scholarly journals Genetic and Epigenetic Therapies for β-Thalassaemia by Altering the Expression of α-Globin Gene

2021 ◽  
Vol 3 ◽  
Author(s):  
Sachith Mettananda
Keyword(s):  
Genome Editing ◽  
Current Knowledge ◽  
Globin Gene ◽  
Genetic Data ◽  
Histone Deacetylation ◽  
Globin Genes ◽  
Clinical Genetic ◽  
Globin Chains ◽  
Epigenetic Drug

β-Thalassaemia is caused by over 300 mutations in and around the β-globin gene that lead to impaired synthesis of β-globin. The expression of α-globin continues normally, resulting in an excess of α-globin chains within red blood cells and their precursors. These unpaired α-globin chains form unstable α-hemichromes that trigger cascades of events to generate reactive oxygen species, leading to ineffective erythropoiesis and haemolysis in patients with β-thalassaemia. The clinical genetic data reported over several decades have demonstrated how the coinheritance of α-thalassaemia ameliorates the disease phenotype of β-thalassaemia. Thus, it is evident that down-regulation of the α-globin gene expression in patients with β-thalassaemia could ameliorate or even cure β-thalassaemia. Over the last few years, significant progress has been made in utilising this pathway to devise a cure for β-thalassaemia. Most research has been done to alter the epigenetic landscape of the α-globin locus or the well-characterised distant enhancers of α-globin. In vitro, pre-clinical studies on primary human erythroid cells have unveiled inhibition of histone lysine demethylation and histone deacetylation as potential targets to achieve selective downregulation of α-globin through epigenetic drug targeting. CRISPR based genome editing has been successfully used in vitro to mutate α-globin genes or enhancers of α-goblin to achieve clinically significant knockdowns of α-globin to the levels beneficial for patients with β-thalassaemia. This review summarises the current knowledge on the regulation of human α-globin genes and the clinical genetic data supporting the pathway of targeting α-globin as a treatment for β-thalassaemia. It also presents the progress of epigenetic drug and genome editing approaches currently in development to treat β-thalassaemia.

scholarly journals Erythropoiesis in Malaria Infections and Factors Modifying the Erythropoietic Response

Anemia ◽  
2016 ◽  
Vol 2016 ◽  
pp. 1-8 ◽  
Author(s):  
Vrushali A. Pathak ◽  
Kanjaksha Ghosh
Keyword(s):  
Malaria Infection ◽  
Globin Gene ◽  
Experimental Studies ◽  
Collective Effect ◽  
Ineffective Erythropoiesis ◽  
Globin Chains ◽  
In Vitro Cell Cultures ◽  
Globin Gene Expression ◽  
Erythropoietic Response

Anemia is the primary clinical manifestation of malarial infections and is responsible for the substantial rate of morbidity. The pathophysiology discussed till now catalogued several causes for malarial anemia among which ineffective erythropoiesis being remarkable one occurs silently in the bone marrow. A systematic literature search was performed and summarized information on erythropoietic response upon malaria infection and the factors responsible for the same. This review summarizes the clinical and experimental studies on patients, mouse models, and in vitro cell cultures reporting erythropoietic changes upon malaria infection as well as factors accountable for the same. Inadequate erythropoietic response during malaria infection may be the collective effect of various mediators generated by host immune response as well as parasite metabolites. The interplay between various modulators causing the pathophysiology needs to be explored further. Globin gene expression profiling upon malaria infection should also be looked into as abnormal production of globin chains could be a possible contributor to ineffective erythropoiesis.

Effective erythropoiesis and HbF reactivation induced by kit ligand in β-thalassemia

Blood ◽  
2008 ◽  
Vol 111 (1) ◽  
pp. 421-429 ◽  
Author(s):  
Marco Gabbianelli ◽  
Ornella Morsilli ◽  
Adriana Massa ◽  
Luca Pasquini ◽  
Paolo Cianciulli ◽  
...  
Keyword(s):  
Fetal Hemoglobin ◽  
Thalassemia Major ◽  
Globin Genes ◽  
Erythroid Progenitors ◽  
Ineffective Erythropoiesis ◽  
Kit Ligand ◽  
Globin Chains ◽  
In Vitro Response ◽  
Globin Synthesis

In human β-thalassemia, the imbalance between α- and non–α-globin chains causes ineffective erythropoiesis, hemolysis, and anemia: this condition is effectively treated by an enhanced level of fetal hemoglobin (HbF). In spite of extensive studies on pharmacologic induction of HbF synthesis, clinical trials based on HbF reactivation in β-thalassemia produced inconsistent results. Here, we investigated the in vitro response of β-thalassemic erythroid progenitors to kit ligand (KL) in terms of HbF reactivation, stimulation of effective erythropoiesis, and inhibition of apoptosis. In unilineage erythroid cultures of 20 patients with intermedia or major β-thalassemia, addition of KL, alone or combined with dexamethasone (Dex), remarkably stimulated cell proliferation (3-4 logs more than control cultures), while decreasing the percentage of apoptotic and dyserythropoietic cells (<5%). More important, in both thalassemic groups, addition of KL or KL plus Dex induced a marked increase of γ-globin synthesis, thus reaching HbF levels 3-fold higher than in con-trol cultures (eg, from 27% to 75% or 81%, respectively, in β-thalassemia major). These studies indicate that in β-thalassemia, KL, alone or combined with Dex, induces an expansion of effective erythropoiesis and the reactivation of γ-globin genes up to fetal levels and may hence be considered as a potential therapeutic agent for this disease.

scholarly journals Sickle Cell Anemia and Babesia Infection

Pathogens ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 1435
Author(s):  
Divya Beri ◽  
Manpreet Singh ◽  
Marilis Rodriguez ◽  
Karina Yazdanbakhsh ◽  
Cheryl Ann Lobo
Keyword(s):  
Sickle Cell ◽  
Sickle Cell Anemia ◽  
Current Knowledge ◽  
Globin Gene ◽  
Environmental Niche ◽  
Parasite Resistance ◽  
The Usa ◽  
Human Infections

Babesia is an intraerythrocytic, obligate Apicomplexan parasite that has, in the last century, been implicated in human infections via zoonosis and is now widespread, especially in parts of the USA and Europe. It is naturally transmitted by the bite of a tick, but transfused blood from infected donors has also proven to be a major source of transmission. When infected, most humans are clinically asymptomatic, but the parasite can prove to be lethal when it infects immunocompromised individuals. Hemolysis and anemia are two common symptoms that accompany many infectious diseases, and this is particularly true of parasitic diseases that target red cells. Clinically, this becomes an acute problem for subjects who are prone to hemolysis and depend on frequent transfusions, like patients with sickle cell anemia or thalassemia. Little is known about Babesia’s pathogenesis in these hemoglobinopathies, and most parallels are drawn from its evolutionarily related Plasmodium parasite which shares the same environmental niche, the RBCs, in the human host. In vitro as well as in vivo Babesia-infected mouse sickle cell disease (SCD) models support the inhibition of intra-erythrocytic parasite proliferation, but mechanisms driving the protection of such hemoglobinopathies against infection are not fully studied. This review provides an overview of our current knowledge of Babesia infection and hemoglobinopathies, focusing on possible mechanisms behind this parasite resistance and the clinical repercussions faced by Babesia-infected human hosts harboring mutations in their globin gene.

scholarly journals Characterization of the DNase I hypersensitive site 3' of the human beta globin gene domain

Blood ◽  
1993 ◽  
Vol 81 (10) ◽  
pp. 2781-2790
Author(s):  
DE Fleenor ◽  
RE Kaufman
Keyword(s):  
Topoisomerase Ii ◽  
Globin Gene ◽  
Dnase I ◽  
Globin Genes ◽  
Beta Globin ◽  
Hypersensitive Site ◽  
Mobility Shift ◽  
Enhancer Activity ◽  
Beta Globin Gene

The members of the human beta globin gene family are flanked by strong DNase I hypersensitive sites. The collection of sites 5' to the epsilon globin gene is able to confer high levels of expression of linked globin genes, but a function has not been assigned to the site 3' to the beta globin gene (3'HS1). Our analysis of this DNase I super hypersensitive site shows that the region is composed of multiple DNase I sites. By examination of the DNA sequence, we have determined that the region is very A/T-rich and contains topoisomerase II recognition sequences, as well as several consensus binding motifs for GATA-1 and AP-1/NF-E2. Gel mobility shift assays indicate that the region can interact in vitro with GATA-1 and AP-1/NF-E2, and functional studies show that the region serves as a scaffold attachment region in both erythroid and nonerythroid cell lines. Whereas many of the physical features of 3'HS1 are shared by 5'HS2 (a component of the 5' locus control region), transient expression studies show that 3' HS1 does not share the erythroid-specific enhancer activity exhibited by 5'HS2.

Recapitulation of the Fetal Pattern of Expression of the Iγ and Vγ-Globin Genes in Cultured Baboon CD34+ Bone Marrow Cells.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1590-1590
Author(s):  
Donald Lavelle ◽  
Kestis Vaitkus ◽  
Mahipal Singh ◽  
Maria Hankewych ◽  
Joseph DeSimone
Keyword(s):  
Bone Marrow ◽  
Amino Acid ◽  
Bone Marrow Cells ◽  
Globin Gene ◽  
Single Amino Acid ◽  
Globin Genes ◽  
Globin Chain ◽  
Hemoglobin Switching ◽  
Globin Chains

Abstract The human Gγ-globin and Aγ-globin genes differ by the presence of a single amino acid, either glycine or alanine, at position 136. The ratio of Gγ/Aγ-globin expression is approximately 7/3 at birth and changes to 2/3 in the adult. The mechanism responsible for this developmental switch is unknown. In the baboon, the duplicated γ-globin genes differ by the presence of a single amino acid at position 75. The Iγ-globin gene contains isoleucine at position 75, while the Vγ-globin gene contains valine at this position. The ratio of expression of the Iγ and Vγ-globin chains also differs in the fetal and adult stages. The Iγ/Vγ ratio is 3/2 in the fetus and 2/3 in the adult. Thus the pattern of expression of the baboon Iγ-globin gene is analogous to the human Gγ-globin gene, and that of the Vγ-globin gene is analogous to the human Aγ-globin gene. During stress erythropoiesis, moderately increased HbF levels are observed (5–10% HbF) and the Iγ/Vγ-globin chains are expressed in the characteristic adult ratio. Decitabine treatment reactivates HbF expression to high levels (50–70% HbF) and Iγ/Vγ ratios of approximately 1:1 have been observed following decitabine treatment. Thus decitabine treatment alters the Iγ/Vγ ratio but does not cause a complete reversion to the fetal pattern of expression. HbF is also reactivated to high levels in cultured baboon BFUe. In this investigation the pattern of expression of the Iγ- and Vγ-globin genes in cultured baboon CD34+ bone marrow (BM) cells was analyzed to determine whether reactivation of HbF in culture was associated with a change in the pattern of expression of the Iγ-and Vγ-globin genes. CD34+ cells were enriched from baboon BM using the 12.8 monoclonal antibody in combination with immunomagnetic microbead columns (Miltenyi) and cultured in Iscove’s media supplemented with 30% fetal bovine serum, stem cell factor (SCF; 200ng/ml), erythropoietin (EPO; 2U/ml), and dexamethasone (Dex; 1μM). The pattern of globin chain expression in d12 cultures, cord blood (CB) of a 58d fetus, and peripheral blood (PB) of adult baboons following phlebotomy and decitabine treatment was compared by HPLC analysis of hemolysates. The baboon 58d CB contained >90% HbF and the ratio of Iγ/Vγ was 1.85. In the adult (phlebotomized) PB the level of HbF was 8.1% and the Iγ/Vγ ratio was 0.75 thus confirming that the ratio of the baboon Iγ and Vγ-globin chains differs in the fetal and adult stages of development in a manner similar to that of the human Gγ and Aγ-globin chains. Following decitabine treatment (PA 7002) an HbF level of 55% was attained with an Iγ/Vγ ratio of 1.1. Hemolysates prepared from d12 cultures of CD34+ baboon (PA 7002) BM cells grown in the presence of SCF, EPO, and Dex contained 57.6% HbF, nearly the same level observed following decitabine treatment in vivo. The Iγ/Vγ ratio was 1.94, markedly different from that observed in this same baboon following decitabine in vivo and, moreover, nearly identical to the fetal ratio. Thus HbF reactivation in cultured adult baboon CD34+ BM cells was associated with a change in the ratio of expression of the two baboon γ-globin genes to that characteristic of the fetal stage. Recapitulation of the fetal pattern of γ-globin chain expression in cultured baboon CD34+ progenitors demonstrates yet another advantage of the baboon model for investigations of hemoglobin switching.

Diagnostic Complications of Thalassemic Hb Showa-Yakushiji (β110[G12] Leu→Pro),Observed in An African American Child with Co-Inherited Hb B2 (δ16[A13]Gly→Arg) and α-Thal-2 (-α3.7 deletion): Effects of Triple Globin Gene Abnormality On Phenotype.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2558-2558
Author(s):  
Chinwe Obiaga ◽  
Niren Patel ◽  
Hernan Sabio ◽  
Natalia Dixon ◽  
Steffen E. Meiler ◽  
...  
Keyword(s):  
African American ◽  
Conflicts Of Interest ◽  
Globin Gene ◽  
Molecular Characteristics ◽  
African American Child ◽  
Globin Genes ◽  
Heterozygous State ◽  
Globin Chain ◽  
American Child ◽  
Globin Chains

Abstract Abstract 2558 Poster Board II-535 Hemoglobinopathies can usually be classified under two major categories. Qualitative abnormalities resulting from missense mutations in the globin genes, leading to the production of mostly asymptomatic Hb variants, and quantitative defects, which result in the synthesis of structurally normal globin chains in reduced quantities (thalassemias). However there are known globin chain variants that cause alterations of the globin structure as well as a decrease in synthesis, leading to a thalassemic phenotype. The occurrence of multiple abnormalities of α, β and δ globin chains can lead to an unusual and complex phenotype. We report here the inheritance of triple globin gene abnormalities in an African American child with a genotype that is heterozygous for three abnormalities: α-thal-2 (-α3.7 deletion), thalassemic Hb Showa-Yakushiji (β110[G12] Leu→Pro), and a δ-chain variant Hb B2 (δ16[A13]Gly→ Arg) . Although Hb Showa Yakushiji presents with a severe hemolytic anemia and a thalassemia-like phenotype in the heterozygous state; when co-inherited with Hb B2 and α-thal-2, a milder phenotype was observed. We report the diagnostic approach, molecular characteristics and genotype/phenotype correlations of this complex hemoglobinopathy syndrome. A 2 year old African American boy presented with anemia which was not responsive to iron therapy. CBC revealed: Hb 9.9 g/dL, Hct 31.3 %, MCV 62.5 fl, MCH 19.8 pg, MCHC 31.7 g/dl. The reticulocyte count was 1.1%. The iron profile showed a TIBC of 368; Iron 119; Transferrin 257, Ferritin 30; and % Iron saturation 32. The peripheral blood smear revealed a microcytic anemia suggestive of a thalassemic phenotype. The patient's hemolysate was analyzed by isoelectric focusing (IEF) showed Hb's A, F, A2, and a minor peak Hb X which was significantly slower than Hb A2 . Quantitative values by high performance liquid chromatography (HPLC) were: Hb F : 5.0%, Hb A: 91.0%, Hb A2: 2. 0% and Hb X (B2): 2.0%. Reverse Phase HPLC was also performed and no additional abnormality was detected. Sequencing of the β-globin genes revealed a heterozygous T→C mutation at the codon 110 consistent with Hb Showa-Yakushiji (β110[G12] Leu →Pro) which was not detectable with IEF and HPLC. Sequencing of the δ-globin genes showed a heterozygous G→C mutation at codon 16, Hb B2 (δ16[A13] Gly →Arg) which was also not detectible by IEF or HPLC unless over applied. A 590 bp long fragment of the β-globin gene (Accession # EU605697/APR-2008) and a 780 bp long fragment of the δ-globin gene (Accession # EU605698/APR-2008) sequences have been submitted to NCBI/GenBank. Detection of alpha thalasemia (α−3.7) deletion by PCR analysis, revealed one alpha gene deletion (−3.7α/αα). The leucine to proline substitution at residue 110 of β-globin chain, disrupts the G helix and the α1β1 contact of the hemoglobin molecule. As a result, an extremely unstable Hb variant will be produced, which leads to a thalassemic phenotype because of the reduced stability/viability of the mutant beta chain. Previously reported cases of Hb Showa-Yakushiji showed a more severe clinical picture in the heterozygous state than that observed in our patient. This is the first time Hb Showa-Yakushiji is identified in an African American child who presented with a moderate anemia and a thalassemia-like phenotype. The milder phenotype observed in our case may be due to the co-inheritance of α-thal-2 (α−3.7) deletion. The decreased production of α- globin chains may ameliorate the effect of the chain imbalances thus leading to milder clinical and hematologic manifestations. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Pathophysiology of β Thalassemia—A Guide to Molecular Therapies

Hematology ◽  
2005 ◽  
Vol 2005 (1) ◽  
pp. 31-37 ◽  
Author(s):  
Swee Lay Thein
Keyword(s):  
Globin Gene ◽  
Pathophysiological Mechanism ◽  
Globin Genes ◽  
Membrane Structures ◽  
Pharmacological Agents ◽  
Globin Chains ◽  
Molecular Therapies ◽  
Chain Imbalance ◽  
Chain Synthesis ◽  
Molecular Therapeutic

Abstract The central mechanism underlying the pathophysiology of the β thalassemias can be related to the deleterious effects of imbalanced globin chain synthesis on erythroid maturation and survival. An imbalance of the α/non-α globin chains leads to an excess of unmatched α globin which precipitates out, damaging membrane structures leading to accelerated apoptosis and premature destruction of the erythroid precursors in the bone marrow (ineffective erythropoiesis). Close observation of the genotype/phenotype relationships confirms the pathophysiological mechanism and provides clues to molecular therapies, all of which aim to reduce the α/non-α chain imbalance. They include inheritance of the milder forms of β thalassemia, co-inheritance of α thalassemia, or genetic factors (quantitative trait loci, QTLs) for increasing γ globin expression. Currently, the most promising molecular therapeutic approaches include increasing β globin gene expression by stem cell gene therapy and increasing γ globin expression using pharmacological agents or by transduction of the γ globin genes.

Haemoglobinopathies that occur with decreased HbA2 levels: a gene mutation set involving the δ gene at a Spanish centre

2016 ◽  
Vol 70 (1) ◽  
pp. 75-80 ◽  
Author(s):  
Ana Villegas ◽  
Fernando Ataúlfo González ◽  
Jorge M Nieto ◽  
Félix de la Fuente-Gonzalo ◽  
Rafael Martínez ◽  
...  
Keyword(s):  
High Performance ◽  
Globin Gene ◽  
Reversed Phase ◽  
Globin Genes ◽  
Reversed Phase Hplc ◽  
Structural Variants ◽  
Globin Chains ◽  
Zone Electrophoresis ◽  
Essential Parameter ◽  
Capillary Zone

AimsHaemoglobin A2 (HbA2) consists of two globin chains, α and β. Alterations in any of these genes influences the level of HbA2. Here, we present cases of structural Hb variants and thalassaemias which present either alone or together and reduce the level of HbA2 at varying degrees. Furthermore, we present a novel structural mutation in the δ globin gene, called Hb A2-Madrid.MethodsThe levels of HbA2 and HbF and the different haemoglobin variants were measured and analysed by ion exchange high performance liquid chromatography (HPLC, VARIANT II), the types of haemoglobins were determined by capillary zone electrophoresis (CZE) (Sebia) and the globin chains were determined by reversed-phase HPLC. Genetic analysis was performed by automatic sequencing of the α and δ genes as well as by multiple PCRs for the α globin genes.ResultsIn α thalassaemia (n=94), the HbA2 levels ranged from 1.39% to 2.43%. Among individuals with δ thalassaemia (n=5), the HbA2 level of those with δ+ thalassaemia was 1.77%, and that of those with δ0 thalassaemia was 1.70%. Among the individuals with δβ thalassaemia (n=13), those who were homozygous lacked HbA2. All structural haemoglobinopathies (n=97) were heterozygous; the α chain variants (n=84) presented with an HbA2 level of 1.76%, while the δ chain variants (n=13) presented with a level of 1.75%.ConclusionHbA2 is an essential parameter in the diagnostics of haemoglobinopathies. HPLC-EC and CZE allow the quantification of HbA2. Here, we show that quantification of HbA2 is critical for the identification of α, δ and βδ thalassaemias. Structural variants are discovered by HPLC. Molecular genetics is required for the proper identification of the mutations. Only with this knowledge is genetic counselling possible.

scholarly journals Correction of β-thalassemia by CRISPR/Cas9 editing of the α-globin locus in human hematopoietic stem cells

2021 ◽  
Vol 5 (5) ◽  
pp. 1137-1153 ◽  
Author(s):  
Giulia Pavani ◽  
Anna Fabiano ◽  
Marine Laurent ◽  
Fatima Amor ◽  
Erika Cantelli ◽  
...  
Keyword(s):  
Oxygen Carrier ◽  
Globin Gene ◽  
Globin Genes ◽  
Blood Disorders ◽  
Erythroid Progenitor ◽  
Gene Deletions ◽  
Hematopoietic Stem ◽  
Globin Chains ◽  
Novel Strategy ◽  
Novel Therapeutic Strategies

Abstract β-thalassemias (β-thal) are a group of blood disorders caused by mutations in the β-globin gene (HBB) cluster. β-globin associates with α-globin to form adult hemoglobin (HbA, α2β2), the main oxygen-carrier in erythrocytes. When β-globin chains are absent or limiting, free α-globins precipitate and damage cell membranes, causing hemolysis and ineffective erythropoiesis. Clinical data show that severity of β-thal correlates with the number of inherited α-globin genes (HBA1 and HBA2), with α-globin gene deletions having a beneficial effect for patients. Here, we describe a novel strategy to treat β-thal based on genome editing of the α-globin locus in human hematopoietic stem/progenitor cells (HSPCs). Using CRISPR/Cas9, we combined 2 therapeutic approaches: (1) α-globin downregulation, by deleting the HBA2 gene to recreate an α-thalassemia trait, and (2) β-globin expression, by targeted integration of a β-globin transgene downstream the HBA2 promoter. First, we optimized the CRISPR/Cas9 strategy and corrected the pathological phenotype in a cellular model of β-thalassemia (human erythroid progenitor cell [HUDEP-2] β0). Then, we edited healthy donor HSPCs and demonstrated that they maintained long-term repopulation capacity and multipotency in xenotransplanted mice. To assess the clinical potential of this approach, we next edited β-thal HSPCs and achieved correction of α/β globin imbalance in HSPC-derived erythroblasts. As a safer option for clinical translation, we performed editing in HSPCs using Cas9 nickase showing precise editing with no InDels. Overall, we described an innovative CRISPR/Cas9 approach to improve α/β globin imbalance in thalassemic HSPCs, paving the way for novel therapeutic strategies for β-thal.

Molecular Analysis of Alpha Hemoglobin Stabilizing Protein (AHSP) in Caucasian Patients with different Beta-Thalassemia Phenotypes.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3770-3770 ◽  
Author(s):  
M. Domenica Cappellini ◽  
Chiara Refaldi ◽  
Daniela Bignamini ◽  
Laura Zanaboni ◽  
Gemino Fiorelli
Keyword(s):  
High Performance ◽  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Clinical Severity ◽  
Beta Thalassemia ◽  
Red Cell ◽  
Globin Genes ◽  
Nucleotide Polymorphisms ◽  
Thai Population ◽  
Globin Chains

Abstract Beta-thalassemia is a inherited hemoglobin disorder characterized by absent or reduced synthesis of the b globin chains. The pathophysiology and the severity of b-thalassemias reflect the degree of globin chain imbalance and the excess of free a globin chains that precipitate and cause oxidative damage in red cell precursors inducing their premature destruction in the bone marrow (ineffective erythropoiesis). Although the phenotype of b thalassemias can be modified by inherited factors such as different number of a globin genes or increased fetal hemoglobin production, other mechanisms appear to be involved. Recently, a protein, named alpha hemoglobin stabilizing protein (AHSP), that acts as a molecular chaperone specifically for free a globin chains, preventing their precipitation in red cell precursors, has been identified. To establish whether AHSP might have a role in modifying the clinical outcome of b thalassemias, we have analyzed the AHSP gene in 70 Caucasian b thalassaemic subjects: 26 patients with b°/b° genotype (Thalassaemia Major),24 patients with Thalassemia Intermedia (b°/b+ or b+/b+) and 20 patients with a Thalassaemia Intermedia phenotype but with only one mutation in the b globin gene, a normal a globin genotype and no other causes of anemia. In all the subjects, we have performed Denaturing High-Performance Liquid Chromatography (DHPLC) of the three exons and the direct genomic sequencing of coding and noncoding regions (~ 1.5 kb) of AHSP gene. No mutations able to modify the structure or function of AHSP have been found, however we identified eight single nucleotide polymorphisms (SNPs) spanned along the whole gene that segregate in four different aplotypes. To evaluate a possible relationship between a particular aplotype and b thalassemia severity, the allele frequency of each single aplotype in the tree groups has been established and compared to that of 33 Caucasian normal controls: no statistically significant association has been proved. Even though the loss of AHSP aggravates the b thalassaemia phenotype in mice, in Thalassemic Caucasian population the AHSP apparently doesn’t make changes in the clinical severity of b thalassemia confirming the results recently found in Thai population.

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