Figure 3.

In vitro and in vivo differentiation potential of hPSC-derived TSC

(A) Morphology, lineage-specific gene expression, and hCG secretion of primary (1049) and hPSC (H9)-derived TSCs differentiated into syncytiotrophoblast (STB). Bar: 312.5 μm. qPCR data were normalized to L19 and shown as fold change over undifferentiated (day 0/D0) 1049 TSCs. hCG secretion was normalized to ng of DNA. Both qPCR and ELISA data represent mean ± SD for n = 3 independent experiments. ^∗p < 0.05; ^∗∗p < 0.01 by Student’s t test.

(B) Morphology, lineage-specific gene expression, and flow-cytometric analysis for surface HLA-G expression of primary (1049) and hPSC (H9)-derived TSCs differentiated into extravillous trophoblast (EVT). Bar: 312.5 μm. qPCR data were normalized to L19 and shown as fold change over undifferentiated (day 0/D0) 1049 TSCs and represent mean ± SD for n = 3 independent experiments. ^∗p < 0.05; ^∗∗p < 0.01; ^∗∗∗p < 0.001 by Student’s t test. Flow-cytometric data are representative of 3 independent experiments.

(C and D) Tumors generated 10 days following injection of primary (1049) and hPSC (H9)-derived TSCs into NOD-SCID mice (representative of n = 2 independent experiments). (C) H&E staining shows the tumor cells invading through muscle (1049-TSC tumor) or forming a tumor with a necrotic center (H9-TSC), both characteristic of human trophoblastic tumors. Scale bars: 250 μm for low-power (left) and 50 μm for high-power (right) images. (D) Immunohistochemical staining of the same tumors using antibodies against EGFR, hCG, and HLAG shows positively stained cells (brown) in the TSC-derived lesions. Scale bars: 50 μm.