Conflicts of interest: none to declare.
Original Research Article
Regulation of intracellular pH in cancer cell lines under normoxia and hypoxia†
Article first published online: 20 DEC 2012
Copyright © 2012 Wiley Periodicals, Inc.
Journal of Cellular Physiology
Volume 228, Issue 4, pages 743–752, April 2013
How to Cite
Hulikova, A., Harris, A. L., Vaughan-Jones, R. D. and Swietach, P. (2013), Regulation of intracellular pH in cancer cell lines under normoxia and hypoxia. J. Cell. Physiol., 228: 743–752. doi: 10.1002/jcp.24221
- Issue published online: 20 DEC 2012
- Article first published online: 20 DEC 2012
- Accepted manuscript online: 4 SEP 2012 08:22AM EST
- Manuscript Accepted: 23 AUG 2012
- Manuscript Received: 21 JUN 2012
- Royal Society
- Medical Research Council
- Association for International Cancer Research
- British Heart Foundation
- Cancer Research UK
Acid-extrusion by active transport is important in metabolically active cancer cells, where it removes excess intracellular acid and sets the intracellular resting pH. Hypoxia is a major trigger of adaptive responses in cancer, but its effect on acid-extrusion remains unclear. We studied pH-regulation under normoxia and hypoxia in eight cancer cell-lines (HCT116, RT112, MDA-MB-468, MCF10A, HT29, HT1080, MiaPaca2, HeLa) using the pH-sensitive fluorophore, cSNARF-1. Hypoxia responses were triggered by pre-incubation in low O2 or with the 2-oxoglutarate-dependent dioxygenase inhibitor dimethyloxalylglycine (DMOG). By selective pharmacological inhibition or transport-substrate removal, acid-extrusion flux was dissected into components due to Na+/H+ exchange (NHE) and Na+-dependent HCO transport. In half of the cell-lines (HCT116, RT112, MDA-MB-468, MCF10A), acid-extrusion on NHE was the dominant flux during an acid load, and in all of these, bar one (MDA-MB-468), NHE-flux was reduced following hypoxic incubation. Further studies in HCT116 cells showed that <4-h hypoxic incubation reduced NHE-flux reversibly with a time-constant of 1–2 h. This was not associated with a change in expression of NHE1, the principal NHE isoform. Following 48-h hypoxia, inhibition of NHE-flux persisted but became only slowly reversible and associated with reduced expression of the glycosylated form of NHE1. Acid-extrusion by Na+-dependent HCO transport was hypoxia-insensitive and comparable in all cell lines. This constitutive and stable element of pH-regulation was found to be important for setting and stabilizing resting pH at a mildly alkaline level (conducive for growth), irrespective of oxygenation status. In contrast, the more variable flux on NHE underlies cell-specific differences in their dynamic response to larger acid loads. J. Cell. Physiol. 228: 743–752, 2013. © 2012 Wiley Periodicals, Inc.