Figure 3.

Exogenous Bmp4 induces gene expression associated with odontoblast differentiation in induced pluripotent stem cell-derived neural crest-like cells (iNCLCs). A Bmp4 expression plasmid was electroporated into second passage iNCLCs followed by 2 days of culture. Bmp4 expression was assessed by real-time polymerase chain reaction (PCR) (A) and Western blot (B); ∗, p < .01; n = 3. (C): Representative phase-contrast micrographs of control and Bmp4-transfected iNCLCs. Scale bar = 100 μm. Expression levels of Pax9 (D), Msx1 (E), Dmp1 (F), and Dspp (G) in Bmp4-overexpressing iNCLCs were determined by real-time PCR; ∗, p < .01; n = 3. (H): The expression of DSP (green) in control and Bmp4-transfected iNCLCs was analyzed by immunofluorescence. Nuclei are stained with DAPI (blue). Scale bar = 20 μm. (I): Expression of DSP was analyzed by Western blot. Total protein from dissected tooth germs of embryonic day 18.5 mice was used as a positive control. (J): Quantification of the DSP protein level relative to β-actin; ∗, p < .01; n = 3. Abbreviations: DAPI, 4′,6-diamidino-2-phenylindole; DSP, dentin sialoprotein.