Journal: Cancer Research

Article Title: Xenoestrogen-Induced Epigenetic Repression of microRNA-9-3 in Breast Epithelial Cells

doi: 10.1158/0008-5472.can-08-4914

Figure Lengend Snippet: Figure 1. Preexposure of MDECs to diethylstilbestrol and immunofluorescence analysis of nuclear ER-a. A, treatment scheme. Breast progenitor cells were propagated as nonadherent spherical colonies, called mammospheres, and treated with 70 nmol/L diethylstilbestrol or DMSO solvent control for 3 wk. To induce differentiation, cells were seeded on a collagen substratum in the absence of diethylstilbestrol for 3 wk. Expression profiling (microRNA microarray), immunofluorescence image (IFA), and epigenetic [chromatin immunoprecipitation-PCR (ChIP-PCR) and MassARRAY] analyses were then done on the progeny epithelial cells. B, increased internalization of ER-a in diethylstilbestrol-preexposed MDECs. After the preexposure to diethylstilbestrol (DES) or DMSO, MDECs were subjected to immunofluorescence analysis. The percentage of subcellular localization of ER-a–positive cells, independently scored by two researchers, is shown in the bar chart. Columns, mean of five independent sets of MDEC samples; bars, SE. P < 0.001 (Student’s t test). C, nuclear trafficking of ER-a in MDECs on diethylstilbestrol treatment. DMSO-preexposed MDECs were exposed to 70 nmol/L diethylstilbestrol in the indicated time points. Translocation of ER-a protein (green) from the cytoplasm to the nucleus was observed, suggesting functional estrogen signaling. Nuclei were stained with 4¶,6-diamidino-2-phenylindole (DAPI; blue). The turquoise signals (merged) highlight the localization of ER-a in the nucleus.

Article Snippet: The MassARRAY platform was used to perform quantitative methylation analysis (Sequenom).

Techniques: Immunofluorescence, Solvent, Control, Expressing, Microarray, Chromatin Immunoprecipitation, Translocation Assay, Functional Assay, Staining