title: Anion exchanger isoform 2 operates in parallel with Na(+)/H(+) exchanger isoform 1 during regulatory volume decrease of human cervical cancer cells.
abstract: Intracellular pH (pH(i)) homeostasis was investigated in human cervical cancer SiHa cells undergoing regulatory volume decrease (RVD) to determine which transport systems were involved. Using isoform-specific primers, mRNA transcripts of Na(+)/H(+) exchanger isoform 1 (NHE1) and isoform 3 were identified by reverse transcriptase polymerase chain reaction (RT-PCR) and the results confirmed by Western immunoblotting. From anion exchanger isoforms 1-3 (AE1-3), only the mRNA transcript of AE2 was identified by RT-PCR and the identity was confirmed by digestion with a specific restriction endonuclease. SiHa cells loaded with the fluorescent dye 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein and resuspended in isotonic media showed a stable pH(i). In contrast, a gradual internal acidification took place following resuspension in hypotonic media. The NHE inhibitors, HOE694 (10 microM) and amiloride (1 mM), showed a similar potency in enhancing the rate and extent of the hypotonicity-induced internal acidification. The absence of extracellular Na(+) also substantially enhanced the acidification during RVD. These results suggest that internal acidification during RVD is mainly compensated by the operation of NHE1. Extracellular Cl(-) was critically necessary for the pH(i) acidification during RVD. The hypotonicity-induced acidification was significantly attenuated by 100 microM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, a concentration inhibiting more than 90% AE activity. This indicates that AE2 mediates a net Cl(-) influx with compensating HCO(3)(-) efflux during RVD. We conclude that AE2 operates in parallel with NHE1 to regulate pH(i) during RVD of human cervical cancer cells.
authors.affiliation: University Laboratory of Physiology, Oxford, UK.
authors.forename: Meng Ru
authors.initials: MR
authors.lastname: Shen
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Wilkins
authors.forename: Cheng Yang
authors.initials: CY
authors.lastname: Chou
authors.forename: J Clive
authors.initials: JC
authors.lastname: Ellory
mesh_terms: D001457:Anion Exchange Protein 1, Erythrocyte / Q000378:metabolism*; D048429:Cell Size; D002584:Cervix Uteri / Q000166:cytology*; D005260:Female; D006706:Homeostasis; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008954:Models, Biological; D009363:Neoplasm Proteins / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002583:Uterine Cervical Neoplasms / Q000473:pathology
chemical_list: D001457:Anion Exchange Protein 1, Erythrocyte; D009363:Neoplasm Proteins; D017923:Sodium-Hydrogen Exchangers; C069139:growth factor-activatable Na-H exchanger NHE-1