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pubmed:abstractText |
Cellular function is dependent on tight regulation of intracellular pH and numerous reports show cancer cells have abnormal pH values in the cytosol and organelles, such as lysosomes. 5-(and-6)-carboxyl seminaphthorhodafluor (SNARF-1) is a commonly used pH sensitive probe and was used here to determine cytosolic pH of HL-60 leukemia cells and a drug-resistant variant overexpressing multidrug-resistance related protein 1 (MRP1). Resistant cells accumulated significantly less SNARF-1 compared to parental cells but near control levels of probe accumulation were observed by preincubating cells with the specific MRP1 inhibitor MK571. Two new drug-resistant cell lines were generated following exposure to doxorubicin or daunorubicin and these upregulated MRP1 or P-glycoprotein expression, respectively. Experiments in these cells showed that reduced SNARF-1 accumulation was specific to MRP1 overexpression, as cells upregulating P-glycoprotein accumulated control levels of the probe. Confirmation that SNARF-1 is a MRP1 substrate was obtained using K562 and KG1a cells that have been shown to, respectively, constitutively express MRP1 and P-glycoprotein. Together, the data suggest that SNARF-1 is a substrate for MRP1 but not P-glycoprotein, and could therefore be used as a probe to distinguish between expression and activity of these 2 efflux proteins. Finally, we confirm that doxorubicin but not daunorubicin challenged MRP1 overexpressing HL-60 cells have elevated cytosolic pH.
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