Medicine (St Vincent's) - Theses

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Type: Masters Research thesis × Subject: cancer stem cell ×

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    Breast cancer stem cell properties in the PMC42 breast cancer cell line system
    WIDODO, EDWIN ( 2010)
    Epithelial to mesenchymal transition, EMT, is a developmental process that may be adopted by tumour cells to facilitate metastasis. EMT changes resulting in a more mesenchymal phenotype have been associated with a “migrating stem cell” pattern in breast and other carcinoma cells. PMC42 human breast cancer cells can be induced by epidermal growth factor (EGF) to a more mesenchymal state and adopt cancer stem cell characteristics. This study investigated the cancer stem cell properties in EGF-induced EMT in the PMC42 system, which consists of the predominantly mesenchymal PMC42-ET cell line and its epithelial-like subline, PMC42-LA. Quantitative reverse transcriptase polymerase chain reaction, qRT-PCR, was used to measure relative gene expression levels following EGF-induced EMT ofPMC42 cells in two- and three-dimensional cell cultures. Stem cell properties were evaluated through immunocytochemistry and two-channel fluorescence-activated cell sorting (FACS). CD44+/CD24- expression, a marker of breast cancer stem cells (BCSC), was measured after EGF-induced EMT. Ten ng/mL EGF treatment increased the number of cells expressing CD44+/CD24- in PMC42-ET but not in PMC42-LA cells as shown by FACS analysis. A mammosphere formation assay, which measures the ability of cells to form complex structures, was used to assess the functionality of the stem like properties exhibited by the different cell populations. This evidence of stemness was found in the BCSC-like population (CD44+/CD24-) of PMC42-ET cells. Expression of embryonic stem cell markers such as ABCG2 and ITGA6, EMT-BCSC related genes generated from EMT and BCSC bioinformatic comparison, and CD24-correlated genes (SerpinE1, EMP1, EMP3, AXL and VIM) were examined in the PMC42 system. EGF-induced EMT increased those gene expressions in the PMC42 system. PLP2 and VIL2, components of the Invasiveness Gene Signature derived from BCSC (Liu et al., 2007), showed no change in expression during EGF-induced EMT of PMC42. In conclusion, transcriptional changes demonstrating an increased stem-like or BCSC-like characteristic was observed in the PMC42 system following an EGF induced EMT. These results suggest the existence of a subpopulation of PMC42 cells which inherently resemble BCSC and provide new insights into the potential role of these subpopulations in EMT-associated metastasis.