Cycling hypoxia up-regulates thioredoxin levels in human MDA-MB-231 breast cancer cells
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The thioredoxin system is a key cellular antioxidant system and is highly expressed in cancer cells, especially in more aggressive and therapeutic resistant tumors. We analysed the expression of the thioredoxin system in the MDA-MB-231 breast cancer cell line under conditions mimicking the tumor oxygen microenvironment. We grew breast cancer cells in either prolonged hypoxia or hypoxia followed by various lengths of reoxygenation and in each case cells were cultured with or without a hypoxic cycling preconditioning (PC) phase preceding the hypoxic growth. Flow cytometry-based assays were used to measure reactive oxygen species (ROS) levels. Cells grown in hypoxia showed a significant decrease in ROS levels compared to normoxic cells, while a significant increase in ROS levels over normoxic cells was observed after 4 hours of reoxygenation. The PC pre-treatment did not have a significant effect on ROS levels. Thioredoxin levels were also highest after 4 hours of reoxygenation, however cells subjected to PC pre-treatment displayed even higher thioredoxin levels. The high level of intracellular thioredoxin was also reflected on the cell surface. Reporter assays showed that activity of the thioredoxin and thioredoxin reductase gene promoters was also highest in the reoxygenation phase, although PC pre-treatment did not result in a significant increase over non-PC treated cells. The use of a dominant negative Nrf-2 negated the increased thioredoxin promoter activity during reoxygenation. This data suggests that the high levels of thioredoxin observed in tumors may arise due to cycling between hypoxia and reoxygenation.
Biochemical and Biophysical Research Communications
© 2012 Elsevier Inc. This is the author-manuscript version of this paper. Reproduced in accordance with the copyright policy of the publisher. Please refer to the journal's website for access to the definitive, published version.
Cancer Cell Biology
Gene Expression (incl. Microarray and other genome-wide approaches)
Biochemistry and Cell Biology not elsewhere classified