Upregulation of fibroblast growth factor receptor type 1 is stably maintained during oncogenic transforming growth factor β signaling. (A) Normal mammary epithelial (NME) cells were left untreated (before transforming growth factor treatment (pre-TGF)) or treated and allowed to recover from exogenous TGF-β1 treatment (post-TGF-Rec) as described in the Methods section. Global gene expression was assessed by microarray analyses. Data are results of RT-PCR analysis confirming stable upregulation of fibroblast growth factors 1 through 3 (FGFR1 to FGFR3) following TGF-β1 treatment and withdrawal. (B) Real-time PCR analyses showing the differential expression of FGFR1 to FGFR4 in control yellow fluorescent protein (YFP), nonmetastatic (NME) and lung metastatic (LM2) cell lines derived from the normal murine mammary gland (NMuMG) cells. (C) Control NMuMG (YFP) and NME cells were not stimulated (NS), stimulated with TGF-β1 (5 ng/ml) for 48 hours (TGF) and subsequently recovered for an additional 48 hours without exogenous ligand (Rec). Expression of FGFR1 was analyzed by real-time RT-PCR. Data in panels B and C are the mean expression values (±SD) of three independent experiments, resulting in the indicated P-values. (D) Human MCF-10A-derived breast cancer (BC) cells of increasing metastatic potential were analyzed by RT-PCR for their expression of FGFR1. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) served as a loading control. Data are representative of three independent experiments. (E) In silico analyses of the Gene Expression Omnibus data set [GEO:GSE20437] demonstrating significant increases in FGFR1 to FGFR4 in prophylactically removed breast tissue (Pro), as well as in ER-negative (ER-) and ER-positive (ER+) tumor samples, as compared to a cohort of normal (Nor) breast tissue samples. Data are the individual values for each sample resulting in the indicated mean values (±SE) and P values.