Figure 4.

Cardiomyocyte and Erythrocyte Progenitors Are Increased in Prkci^−/− EBs

(A–F′) In (A, A′, E, and E′), Prkci^−/− EBs cultured without LIF have more ISL1 (cardiac progenitor marker) and α-ACTININ-positive cells compared to heterozygous EBs. (C) At day (d) 9, the pixel count ratio for ISL1 expression indicates that null EBs (green) have larger ISL1 populations than heterozygous EBs (black) (three independent experiments, n = 20 heterozygous EBs, 21 null EBs total; mean ± SEM; ^∗p < 0.05). In (B, B′, D, F, and F′), RA treatment induces more NKX2-5 (both nuclear and cytoplasmic) and α-ACTININ expression in null EBs. Arrows point to fibers in (F′).

(G) Null EBs (green) generate more beating EBs with RA treatment compared to heterozygous EBs (black) (four independent experiments; mean ± SEM; ^∗p < 0.05, ^∗∗∗p < 0.001).

(H) Dissociated null EBs of different stages (green) generate more erythrocytes in a colony-forming assay (CFU-E) (four independent experiments; mean ± SEM; ^∗∗p < 0.01).

(I) Examples of red colonies.

(J) Gene expression for primitive HSC markers is upregulated in null EBs (relative to heterozygous EBs) (three independent experiments; mean ± SEM).

Scale bars, 50 μm in (A, B, and E); 100 μm in (F), and 25 μm in (I).

See also Figure S4.