Abstract. To establish a model of endocrine resistant breast cancer that is associated with loss of estrogen receptor (ER), MCF7 cells were transfected with several plasmid constructs intended to produce intracellular double stranded hairpin RNA to be processed into siRNA directed against different regions of the ER· mRNA. Stably transformed cells were propagated in long-term culture. One of these lines, designated pII, was selected for further analysis. pII cells exhibited reduced levels of ER· mRNA and protein as well as several estrogen-regulated genes assessed by real-time PCR and were unresponsive to addition of estradiol and tamoxifen.
BackgroundAstrocytomas are cancers of the brain in which high levels of extracellular glutamate plays a critical role in tumor growth and resistance to conventional treatments. This is due for part to a decrease in the activity of the glutamate transporters, i.e. the Excitatory Amino Acid Transporters or EAATs, in relation to their nuclear mislocalization in astrocytoma cells. Although non-astrocytoma cancers express EAATs, the localization of EAATs and the handling of L-glutamate in that case have not been investigated.MethodsWe looked at the cellular localization and activity of EAATs in human astrocytoma and non-astrocytoma cancer cells by immunofluorescence, cell fractionation and L-glutamate transport studies.ResultsWe demonstrated that the nuclear mislocalization of EAATs was not restricted to astrocytoma and happened in all sub-confluent non-astrocytoma cancer cells we tested. In addition, we found that cell-cell contact caused the relocalization of EAATs from the nuclei to the plasma membrane in all human cancer cells tested, except astrocytoma.ConclusionsTaken together, our results demonstrated that the mislocalization of the EAATs and its associated altered handling of glutamate are not restricted to astrocytomas but were also found in human non-astrocytoma cancers. Importantly, we found that a cell contact-dependent signal caused the relocalization of EAATs at the plasma membrane at least in human non-astrocytoma cancer cells, resulting in the correction of the altered transport of glutamate in such cancer cells but not in astrocytoma.
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