Changes in the expression of genes involved in ESC self-renewal and differentiation, following retinoic acid (RA) treatment.

<p>(A) Alkaline phosphatase staining of J1 mESCs cultured on gelatin-coated plates without LIF and treated with 1 μM RA or DMSO for 24 h. Alkaline phosphatase staining is an indicator of pluripotency. Scale bars = 100 μm. (B, C, D) qPCR, immunofluorescence, and western blot analysis of Oct4 and Nanog expression in J1 mESCs. DAPI was used for nuclear staining. Scale bars represent 100 μm. (E) Expression pattern of the representative upregulated differentiation-associated genes and downregulated pluripotency-related genes. Low intensities are shown in green and high intensities in red. mESCs were treated with DMSO or RA. (F) Representative differentiation-associated genes. The Hoxb gene was upregulated significantly by RA treatment. (G) Representative pluripotency-related genes downregulated in the presence of RA were validated by qPCR. (H) Heatmap diagram of three germ cell marker genes treated with or without RA. Epiblast marker expression was significantly altered by RA. (I) qPCR analysis validated these trends for the altered expression of genes related to growth and development of mESCs. J1 mESCs cultured with DMSO (vehicle) or 1 μM RA for 24 h. Data are presented as the mean ± SEM of three independent experiments (*p < 0.05; **p < 0.01).