Studio sui determinanti genetici correlati con la sintesi di emoglobina fetale nella vita adulta

The principal cause of β-thalassemia (or β-thalassemia major) is the presence of free α chains that cause severe damage to red blood cells. Clinical manifestations of the disease are attenuated in cases known as β-thalassemia intermedia that do not require transfusions. While some cases of β- thalassemia intermedia correlate with the presence of 2 β+alleles, i.e. alleles that do not abolish synthesis of β chains, other patients affected with this mild condition bear 2 β0alleles and for them the explanation for the mild phenotype is the synthesis of fetal γ-globin that persists in adult life and that associating with free α chains produces fetal hemoglobin (HbF). It has been proposed that this unusual production might be due to polymorphic sequences located on the β-globin cluster that might affect transcription factor binding. These polymorphic sites are represented by single nucleotide polymorphisms (SNPs) and microsatellites located in intragenic regions like (TG)m(CG)nin IVS2 of fetal genes, or in their proximal and distal promoters; the microsatellite (AT)XTYat -540 of the β globin gene; and the microsatellite (AT)XNY(AT)Zlocated in the hypersensitive site (HS2) of the β globin locus control region LCR. We investigated these polymorphic sites and their linkage to the β-thalassemic mutations typical of the β0-Sardinian thalassemic patients. In fact, 10% of them manifest intermediate thalassemia due to persistent production of HbF in adults. We analyzed four categories of patients: 1) β-thalassemic patients affected with transfusion-dependent Thalassemia Major (genotypes β039/β039 and β039/β+-87); 2) patients homozygous for transfusion-independent Thalassemia Intermedia (β039/β039); 3) heterozygotes for different β-thalassemic mutations (β039, β+IVS1-nt6) that present high levels of HbF; and 4) normal subjects. We identified a new (never described in literature) polymorphic microsatellite in the IVS2 of the Gγ and Aγ genes, and in the distal promoter of β gene. None of the patients examined presented the ndHPFH mutations and in all we ruled out the presence of the α-thalassemia mutations more widespread in Sardinia and the Mediterranean. We confirmed that the Thalassemia Intermedia (mild) patients were homozygous for the mutation β039 (CAG→ TAG), the same presented in 90% of patients affected by Thalassemia Major. The β039 mutation resulted distributed on different polymorphic configurations, none of which identical to those described in literature: two of them in association with the II haplotype of Orkin and theAγTallele, and two with haplotype I and theAγIallele. The β+-87 mutation located on chromosome with the polymorphic configuration 7a, resulted linked to the haplotype VIII of Orkin, while the locus β+IVS1-nt6 (8a) to the haplotipe II. Presence of the same polymorphic configuration (1a) on 75% of the chromosomes of the intermediate patients and 67,5% of the transfused patients rules out the hypothesis that this configuration can carry out a role of modulation of the expression of HbF in Sardinian patients.