Expression analyses of PR in differentiating HDSC and THESC. Quantitative real-time PCR and Western blot analyses were performed as described in Materials and Methods. Representative examples of Western blots are shown. A) PR mRNA expression at day 3, 6, 9, and 12 of cAMP or E2P4 stimulation. For relative quantification of total PR mRNA expression, n.c. of day three was arbitrarily set at 1 (calibrator). Bars indicate mean values ± SEM of seven and five different experiments performed in HDSC and THESC, respectively. PCR reactions were done in duplicates. * indicates p < 0.05 compared to the n.c. of the same day; n.s., not significant. B) Western blot analyses showing PR protein expression in total cellular extracts of dezidualizing HDSC. Specific signals detecting PR-A (90 kD) and PR-B (118 kD) in the absence or presence of cAMP or E2P4 (day 9) are depicted. Total extracts prepared from T47D cells were used as a positive control. Marker bands are indicated on the left. GAPDH was used as a loading control. C) Western blot analyses of cytoplasmic and nuclear extracts isolated from unstimulated and differentiated THESC which had been treated with cAMP and/or E2P4 for 9 days. Specific signals detecting PR-A (90 kD) and PR-B (118 kD) in T47D are depicted. Marker bands are indicated on the left and unspecific signals are indicated by stars. GAPDH and TopoIIβ were used as loading controls for cytoplasmic and nuclear extracts, respectively.