title: Oxidative activation of K-Cl cotransport by diamide in erythrocytes from humans with red cell disorders, and from several other mammalian species.
abstract: Red blood cells (RBCs) from different mammalian species were investigated for the presence of diamide-induced oxidative activation of K-Cl cotransport reported to be present in sheep but absent in human RBCs. K efflux was measured in RBCs from human with hemoglobin (Hb) A or S, glucose-phosphate dehydrogenase (G6PDH) and a cytoskeletal deficiency, and from rat, mouse and rabbit. RBCs were incubated with diamide (0-1.0 mm) in K-free Cl or NO3 media of variable osmolalities (200-450 mOsM). Cl-dependent K efflux or K-Cl cotransport (estimated as the difference between K efflux rate constants in Cl and NO3) was activated by diamide in a sigmoidal fashion. Relative maximum K-Cl cotransport followed the sequence: human HbA (1) < rabbit (1.8) < sheep (6.9) < human HbS (9.5) approximately rat (9.7). Relative diamide concentrations for half maximal activation of K-Cl cotransport followed the sequence: sheep (1.9) > human Hb A (1) > rabbit (0.75) > human HbS and rat (0.67). Cell swelling in 200 mOsM doubled K-Cl cotransport in diamide, both in human HbA and S cells but reduced that in rat RBCs. In contrast, cell shrinkage at 450 mOsM obliterated K-Cl cotransport in human HbA and S but not in rat RBCs. Human RBCs with G6PDH and a cytoskeleton deficiency behaved like HbA RBCs. In mouse RBCs, diamide-activated K-Cl cotransport was 30% higher in isotonic than in hypotonic medium. In human HbA and S, and in low or high K sheep RBCs fractionated by Percoll density gradient, diamide increased the activity of K-Cl cotransport, an effect inversely correlated with cell density. Analysis of pooled data reveals that K-Cl cotransport accounted for about 80% of all K flux in Cl. There was a statistically significant correlation between K-Cl cotransport and K efflux in Cl (P < 0. 00001) and in NO3 (P < 0.00001). In conclusion, a diamide-activated K-Cl cotransport was present in human RBCs and in all other mammalian RBCs tested, with a large inter-, and for human and sheep, intraspecies variability for its maximum activity.
authors.affiliation: Department of Pharmacology & Toxicology, Wright State University School of Medicine, Dayton, Ohio 45401-0927, USA.
authors.forename: N C
authors.initials: NC
authors.lastname: Adragna
authors.forename: P K
authors.initials: PK
authors.lastname: Lauf
mesh_terms: D000755:Anemia, Sickle Cell / Q000503:physiopathology*; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D048429:Cell Size; D002648:Child; D002712:Chlorides / Q000378:metabolism; D003958:Diamide / Q000032:analysis / Q000494:pharmacology*; D004912:Erythrocytes / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D008297:Male; D051379:Mice; D010084:Oxidation-Reduction; D011188:Potassium / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D017207:Rats, Sprague-Dawley; D013045:Species Specificity; D013439:Sulfhydryl Reagents / Q000032:analysis / Q000494:pharmacology*; D027981:Symporters; D000096922:K Cl- Cotransporters
chemical_list: D002352:Carrier Proteins; D002712:Chlorides; D003958:Diamide; D011188:Potassium; D013439:Sulfhydryl Reagents; D027981:Symporters; D000096922:K Cl- Cotransporters