The fetal hemoglobin, Hb F, formed by two alpha and two gamma globin chains, is characteristic of the fetal development period and has diminished synthesis in the post-natal period. In some hereditary alterations Hb F remains elevated, as in delta-beta thalassemia, beta thalassemia and hereditary persistence of Hb F (HPFH). The synthesis of gamma globin can also be stimulated by external factors including leukemia, blood marrow transplantation and chemical induction, amongst others. Through the observation of elevated levels of Hb F in blood samples using electrophoretic procedures, we aimed at evaluating the amount of Hb F in blood donor samples in order to establish limits of normality in the population from São José do Rio Preto and region. Also using alkaline denaturation and high-pressure liquid chromatography (HPLC), we compared the applied methodologies and, in individuals with increased Hb F, we used molecular studies to identify mutations that modify the gamma globin gene expression. Blood samples from 119 donation candidates and, as a control group, 89 from individuals without symptoms of anemia or hematological findings and with increased Hb F, were analyzed. Of these 119 samples from blood donation candidates, 110 were used to trace the profile of normal Hb F, comparing the applied measurement methodologies, giving an average of 1.48% for alkaline denaturation and 0.6%, for HPLC. Statistical analysis using linear regression showed a significant difference between the two applied methodologies. HPLC was the best method to measure Hb F. In the 110 blood samples that were evaluated in the screening tests for abnormal hemoglobins 16.4% presented with alpha thalassemia, 0.9% with increased Hb F, 0.9% with beta thalassemia and 0.9% with a hemoglobin variant of the delta globin chain. The other nine blood donors presented with Hb F levels above 10% in electrophoresis with an average of 32.28% observed in alkaline denaturation and 26.4% in HPLC. Molecular analysis by PCR-ASO was carried out, screening the 16 mutations that give rise to beta-type thalassemia, in an attempt to identify a genetic defect that could explain the increase of Hb F. We found 5.3% of heterozygotes with the CD 6-A mutation and 1.75% with the CD 39, IVS1:6, -87 and IVS2:654 mutations. The results in this study suggested a necessity of better characterization of the hemoglobin profiles obtained by the classic methods and the importance of its molecular characterization.
Fetal Hemoglobin; abnormal hemoglobins; molecular diagnosis
