Figure 3.

TCM delays mesoderm and specifically cardiac mesoderm differentiation. (A) Schematic depiction of the progression of differentiation from ESCs to cardiomyocytes including representative marker genes. (B) RT-qPCR analysis of differentiation stage-specific marker genes representative at various days of differentiation, as indicated. The most affected processes are the specification of cardiac mesoderm (Mesp1) at D4 and of cardiac progenitors (Nkx2.5, Tbx5) at D5. Data of three independent biological replicates are normalized to BM conditions and are displayed as mean ± SEM. Statistical analysis was by unpaired two-tailed Student’s t-test. ** p < 0.01, *** p < 0.001. (C) Progression of mesoderm (Brachyury) and cardiac mesoderm (Mesp1) specification at D4 and D5 of differentiation in EBs from the same starting samples. In BM-grown EBs, Brachyury peaks at D4 and has declined in expression levels at D5. In TCM, Brachyury reaches peak expression only at D5. Mesp1 activation is even more delayed. (D) Diagrammatic depiction of the time frame most impacted by TCM.