Evaluation of the free α-hemoglobin pool in red blood cells: a new test providing a scale of β-thalassemia severity.
β-Thalassemias are characterized by an imbalance of globin chains with an excess of α-chains which precipitates in erythroid precursors and red blood cells (RBCs) leading to inefficient erythropoiesis. The severity of the disease correlates with the amount of unpaired α-chains.Our goal was to develop a simple test for evaluation of the free α-hemoglobin pool present in RBC lysates. Alpha-Hemoglobin Stabilizing Protein (AHSP), the chaperone of α-Hb, was used to trap excess a-Hb. A recombinant GST-AHSP fusion protein was bound to an affinity micro-column and then incubated with hemolysates of patients. After washing, the α-Hb was quantified by spectrophotometry in the elution fraction. This assay was applied to 54 patients: 28 without apparent Hb disorder, 20 β-thalassemic and 6 α-thalassemic. The average value of free α-Hb pool was 93 ± 21 ppm (ng of free α-Hb per mg of Hb subunits)in patients without Hb disorder, while it varies from 119 to 1,756 ppm, in β-thalassemic patients and correlated with genotype. In contrast,the value of the free α-Hb pool was decreased in α-thalassemic patients (65 ± 26 ppm). This assay may help to characterize β-thalassemia phenotypes and to follow the evolution of the globin chain imbalance(α/β+γ ratio) in response to treatment.
Institut National de la Santé et de la Recherche Médicale, Université Paris XI, Le Kremlin-Bicêtre, France.
Corinne
C
Vasseur
Serge
S
Pissard
Elisa
E
Domingues-Hamdi
Michael C
MC
Marden
Frédéric
F
Galactéros
Véronique
V
Baudin-Creuza
D000328:Adult; D000369:Aged, 80 and over; D001798:Blood Proteins / Q000235:genetics / Q000378:metabolism; D002248:Carbon Monoxide / Q000378:metabolism; D002846:Chromatography, Affinity; D004912:Erythrocytes / Q000378:metabolism*; D005260:Female; D005838:Genotype; D006454:Hemoglobins / Q000032:analysis* / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008297:Male; D008875:Middle Aged; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism; D021122:Protein Subunits / Q000097:blood / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012720:Severity of Illness Index; D055815:Young Adult; D017085:alpha-Thalassemia / Q000097:blood / Q000235:genetics / Q000378:metabolism; D017086:beta-Thalassemia / Q000097:blood* / Q000235:genetics / Q000378:metabolism / Q000503:physiopathology
C468768:AHSP protein, human; D001798:Blood Proteins; D006454:Hemoglobins; D018832:Molecular Chaperones; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; D002248:Carbon Monoxide