Hemoglobin biosynthetic characterization of a classical and a mild-beta-thalassemia in Thailand

Abstract

Thalassemia if defined as a hereditary disorder with a decreased rate of a globin chain synthesis and presents as a hypochromic micro-cytic anemia without iron deficiency. β-thalassemia and hemoglobin E (Hb E) (∝₂ β₂[superscript 26Glu→Lys) are very prevalent in Thailand, with the incidence of 5 and 13 % respectively. Interaction of the two abnormal genes, known as β-thalassemia/Hb E disease, results in a clinical disorder of chronic hemolytic anemia, which poses not only in medical care but also in socio-economic problems. It is evident that the β-thalassemia can be classified into two types: classical β-thalassemia (β-thal₁), and mild β-thalassemia (β-thal₂). Upon the interaction with Hb E, β-thal₁/Hb E disease reveals hemoglobin types of E+F, while the β-thal₂/Hb E disease has hemoglobin type of E+F+A without a history of blood transfusion. It is very difficult to designate the β-thal₁ and β-thal₂ trait in adult because the hematologic data of both are similar interms of hypochromic microcytic res cells with elevated Hb A₂. This study attempts to characterize the heterozygotes by measureing the globin chain synthesis in reticulocytes in obligatory cases of β-thal₁ and β-thal₂ traits, and also to study the globin chain synthesis in Hb E traits. The hemoglobin synthesis in reticulocytes by incubation at 37℃ for 3 hours with 3H leucine were carried out. Globin prepared from washed red cells was fractionated by CMC Chromatography, in sodium phosphate buffer at pH 6.7 containing 8M urea and 0.05M mercaptoethanol. The globin chains were eluted by a linear gradient of sodium ions. The optical density and radioactivity of each peak were determined. The total radioactivity and specific activity (cpm/min/OD) of β/∝ globin chain ratio were calculated. A mean radioactivity β / ∝ ratio of 6 cases of normal controls was 0.92 ± 0.05 (or ∝ / β ratio = 1.08 ± 0.05). The hematolgic data of 7 cases of obligatory β-thal₁ trait were similar to that of 9 cases of obligatory β-thal₂ trait, but the mean radioactivity ratio of the β-thal₁ being 0.44 ± 0.14 revealed significant difference from that of the β-thal₂ trait with 0.50 ± 0.02. This indicated that the measurement of globin chain synthesis is a sensitive method for precise diagnosis of β-thal₁ and β-thal₂ trait. From the studies of incorporation in reticulocytes in β-thalassemia/ Hb E disease, the mean radioactivity of β chain/∝ chain ratio, β[superscript E]/∝ (no βA) of 6 patients with β-thal₁/Hb E disease was 0.40 ± 0.08, but the β chains/∝ chain ratio, β[superscript A] + β[superscript E]/∝ of 5 cases of β-thal₂/Hb E disease was 0.51 ± 0.07. Since the β / ∝ ratio of the β-thal₂ trait indicated suppression of β chain synthesis less than the β-thal₁ trait, this was consistent with the presence of β[superscript A] chain, presumably due to partial suppression by β-thal₂ gene, in β-thal₂ / Hb E disease. In Hb E trait, although the amount of Hb E is less than Hb A, the radioactivity of β chains/∝ chain ratio; β+β[superscript E] / ∝ was 0.97 ± 0.04, which was in normal ranges.