Figure 4From: Transcriptional implications of intragenic DNA methylation in the oestrogen receptor alpha gene in breast cancer cells and tissuesIntragenic DNA methylation in ESR1 in exfoliated breast epithelial cells from expressed breast milk (EBM). (A) Flow cytometry of epithelial membrane antigen (EMA; median positive staining, 97.8%; 95.3%-99.4%; n = 60) and intracellular ESR1 (median, 90.5%; IQR, 84.4%-93.3%; n = 6). (B) qRT-PCR analysis of 11 EBM epithelial cell RNA samples showing high levels of ESR1 expression normalised against ER-pos MCF7 and ER-neg MDA-MB-231 cell lines, (range 2.2- 19-fold higher). (C) Methylation values from 450 K analysis of 36 DNA samples extracted from breast epithelial cells from expressed breast milk. The overall pattern matched those of ER-positive cancer cell lines, and regions of variability were found at regions that mapped to p2 and p7. (D) Validation of the methylation levels using pyrosequencing in a separate set of 53 EBM samples (black solid line). The pattern of methylation is very similar to that of the 450 K analysis of EBM samples and of ER-pos cancer cell lines (dashed line) with high levels of variability at p2 for both.Back to article page