title: Potential role of sodium-proton exchangers in the low concentration arsenic trioxide-increased intracellular pH and cell proliferation.
abstract: Arsenic main inorganic compound is arsenic trioxide (ATO) presented in solution mainly as arsenite. ATO increases intracellular pH (pHi), cell proliferation and tumor growth. Sodium-proton exchangers (NHEs) modulate the pHi, with NHE1 playing significant roles. Whether ATO-increased cell proliferation results from altered NHEs expression and activity is unknown. We hypothesize that ATO increases cell proliferation by altering pHi due to increased NHEs-like transport activity. Madin-Darby canine kidney (MDCK) cells grown in 5 mmol/L D-glucose-containing DMEM were exposed to ATO (0.05, 0.5 or 5 µmol/L, 0-48 hours) in the absence or presence of 5-N,N-hexamethylene amiloride (HMA, 5-100 µmol/L, NHEs inhibitor), PD-98059 (30 µmol/L, MAPK1/2 inhibitor), Gö6976 (10 µmol/L, PKCα, βI and μ inhibitor), or Schering 28080 (10 µmol/L, H(+)/K(+)ATPase inhibitor) plus concanamycin (0.1 µmol/L, V type ATPases inhibitor). Incorporation of [(3)H]thymidine was used to estimate cell proliferation, and counting cells with a hemocytometer to determine the cell number. The pHi was measured by fluorometry in 2,7-bicarboxyethyl-5,6-carboxyfluorescein loaded cells. The Na(+)-dependent HMA-sensitive NHEs-like mediated proton transport kinetics, NHE1 protein abundance in the total, cytoplasm and plasma membrane protein fractions, and phosphorylated and total p42/44 mitogen-activated protein kinases (p42/44(mapk)) were also determined. Lowest ATO (0.05 µmol/L, ~0.01 ppm) used in this study increased cell proliferation, pHi, NHEs-like transport and plasma membrane NHE1 protein abundance, effects blocked by HMA, PD-98059 or Gö6976. Cell-buffering capacity did not change by ATO. The results show that a low ATO concentration increases MDCK cells proliferation by NHEs (probably NHE1)-like transport dependent-increased pHi requiring p42/44(mapk) and PKCα, βI and/or μ activity. This finding could be crucial in diseases where uncontrolled cell growth occurs, such as tumor growth, and in circumstances where ATO, likely arsenite, is available at the drinking-water at these levels.
authors.affiliation: Cellular Physiology Laboratory, Biomedical Department, Faculty of Health Sciences, Universidad de Antofagasta, Antofagasta, Chile.
authors.forename: Carmen
authors.initials: C
authors.lastname: Aravena
authors.forename: Ana R
authors.initials: AR
authors.lastname: Beltrán
authors.forename: Marcelo
authors.initials: M
authors.lastname: Cornejo
authors.forename: Viviana
authors.initials: V
authors.lastname: Torres
authors.forename: Emilce S
authors.initials: ES
authors.lastname: Díaz
authors.forename: Enrique
authors.initials: E
authors.lastname: Guzmán-Gutiérrez
authors.forename: Fabián
authors.initials: F
authors.lastname: Pardo
authors.forename: Andrea
authors.initials: A
authors.lastname: Leiva
authors.forename: Luis
authors.initials: L
authors.lastname: Sobrevia
authors.forename: Marco A
authors.initials: MA
authors.lastname: Ramírez
mesh_terms: D000584:Amiloride / Q000031:analogs & derivatives; D000704:Analysis of Variance; D000818:Animals; D000077237:Arsenic Trioxide; D001152:Arsenicals / Q000378:metabolism* / Q000494:pharmacology; D015153:Blotting, Western; D002227:Carbazoles; D002452:Cell Count; D002458:Cell Fractionation; D049109:Cell Proliferation / Q000187:drug effects*; D004285:Dogs; D005419:Flavonoids; D005470:Fluorometry; D006863:Hydrogen-Ion Concentration; D007093:Imidazoles; D018942:Macrolides; D061985:Madin Darby Canine Kidney Cells; D010087:Oxides / Q000378:metabolism* / Q000494:pharmacology; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
chemical_list: D001152:Arsenicals; D002227:Carbazoles; D005419:Flavonoids; D007093:Imidazoles; D018942:Macrolides; D010087:Oxides; D017923:Sodium-Hydrogen Exchangers; C035235:Sch 28080; C081021:Go 6976; C057360:5-(N,N-hexamethylene)amiloride; D000584:Amiloride; C036978:concanamycin A; D000077237:Arsenic Trioxide; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one