differentiation.


Osteogenic differentiation MicroRNA Satb2 Bone marrow stromal cell Transforming growth factor β BMP 



This work was supported by a grant from Major Scientific and Technological Research Projects of Science and Technology Commission of Shanghai Municipality (No. 10JC1402600).

Conflict of interest

All authors have no conflicts of interest.


  1. 1.
    Dennis JE, Haynesworth SE, Young RG, Caplan AI (1992) Osteogenesis in marrow-derived mesenchymal cell porous ceramic composites transplanted subcutaneously: effect of fibronectin and laminin on cell retention and rate of osteogenic expression. Cell Transplant 1:23–32PubMedGoogle Scholar
  2. 2.
    Franceschi RT (2005) Biological approaches to bone regeneration by gene therapy. J Dent Res 84:1093–1103PubMedCrossRefGoogle Scholar
  3. 3.
    Partridge K, Yang X, Clarke NM, Okubo Y, Bessho K, Sebald W, Howdle SM, Shakesheff KM, Oreffo RO (2002) Adenoviral BMP-2 gene transfer in mesenchymal stem cells: in vitro and in vivo bone formation on biodegradable polymer scaffolds. Biochem Biophys Res Commun 292:144–152PubMedCrossRefGoogle Scholar
  4. 4.
    Samee M, Kasugai S, Kondo H, Ohya K, Shimokawa H, Kuroda S (2008) Bone morphogenetic protein-2 (BMP-2) and vascular endothelial growth factor (VEGF) transfection to human periosteal cells enhances osteoblast differentiation and bone formation. J Pharmacol Sci 108:18–31PubMedCrossRefGoogle Scholar
  5. 5.
    Zhang J, Tu Q, Grosschedl R, Kim MS, Griffin T, Drissi H, Yang P, Chen J (2011) Roles of SATB2 in osteogenic differentiation and bone regeneration. Tissue Eng A 17:1767–1776CrossRefGoogle Scholar
  6. 6.
    Dobreva G, Chahrour M, Dautzenberg M, Chirivella L, Kanzler B, Fariñas I, Karsenty G, Grosschedl R (2006) SATB2 is a multifunctional determinant of craniofacial patterning and osteoblast differentiation. Cell 125:971–986PubMedCrossRefGoogle Scholar
  7. 7.
    Kim IS, Jeong SJ, Kim SH, Jung JH, Park YG, Kim SH (2012) Special AT-rich sequence-binding protein 2 and its related genes play key roles in the differentiation of MC3T3-E1 osteoblast like cells. Biochem Biophys Res Commun 17:697–703CrossRefGoogle Scholar
  8. 8.
    Ye JH, Xu YJ, Gao J, Yan SG, Zhao J, Tu Q, Zhang J, Duan XJ, Sommer CA, Mostoslavsky G, Kaplan DL, Wu YN, Zhang CP, Wang L, Chen J (2011) Critical-size calvarial bone defects healing in a mouse model with silk scaffolds and SATB2-modified iPSCs. Biomaterials 32:5065–5076PubMedCentralPubMedCrossRefGoogle Scholar
  9. 9.
    Yan SG, Zhang J, Tu QS, Ye JH, Luo E, Schuler M, Kim MS, Griffin T, Zhao J, Duan XJ, Cochran DJ, Murray D, Yang PS, Chen J (2011) Enhanced osseointegration of titanium implant through the local delivery of transcription factor SATB2. Biomaterials 32:8676–8683PubMedCentralPubMedCrossRefGoogle Scholar
  10. 10.
    Stefani G, Slack FJ (2008) Small non-coding RNAs in animal development. Nat Rev Mol Cell Biol 9:219–230PubMedCrossRefGoogle Scholar
  11. 11.
    Guo L, Zhao RC, Wu Y (2011) The role of microRNAs in self-renewal and differentiation of mesenchymal stem cells. Exp Hematol 39:608–616PubMedCrossRefGoogle Scholar
  12. 12.
    Zhang Y, Xie RL, Croce CM, Stein JL, Lian JB, van Wijnen AJ, Stein GS (2011) A program of microRNAs controls osteogenic lineage progression by targeting transcription factor Runx2. Proc Natl Acad Sci USA 108:9863–9868PubMedCrossRefGoogle Scholar
  13. 13.
    Wu T, Zhou H, Hong Y, Li J, Jiang X, Huang H (2012) miR-30 family members negatively regulate osteoblast differentiation. J Biol Chem 287:7503–7511PubMedCrossRefGoogle Scholar
  14. 14.
    Huang J, Zhao L, Xing L, Chen D (2010) MicroRNA-204 regulates Runx2 protein expression and mesenchymal progenitor cell differentiation. Stem Cells 28:357–364PubMedCentralPubMedGoogle Scholar
  15. 15.
    Hassan MQ, Gordon JA, Beloti MM, Croce CM, van Wijnen AJ, Stein JL, Stein GS, Lian JB (2010) A network connecting Runx2, SATB2, and the miR-23a~27a~24-2 cluster regulates the osteoblast differentiation program. Proc Natl Acad Sci USA 107:19879–19884PubMedCrossRefGoogle Scholar
  16. 16.
    Deng Y, Wu S, Zhou H, Bi X, Wang Y, Hu Y, Gu P, Fan X (2013) Effects of a miR-31, Runx2, and Satb2 regulatory loop on the osteogenic differentiation of bone mesenchymal stem cells. Stem Cells Dev 22:2278–2286PubMedCrossRefGoogle Scholar
  17. 17.
    Wei J, Shi Y, Zheng L, Zhou B, Inose H, Wang J, Guo XE, Grosschedl R, Karsenty G (2012) miR-34s inhibit osteoblast proliferation and differentiation in the mouse by targeting SATB2. J Cell Biol 197:509–521PubMedCrossRefGoogle Scholar
  18. 18.
    Zhao Z, Zhao M, Xiao G, Franceschi RT (2005) Gene transfer of the Runx2 transcription factor enhances osteogenic activity of bone marrow stromal cells in vitro and in vivo. Mol Ther 12:247–253PubMedCrossRefGoogle Scholar
  19. 19.
    Suomi S, Taipaleenmäki H, Seppänen A, Ripatti T, Väänänen K, Hentunen T, Säämänen AM, Laitala-Leinonen T (2008) MicroRNAs regulate osteogenesis and chondrogenesis of mouse bone marrow stromal cells. Gene Regul Syst Biol 2:177–191Google Scholar
  20. 20.
    Tiscornia G, Singer O, Verma IM (2006) Design and cloning of lentiviral vectors expressing small interfering RNAs. Nat Protoc 1:234–240PubMedCrossRefGoogle Scholar
  21. 21.
    Gene Ontology Consortium (2006) The Gene Ontology (GO) project in 2006. Nucleic Acids Res 34:D322–D326PubMedCentralCrossRefGoogle Scholar
  22. 22.
    Dennis G Jr, Sherman BT, Hosack DA, Yang J, Gao W, Lane HC, Lempicki RA (2003) DAVID: Database for Annotation, Visualization, and Integrated Discovery. Genome Biol 4:P3PubMedCentralPubMedCrossRefGoogle Scholar
  23. 23.
    Mizuno Y, Tokuzawa Y, Ninomiya Y, Yagi K, Yatsuka-Kanesaki Y, Suda T, Fukuda T, Katagiri T, Kondoh Y, Amemiya T, Tashiro H, Okazaki Y (2009) miR-210 promotes osteoblastic differentiation through inhibition of AcvR1b. FEBS Lett 583:2263–2268PubMedCrossRefGoogle Scholar
  24. 24.
    Zhou M, Ma J, Chen S, Chen X, Yu X (2013) MicroRNA-17-92 cluster regulates osteoblast proliferation and differentiation. Endocrine (Epub ahead of print)Google Scholar
  25. 25.
    Zhang JF, Fu WM, He ML, Xie WD, Lv Q, Wan G, Li G, Wang H, Lu G, Hu X, Jiang S, Li JN, Lin MC, Zhang YO, Kung HF (2011) miRNA-20a promotes osteogenic differentiation of human mesenchymal stem cells by co-regulating BMP signaling. RNA Biol 8:829–838PubMedCrossRefGoogle Scholar
  26. 26.
    Liu Y, Liu W, Hu C, Xue Z, Wang G, Ding B, Luo H, Tang L, Kong X, Chen X, Liu N, Ding Y, Jin Y (2011) MiR-17 modulates osteogenic differentiation through a coherent feed-forward loop in mesenchymal stem cells isolated from periodontal ligaments of patients with periodontitis. Stem Cells 29:1804–1816PubMedCrossRefGoogle Scholar
  27. 27.
    Schoolmeesters A, Eklund T, Leake D, Vermeulen A, Smith Q, Force Aldred S, Fedorov Y (2009) Functional profiling reveals critical role for miRNA in differentiation of human mesenchymal stem cells. PLoS ONE 4:e5605PubMedCentralPubMedCrossRefGoogle Scholar
  28. 28.
    Wang T, Xu Z (2010) miR-27 promotes osteoblast differentiation by modulating Wnt signaling. Biochem Biophys Res Commun 402:186–189PubMedCrossRefGoogle Scholar
  29. 29.
    Zhang ZJ, Zhang H, Kang Y, Sheng PY, Ma YC, Yang ZB, Zhang ZQ, Fu M, He AS, Liao WM (2012) miRNA expression profile during osteogenic differentiation of human adipose-derived stem cells. J Cell Biochem 113:888–898PubMedCrossRefGoogle Scholar
  30. 30.
    Mizuno Y, Yagi K, Tokuzawa Y, Kanesaki-Yatsuka Y, Suda T, Katagiri T, Fukuda T, Maruyama M, Okuda A, Amemiya T, Kondoh Y, Tashiro H, Okazaki Y (2008) miR-125b inhibits osteoblastic differentiation by down-regulation of cell proliferation. Biochem Biophys Res Commun 368:267–272PubMedCrossRefGoogle Scholar
  31. 31.
    Chen G, Deng C, Li YP (2012) TGF-β and BMP signaling in osteoblast differentiation and bone formation. Int J Biol Sci 8:272–288PubMedCentralPubMedCrossRefGoogle Scholar
  32. 32.
    Liu DD, Zhang JC, Zhang Q, Wang SX, Yang MS (2013) TGF-β/BMP signaling pathway is involved in cerium-promoted osteogenic differentiation of mesenchymal stem cells. J Cell Biochem 114:1105–1114PubMedCrossRefGoogle Scholar
  33. 33.
    Gazzerro E, Canalis E (2006) Bone morphogenetic proteins and their antagonists. Rev Endocr Metab Disord 7:51–65PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Yiming Gong
    • 1
  • Fei Xu
    • 2
  • Ling Zhang
    • 3
  • Yanyan Qian
    • 2
  • Jake Chen
    • 4
  • Huijun Huang
    • 2
  • Youcheng Yu
    • 1
    Email author
  1. 1.Department of Stomatology, Zhongshan HospitalFudan UniversityShanghaiChina
  2. 2.Department of Biochemistry and Molecular Biology, Shanghai Medical CollegeFudan UniversityShanghaiChina
  3. 3.Fusen LaboratoryFusen Biological Technology Co., LtdShanghaiChina
  4. 4.Division of Oral BiologyTufts School of Dental MedicineBostonUSA

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