Abstract
Congenital haemolytic anaemias are inherited disorders caused by red blood cell membrane and cytoskeletal protein defects, deviant hemoglobin synthesis and metabolic enzyme deficiencies. In many cases, although the causing mutation might be known, the pathophysiology and the connection between the particular mutation and the symptoms of the disease are not completely understood. Thus effective treatment is lagging behind. As in many cases abnormal red blood cell cation content and cation leaks go along with the disease, by direct electrophysiological measurements of the general conductance of red blood cells, we aimed to assess if changes in the membrane conductance could be a possible cause. We recorded whole-cell currents from 29 patients with different types of congenital haemolytic anaemias: 14 with hereditary spherocytosis due to mutations in α-spectrin, β-spectrin, ankyrin and band 3 protein; 6 patients with hereditary xerocytosis due to mutations in Piezo1; 6 patients with enzymatic disorders (3 patients with glucose-6-phosphate dehydrogenase deficiency, 1 patient with pyruvate kinase deficiency, 1 patient with glutamate-cysteine ligase deficiency and 1 patient with glutathione reductase deficiency), 1 patient with β-thalassemia and 2 patients, carriers of several mutations and a complex genotype. While the patients with β-thalassemia and metabolic enzyme deficiencies showed no changes in their membrane conductance, the patients with hereditary spherocytosis and hereditary xerocytosis showed largely variable results depending on the underlying mutation.
Highlights
Haemolytic anaemias, characterized by the abnormal breakdown of red blood cells (RBCs), could be either acquired or inherited
All patients included in the study were genetically screened for mutations by next-generation sequencing and diagnosed with the following types of anemia: 14 patients were diagnosed with hereditary spherocytosis (HS), using golden standard techniques (EMAbinding, osmotic gradient ektacytometry and osmotic fragility test), 6 patients were diagnosed with hereditary xerocytosis, 6 patients had enzymatic disorders (3 patients with glucose-6-phosphate dehydrogenase deficiency, 1 patient with pyruvate kinase deficiency, 1 patient with glutamatecysteine ligase deficiency and 1 patient with glutathione reductase deficiency), 1 patient had β-thalassemia and 2 were carriers of several mutations and a complex genotype
We have compared the currents measured from the RBCs of our patients once with their transportation control, i.e., currents measured from the RBCs of a healthy subject, whose blood was delivered together with the blood of the patient, and once with a general, pooled, control, i.e., currents measured from the cells of all healthy subjects delivered throughout the study (Table 1)
Summary
Haemolytic anaemias, characterized by the abnormal breakdown of red blood cells (RBCs), could be either acquired or inherited. The latter are a diverse group of diseases that could be classified based on the affected RBC component into membranopathies, haemoglobinopathies, and enzymopathies (Dhaliwal et al, 2004). Most of the research has been done on membranopathies, understandable, primarily on the ones linked to mutations in ion channels or transporters such as band 3 protein, Rh-associated glycoprotein (RhAG), the glucose transporter GLUT1, Piezo and the Gardos channel (KCNN4) and accompanied by abnormal RBC cation content and disrupted volume homeostasis (Badens and Guizouarn, 2016). On most occasions, the RBC cation content linked to the particular mutation has been extensively described (e.g., Stewart et al, 2011 for R730C in band 3 protein or Fermo et al, 2017 for R352H in the Gardos channel) and the defective channels, when known, expressed and studied in heterologous systems (e.g., Glogowska et al, 2017 for a number of Piezo mutations) with a few exceptions (Stewart et al, 2011; Andolfo et al, 2013; Shmukler et al, 2014; Fermo et al, 2017; Rotordam et al, 2018), direct electrophysiological measurements of membrane conductance in mutated RBCs have been scarce
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