Effects of Y-27632 on pCTB differentiation.

<p>(A, B) pCTBs were cultured in the absence (A) and presence (B) of 10 μM Y-27632 for 96 h. Plasma membrane protein desmoplakin I was stained with Alexa Fluor 555, and nuclei were stained with DAPI. Arrowheads indicate fused, large, cytoplasmic multinucleated syncytiotrophoblast-like cells. Scale bar = 50 μm. (C) The fusion index was calculated using pCTBs from 6 different donors. Each point shows the mean of 10 fields in one well. (D-G) Total RNA was extracted from pCTBs cultured in the presence and absence of 10 μM Y-27632 for 96 h, and qPCR was used to quantify mRNA for GCM1 (D), syncytin-1 (E), syncytin-2 (F) and CGB (G). The experiments were performed in triplicate, and the data are representative of 3 separate experiments using pCTBs from 3 different donors. (H) pCTBs were cultured in the presence and absence of 10 μM Y-27632 for 96 h. Protein levels of hCG-β in the culture supernatant were measured by ELISA. The concentration of hCG-β was normalized against the protein concentration in the whole cell lysate (as mIU/ml/mg protein). The experiments were performed in triplicate, and the data are representative of 3 separate experiments using pCTBs from 3 different donors. Data are expressed as the mean ± SD. *, <i>P</i> < 0.05.