MFhHF mediated NANOG delivery to hHF-MSCs. (A) Schematics of plasmids used in the experiments. NANOG expression was driven by CMV promoter and followed by IRES–egfp to enable quantitation of the transfection efficiency. Empty vector without NANOG was used as control for comparison. (B) Gene delivery was confirmed by flow cytometry (%EGFP+ cells). NANOG overexpression was demonstrated by using (C) quantitative real-time PCR (qRT-PCR), (D) reverse transcription polymerase chain reaction (RT-PCR), (E) Western blot, (F) immunocytochemistry, and (G) luciferase reporter assay. For the latter, hHF-MSCs were modified to express luciferase under the control of NANOG response element (NANOG-RE: NANOG-binding DNA motif). (D) RPL32 and (E) GAPDH served as a loading control for qRT-PCR and Western blot, respectively. The symbol * denotes p < 0.05 between control and NANOG-expressing cells. All values are the mean ± SD of triplicate samples in a representative experiment (n = 3).