CsA and DMSO synergistically increase cardiac differentiation of P19 cells during EB formation.

<p>(A) Schematics for cardiac differentiation of P19 cells treated with 0.32 μM CsA, 1% DMSO, or CsA plus DMSO are shown. (B) Real-time PCR results showing the synergistic effect on cardiac differentiation of P19 cells at day 16 of differentiation after cotreatment with 0.32 μM CsA and 1% DMSO during EB formation. Data represent mean ± SD from three independent experiments (*<i>p</i><0.05; **<i>p</i><0.01; ***<i>p</i><0.001). (C) Representative Western blot showing the synergistic effect on cardiac differentiation of P19 cells of cotreatment with 0.32 μM CsA and 1% DMSO. The cell lysates were probed with anti-α-MHC antibody, and with anti-GAPDH antibody as a loading control. (D) The relative expression levels of α-MHC protein were quantified by densitometric analysis and normalized against the protein expression levels of GAPDH, used as a loading control. The results shown represent mean ± SD from three independent experiments (***<i>p</i><0.001). (E) Graph showing the percentages of 6-well culture plates containing beating cells with 0.32 μM CsA, 1% DMSO, or CsA plus DMSO. (F) Real-time PCR results showing the synergistic effect of Wnt3a, Wnt5a, Wnt8a, Mixl1, Mesp1 and Mesp2 mRNA expression in EBs treated for 48 h with 0.32 μM CsA and 1% DMSO. Data represent mean ± SD from three independent experiments (*p<0.05; **p<0.01; ***p<0.001).