Figure 2.

RMC-4550 and SHP2 knockdown inhibit artificially induced decidualization in hESCs. (A) SHP2 activity assay was performed. The hESCs were incubated for 1 h with or without RMC-4550. Error bars denote the s.d. (n = 3). (B) Real-time PCR analysis of the transcriptional levels of IGFBP1 and PRL in hESCs with or without induced decidualization or treatment with RMC-4550. Error bars denote the s.d. (n = 3). Data were calibrated to the most highly expressing sample in hESCs (yellow color). (C) Immunoblotting of IGFBP1 and PRL protein levels in hESCs with or without induced decidualization or treatment with RMC-4550. Bar figures represent the ratios of densities (IGFBP1/B-actin and PRL/B-actin). Error bars denote the s.d. (n = 3). (D–E) The morphology of hESCs after decidualization with and without RMC-4550 treatment. The number of round cells from five different fields was counted and plotted as a histogram. Bar figures represent the ratios of round cells. Scale bars = 50 μm. (F) Real-time PCR analysis of the transcriptional levels of FOXO1, HOXA10, CEBPB, and STAT3 in hESCs with or without induced decidualization or treatment with RMC-4550. Error bars denote the s.d. (n = 3). Data were calibrated to the most highly expressing sample in hESCs-4d (red color). (G) Real-time PCR analysis of the transcriptional levels of SHP2, IGFBP1, and PRL in hESCs, in which decidualization was induced for 4 days after transfection with sh-SHP2 or sh-Ctrl for 72 h. Error bars denote the s.d. (n = 3). Data were calibrated to the most highly expressing sample in sh-Ctrl (red color). (H) Immunoblotting of SHP2, IGFBP1, and PRL protein levels in hESCs, in which decidualization was induced for 4 days after transfection with sh-SHP2 or sh-Ctrl for 72 h. Bar figures represent the ratios of densities (SHP2/B-actin, IGFBP1/B-actin, and PRL/B-actin). Error bars denote the s.d. (n = 3). (I–J) The morphology of hESCs after decidualization, in which decidualization was induced for 4 days after transfection with sh-SHP2 or sh-Ctrl for 72 h. The number of round cells from five different fields were counted and plotted as histogram. Bar figures represent the ratios of round cells. Scale bars = 50 μm. K. Real-time PCR analysis of SHP2 and the transcriptional levels of FOXO1, HOXA10, CEBPB and STAT3 in hESCs, in which decidualization was induced for 4 days after transfection with sh-SHP2 or sh-Ctrl for 72 h. Error bars denote the s.d. (n = 3). Data were calibrated to the most highly expressing sample in sh-Ctrl (red color). ^*P < 0.05, ^**P < 0.01, and ^***P < 0.001.