Evidence for a protective role of the Gardos channel against hemolysis in murine spherocytosis

Abstract

It has been shown that mice with complete deficiency of all 4.1R protein isoforms (4.1-/-) exhibit moderate hemolytic anemia, with abnormal erythrocyte morphology (spherocytosis) and decreased membrane stability. Here, we characterized the Gardos channel function in vitro and in vivo in erythrocytes of 4.1-/- mice. Compared with wild-type, the Gardos channel of 4.1-/- erythrocytes showed an increase in Vmax (9.75 +/- 1.06 vs 6.08 +/- 0.09 mM cell x minute; P < .04) and a decrease in Km (1.01 +/- 0.06 vs 1.47 +/- 1.02 microM; P < .03), indicating an increased sensitivity to activation by intracellular calcium. In vivo function of the Gardos channel was assessed by the oral administration of clotrimazole, a well-characterized Gardos channel blocker. Clotrimazole treatment resulted in worsening of anemia and hemolysis, with decreased red cell survival and increased numbers of circulating hyperchromic spherocytes and microspherocytes. Clotrimazole induced similar changes in 4.2-/- and band 3+/- mice, indicating that these effects of the Gardos channel are shared in different models of murine spherocytosis. Thus, potassium and water loss through the Gardos channel may play an important protective role in compensating for the reduced surface-membrane area of hereditary spherocytosis (HS) erythrocytes and reducing hemolysis in erythrocytes with cytoskeletal impairments.

A novel erythroid anion exchange variant (Gly796Arg) of hereditary stomatocytosis associated with dyserythropoiesis

“…The Na-H exchange and K-Cl co-transport might be secondarily activated either by possible abnormal functional interactions between membrane proteins and the transmembrane ion transport pathway or by perturbations in signal transduction pathways modulating ion movements through the membrane, as previously described in mouse red cells genetically lacking band 4.1. 10,29 Since the activity of the membrane cation transport pathways is related to cyclic phosphorylation-dephosphorylation events by kinase phosphatases, we evaluated the membrane tyrosine phosphorylation profile in diseased red cells. In diseased red cells, we observed an increase of band 3 tyrosine phosphorylation, most likely due to increased membrane association of Syk and Lyn kinases which have already been reported to tyrosine-phosphorylate band 3.…”

“…With this procedure the internal sites for both transport systems were saturated. 10,12,13 The nystatin loading solution contained 70 mmol/L NaCl, 70 mmol/L KCl and 55 mmol/L sucrose. The Na/K pump activity was estimated as the ouabain-sensitive fraction of Na + efflux into a medium containing 130 mmol/L choline chloride and 10 mmol/L KCl.…”

“…Cation transport activities were estimated according to previously published methods. 10,11 Briefly, the maximal rates of Na/K pump and Na/K/Cl co-transport activities were measured in cells containing equal amounts of Na + and K + (50 mmol/L of cells, obtained with the nystatin technique). With this procedure the internal sites for both transport systems were saturated.…”

Concomitant inactivation of Rb and E2f8 in hematopoietic stem cells synergizes to induce severe anemia

“…38 Hemolysis may also arise from impaired glucose metabolism, hemoglobin synthesis, or defective erythrocyte membrane permeability. [39][40][41][42] However, it remains largely unknown how these critical proteins are regulated. In the present study, we have shown that inactivation of Rb and E2f8 in erythroid cells synergizes to trigger hemolysis ( Figure 2).…”

Neurodegeneration in the elderly – When the blood type matters: An overview of the McLeod syndrome with focus on hematological features

“…Especially, the proper function of Gardos channel is pivotal for the integrity and function of RBCs [75][76][77][78] . Gardos channels are ubiquitously expressed in body tissue 79 and their functionality may depend on intact XK protein.…”