upregulated IDO expression in fibroblasts via the EP4/STAT3 pathway. (A) RMF-EG cells were treated with DMSO or PGE2 (2 μM) in 1% FCS medium for 48 hours. IDO mRNA (i) and protein (ii) were determined by quantitative RT-PCR and Western blotting. *P < 0.05. (B) RMF-EG cells were treated with 17-phenyl-trinor-PGE2 (an EP1 and EP3 receptor agonist), butaprost (an EP2 agonist), or PGE2-alcohol (an EP4 agonist) for 48 hours, and IDO expression was studied with quantitative RT-PCR. *P < 0.05. (C) RMF-EG cells were pretreated with EP4 shRNA for 24 hours and then cultured with MCF-7 (RMF-M) and COX-2-overexpressing MCF-7 (RMF-COX/M) cells for another 48 hours. (i) The IDO mRNA level was determined with quantitative RT-PCR. (ii) Protein level of IDO and EP4 was studied with Western blotting. *P < 0.05. (D) RMF-EG cells were pretreated with STAT3 siRNA for 24 hours and then cultured with PGE2-alcohol for another 48 hours. Protein levels of STAT3 and IDO were determined. (E) RMF-EG cells were transfected with pcDNA or STAT3 expression vector for 48 hours. Expression of STAT3 and IDO was studied with Western blotting.