4 research outputs found
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Polycyclic aromatic hydrocarbons induce estrogen receptor-mediated luciferase, but do not upregulate estrogen responsive genes in MCF-7 cells
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PLD1 is overexpressed in an ER-negative MCF-7 cell line variant and a subset of phospho-Akt-negative breast carcinomas
We have used a novel variant of the human oestrogen receptor (ER)-positive MCF-7 cell line, TMX2-28, as a model to study breast cancer. TMX2-28 cells show no detectable levels of mRNA or protein expression for the ER and express basal cytokeratins (CKs) 5, 14, and 17. cDNA microarray comparison between TMX2-28 and its parent cell line, MCF-7, identified 1402 differentially expressed transcripts, one of which was, phospholipase D1 (PLD1). Using real-time RT-PCR, we confirmed that PLD1 mRNA levels are 10-fold higher in TMX2-28 cells than in MCF-7 cells. We next examined PLD1 expression in human breast carcinomas. Phospholipase D1 mRNA levels were higher in breast tumours that expressed high-mRNA levels of basal CKs 5 and/or 17, but PLD1 mRNA levels were not significantly higher in ER-negative tumours. Phospholipase D1 protein was overexpressed in 10 of 42 (24%) breast tumours examined by IHC. Phospholipase D1 was overexpressed in 6 of 31 ER-positive tumours and 4 of 11 ER-negative tumours. Phospholipase D1 was overexpressed in three of the four tumours that showed high CK5/17 expression. Five PLD1-positive tumours were negative for phospho-Akt expression, but positive for phospho-mammalian target of rapamycin (mTOR) expression. The other five PLD1-positive breast tumours showed positive expression for phospho-Akt; however, only two of these cases were positive for phospho-mTOR. In this study, we report that PLD1 and phospho-mTOR are coexpressed in a subset of phospho-Akt-negative breast carcinomas
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Use of an Aggressive MCF-7 Cell Line Variant, TMX2-28, to Study Cell Invasion in Breast Cancer
An estrogen receptor鈥搉egative variant of the MCF-7 breast cancer cell line, TMX2-28, was used as a model in which to study breast cancer cell invasion. Using a reconstituted basement membrane (Matrigel) assay to evaluate cell invasion, we determined that TMX2-28 cells are more invasive than MCF-7 cells and that the invasiveness of TMX2-28 is similar to that of the aggressive MDA-MB-231 breast cancer cell line. TMX2-28 cells displayed a rounded, epithelial cell鈥搇ike morphology, suggesting an amoeboid mode of cell invasion, in contrast to the mesenchymal mode of invasion characteristic of spindle-shaped, fibroblast-like MDA-MB-231 cells. Using real-time reverse transcription-PCR, we found that mitogen-inducible gene 2 (MIG2) is expressed at a 17-fold higher level in TMX2-28 cells than in nonaggressive MCF-7 cells and that MIG2 mRNA levels are low in the nontumorigenic human mammary epithelial cell line, 184. We determined that MIG2 plays a role in cell invasion by using small interfering RNA (siRNA) to suppress the expression of MIG2 mRNA levels in TMX2-28 cells. TMX2-28 cell invasion was reduced by 48% when the cells were transfected with siRNAs targeting MIG2, relative to cells transfected with siRNAs against glyceraldehyde-3-phosphate dehydrogenase. Finally, MIG2 expression was evaluated in reductive mammoplasty and breast tumor tissue. Although all 21 normal tissues from reduction mammoplasty showed immunoreactivity for MIG2, ranging from weak (62%) to strong (24%), only half of the 34 formalin-fixed breast tumors showed immunoreactivity for MIG2. Of these 17 positive cases, 10 were considered to overexpress MIG2 (moderate to strong staining). Examination of 30 frozen breast tumors supported the finding that MIG2 is overexpressed in a subset of breast cancers. We suggest that MIG2\u27s normal regulation and function are disrupted in breast cancer