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

The principal cause of ?-thalassemia (or ?-thalassemia major) is the presence of free ? chains that cause severe damage to red blood cells. Clinical manifestations of the disease are attenuated in cases known as ?-thalassemia intermedia that do not require transfusions. While some cases of ?- thalassemia intermedia correlate with the presence of 2 ?⁺alleles, i.e. alleles that do not abolish synthesis of ? chains, other patients affected with this mild condition bear 2 ?⁰ alleles and for them the explanation for the mild phenotype is the synthesis of fetal ?-globin that persists in adult life and that associating with free ? chains produces fetal hemoglobin (HbF). It has been proposed that this unusual production might be due to polymorphic sequences located on the ?-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)_n in IVS2 of fetal genes, or in their proximal and distal promoters; the microsatellite (AT)_XT_Y at -540 of the ? globin gene; and the microsatellite (AT)_XN_Y(AT)_Z located in the hypersensitive site (HS2) of the ? globin locus control region LCR. We investigated these polymorphic sites and their linkage to the ?-thalassemic mutations typical of the ?⁰-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) ?-thalassemic patients affected with transfusion-dependent Thalassemia Major (genotypes ?⁰39/?⁰39 and ?⁰39/? ⁺-87); 2) patients homozygous for transfusion-independent Thalassemia Intermedia (?⁰39/?⁰39); 3) heterozygotes for different ?-thalassemic mutations (?⁰39, ?⁺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 G? and A? genes, and in the distal promoter of ? gene. None of the patients examined presented the ndHPFH mutations and in all we ruled out the presence of the ?-thalassemia mutations more widespread in Sardinia and the Mediterranean. We confirmed that the Thalassemia Intermedia (mild) patients were homozygous for the mutation ?⁰39 (CAG? TAG), the same presented in 90% of patients affected by Thalassemia Major. The ?⁰39 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 the ^A?^T allele, and two with haplotype I and the ^A?^I allele. The ?⁺-87 mutation located on chromosome with the polymorphic configuration 7a, resulted linked to the haplotype VIII of Orkin, while the locus ?⁺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.