Expression of the transcriptional repressor ICER. (A) hMCs were stimulated with PGE₂ at the indicated concentrations for 3 hours. Lysates of the cells were resolved by SDS-PAGE, and immunoblotting was performed with a CREM-specific antiserum that recognizes ICER. (B) Time-dependent induction of ICER expression in response to PGE₂ or EP receptor-selective analogs. Lysates were generated from hMCs stimulated with the indicated agonist (1 μM each) for 5 minutes to 3 hours. The samples were subjected to SDS-PAGE and immunoblotting with anti-CREM. The same blot was stripped and probed for β-actin. Data in panels A and B are from single experiments representative of 3 performed for each. (C) Effects of H89 (10 μM), UO126 (5 μg/mL), AH6809 (50 μM), and AE-248 (10 μM) on the induced expression of ICER in response to stimulation with PGE₂ (1 μM) for 3 hours. Results in a second experiment were similar. (D) Quantitative densitometry showing induction of ICER following stimulation with the indicated agonists (1 μM) or their respective buffer controls for 3 hours. Results are the means ± SEM from a minimum of 4 separate experiments, except for the combined effect of H89 with UO126 (top), and the effects of the EP₂ and EP₄ receptor-selective antagonists AH6809 and AE-208, both of which represent mean ± 1/2 range from 2 of these experiments. ^*P < .05 relative to control (buffer or DMSO alone).