title: Regulation of intracellular pH in cancer cell lines under normoxia and hypoxia.
abstract: 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 O(2) 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(3)(-) 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(3)(-) 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.
authors.affiliation: Department of Physiology, Anatomy, and Genetics, Oxford University, Oxford, UK.
authors.forename: Alzbeta
authors.initials: A
authors.lastname: Hulikova
authors.forename: Adrian L
authors.initials: AL
authors.lastname: Harris
authors.forename: Richard D
authors.initials: RD
authors.lastname: Vaughan-Jones
authors.forename: Pawel
authors.initials: P
authors.lastname: Swietach
mesh_terms: D000143:Acids / Q000378:metabolism; D001692:Biological Transport; D027682:Cation Transport Proteins / Q000378:metabolism; D015687:Cell Hypoxia / Q000502:physiology*; D045744:Cell Line, Tumor; D015536:Down-Regulation; D045325:HCT116 Cells; D019073:HT29 Cells; D006367:HeLa Cells; D006706:Homeostasis; D006801:Humans; D006863:Hydrogen-Ion Concentration; D009369:Neoplasms / Q000378:metabolism* / Q000473:pathology; D010100:Oxygen / Q000378:metabolism; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism
chemical_list: D000143:Acids; D027682:Cation Transport Proteins; C490998:SLC9A1 protein, human; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers; D010100:Oxygen