Mesenchymal stem cells (MSCs) respond to diverse growth factors and signaling cascades that coordinately regulate their osteogenic differentiation potential. Bone Morphogenetic Protein 9 (BMP9), a member of the TGF-β superfamily, is one of the most potent inducers of osteogenic differentiation of MSCs.1 DNA methylation plays a role in osteogenesis as an epigenetic regulatory mechanism. Previous studies demonstrated that hypermethylation of the BMP2 promoter regions silenced osteogenic genes.2 The tet-eleven translocation (Tet) family of dioxygenases promotes DNA demethylation through oxidizing 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC) and its derivatives in an Fe(II)/α-ketoglutarate-dependent oxidation reaction.3 While all three TET family members (Tet1, Tet2, Tet3) are expressed in MSCs, Tet2 has been identified as the most significantly upregulated member upon osteogenic induction and is critically involved in lineage determination. Furthermore, studies have shown that Tet2, rather than Tet1 or Tet3, plays a dominant role in the transcriptional activation of key osteogenic factors like Runx2.4 Here, by employing a multiplex-siRNA system to effectively silence Tet2 expression in MSCs, we demonstrate that the loss of Tet2 expression significantly hampers BMP9-induced osteogenic differentiation and ectopic bone formation, confirming our hypothesis in which Tet2-mediated DNA demethylation plays a significant regulatory role in osteogenic differentiation of MSCs.