Erythrocytes anion transport and oxidative change in β‐thalassaemias

Abstract

beta-Thalassaemia is characterized by a decrease in globin beta-chain synthesis and an excess in free alpha-globin chains. This induces alterations in membrane lipids and proteins resulting from a reduction in spectrin/band 3 ratio, partial oxidation of band 4.1 and clustering of band 3. The membrane injury provokes hyperhaemolysis and bone marrow hyperplasia. The pathophysiology of thalassaemia is associated with iron overload that generates oxygen free radicals and oxidative tissue injury with ocular vessel alterations. The aim of this research is to investigate the influence of oxidative stress on band 3 efficiency, which is an integral membrane protein of RBCs (red blood cells). Band 3 protein, of which there are more than 1 million copies per cell, is the most abundant membrane protein in human RBCs. It mediates the anion exchange and acid-base equilibrium through the RBC membrane. Some experiments were performed on thalassaemic cells and beta-thalassaemia-like cells and tested for sulfate uptake. To test the antioxidant effect of Mg(2+), other experiments were performed using normal and pathological cells in the presence of Mg(2+). The oxidant status in thalassaemic cells was verified by increased K(+) efflux, by lower GSH levels and by increased G6PDH (glucose-6-phosphate dehydrogenase) activity. The rate constant of SO(4)(2-) uptake decreases in thalassaemic cells as well as in beta-thalassaemia-like cells when compared with normal cells. It increases when both cells are incubated with Mg(2+). Our data show that oxidative stress plays a relevant role in band 3 function of thalassaemic cells and that antioxidant treatment with Mg(2+) could reduce oxidative damage to the RBC membrane and improve the anion transport efficiency regulated by band 3 protein.

Sulphate and Chloride-Dependent Potassium Transport in Human Erythrocytes are Affected by Crude Venom from Nematocysts of the JellyfishPelagia noctiluca

“…In both cases, erythrocyte shrinkage was detected. Oxidative damage to erythrocyte membranes may increase the permeability to K + ions via increased activity of KCC, as reported by Crupi et al [26]. …”

“…For K + efflux measurements, according to Crupi et al [26] with slight modifications, erythrocytes were re-suspended to 20% hematocrit with either isotonic media A (150 mM choline chloride, 20 mM HEPES, 15 mM glucose, 0.1 mM ouabain, 307 mOsm/Kg H2O pH 7.4) or B (150 mM choline nitrate, 20 mM HEPES, 15 mM glucose, 0.1 mM ouabain, 307 mOsm/Kg H2O , pH 7.4) containing or not containing varying concentrations of venom or 2 mM N-ethylmaleimide (NEM) for 1 h at 25°C. The suspensions were spun at 4 °C for 5 min at 3000 x g and 50 µl of the supernatants were placed in 10 ml 15mM LiNO 3 .…”

Glutathione Redox System in β‐Thalassemia/Hb E Patients

“…The critical mechanism underlying the anemia is thought to be ROS overproduction catalyzed by iron-induced Fenton or Haber-Weiss reactions. Oxidant injury to membrane proteins causes cytoskeletal abnormalities including reduced spectrin/band 3 ratio, partial oxidation of band 4.1, phosphatidylserine (PS) exposure, and clustering of band 3 [13], leading to RBC deformity and complement-mediated erythrophagocytosis [14]. The hyperhemolysis state was reflected in low levels of hemoglobin at steady state and compensation for anemia was shown by an approximately 6-fold increase in the percentage of reticulocytes in β -thalassemia/Hb E patients.…”

Global transformation of erythrocyte properties via engagement of an SH2-like sequence in band 3

“…Not surprisingly, most of these diseases are also characterized by an abnormal erythrocyte membrane stability/morphology, an altered rate of erythrocyte glucose metabolism, and aberrant ion transport properties (36)(37)(38)(39)(40), raising the question of whether tyrosine phosphorylation of band 3 might also contribute to the development of these diseases. Indeed, the membrane blebbing and vesiculation seen in most of the above pathologies are now thought to contribute prominently to the vasoocclusive/thrombotic events that plague many of these patients (41), opening the question of whether a SYK kinase inhibitor might constitute a method for treating these diseases.…”