Inhibition of T cell factor (Tcf3) transcription factor using oligodeoxynucleotide decoy strategy increases embryonic stem cell stemness

IntroductionMouse embryonic stem cells (mESCs) has a molecular pathway involved in the maintenance of stemness, including Hox genes, Wnt, BMP, Sonic Hedgehog, and Notch signaling. Wnt ligands promote nuclear accumulation of β-catenin, which associates with DNA-bound Tcf/Lef factors and activates transcription. In this base, Tcf3 is one member of the larger basic helix loop helix (bHLH) family transcription factors. The Tcf3 localizes to many pluripotency genes in the ES cells that promote cell-specific differentiation. In this study, inhibition of the Tcf3 transcription factor by using oligodeoxynucleotide decoy strategy was done to assess the stemness in mouse embryonic stem cell.MethodsTcf3 decoy and its scramble ODNs designed and synthesized by our group. Also, EMSA assay was used to confirm specificity interaction of the Tcf3 transcription factor with its protein. Self-renewal and pluripotency of mESCs were analyzed by MTT assay, alkaline phosphatase (ALP), embryoid body (EB) formation and real time- PCR assays.ResultsOur result showed that knockdown of the Tcf3 by decoy ODNs transfection in the mESCs lead to increase in cell proliferation, ALP enzyme activity, and master regulatory stemness genes and decrease in the number and diameter of EB.ConclusionIn conclusion, these findings suggest that Tcf3 decoy strategy can be used to maintain stemness in the ESCs being subjected to a long-term expansion regimen prior to tissue engineering applications

Keywords:

Tcf3 , Mouse embryonic stem cell , Transcription factor decoy ODNs , Stemness