diethyl-maleate and Breast-Neoplasms

The system x. [

Topics: Amino Acid Transport System y+; Amino Acids, Dicarboxylic; Breast Neoplasms; Cell Line, Tumor; Female; Glutathione; Humans; Maleates; Positron-Emission Tomography; Reactive Oxygen Species; Receptors, Estrogen; Receptors, Progesterone; RNA, Messenger; Time Factors; Tissue Distribution; Tumor Burden; Xenograft Model Antitumor Assays

Glutathione(GSH) maintains an optimum cellular redox potential. Elevated levels of GSH render some types of cancer cells resistant against anti-cancer drugs. The aim of this study was to determine the effect of a thiol-depleting agent, diethylmaleate (DEM), on the sensitivity of human breast cancer cells to ADM.. The ADM-resistant human breast cancer MCF-7/ADM cell lines and ADM-sensitive MCF-7/S cell lines were treated by thiol-depleting agent DEM for 3 h respectively. The changes of sensitivity to ADM were then measured by MTT assay. The intracellular GSH contents were examined by fluorescent-spectrophotometry and the correlation between the changes of sensitivity to ADM and the intracellular GSH content was analyzed.. Treatment of MCF-7/ADM and MCF-7/S cells by 0.1 micromol/L DEM for 3 h decreased 37.4% and 29.7% of the intracellular GSH content respectively (P < 0.01). ADM also decreased intracellular GSH content in a ADM-concentration-dependent manner. The combined use of DEM and ADM depleted the intracellular GSH content in both cells significantly more than the sum of single use of ADM and DEM alone. The sensitivity of both cells to ADM increased with the decline of intracellular GSH content.. The depletion effect of DEM on the intracellular GSH could be enhanced by ADM and such depletion may be involved in the changes of the sensitivity of MCF/7 cells to ADM.

Topics: Antibiotics, Antineoplastic; Breast Neoplasms; Cell Line, Tumor; Doxorubicin; Drug Resistance, Neoplasm; Glutathione; Humans; Maleates

The currently available flow cytometric stains for cellular glutathione were evaluated, examining the labelling of both human and rodent cell lines under various conditions of concentration, time, and temperature. Procedures were used that depleted glutathione (GSH) while having a minimal effect on other cellular sulphydryls in order to estimate linearity and the extent of background staining. As previously reported, monochlorobimane was highly specific for GSH in rodent cells but failed to label human cells adequately because of its low affinity for human glutathione S-transferases. Higher concentrations of monochlorobimane achieved more complete labelling of the human cellular GSH pool but gave increased background fluorescence due to non-GSH binding. The analogue monobromobimane, which binds nonenzymatically to sulphydryls, reacted more readily with GSH than with protein sulphydryls and, provided that stain concentration and incubation time were controlled, gave reproducible staining of human cells with approximately 20% of total fluorescence due to background staining. Of the currently available stains for measuring GSH in human cells, monobromobimane is the agent of choice. Mercury orange also binds more readily to GSH than to protein, giving a degree of specificity, and it has the additional advantage of being excited at 488 nm. However, the reproducibility of staining with mercury orange was less consistent than that using monobromobimane, and a higher background fluorescence was seen. Two additional stains, o-phthaldialdehyde and chloromethyl fluorescein, could also be used to label cellular GSH, but both gave an unacceptably high level of background staining. It is recommended that flow cytometric GSH assays should routinely include a sample of cells that have been depleted of GSH in order to determine the extent of background labeling.

Topics: Animals; Breast Neoplasms; Bridged Bicyclo Compounds; Buthionine Sulfoximine; Carcinoma; Carcinosarcoma; CHO Cells; Colonic Neoplasms; Cricetinae; Ethylmaleimide; Eukaryotic Cells; Evaluation Studies as Topic; Flow Cytometry; Fluoresceins; Fluorescent Dyes; Glutathione; Humans; Maleates; Mammary Neoplasms, Experimental; Methionine Sulfoximine; Mice; o-Phthalaldehyde; Phenylmercury Compounds; Pyrazoles; Species Specificity; Tumor Cells, Cultured; Urinary Bladder Neoplasms