[Show abstract][Hide abstract] ABSTRACT: Thalassemia and abnormal hemoglobin are the most common genetic disorders and are considered health problems in many developing countries. In the last few years, there has been much progress in laboratory diagnosis, treatment and control of thalassemia. The variation in the clinical severity in both α- and β-thalassemia reflects a genotype-phenotype interaction. This is important for future therapeutic intervention and should be well characterized in each population. The quality of life of the patients is much improved with regular blood transfusion and novel iron chelators. The cure for thalassemia is possible by stem cell transplantation and future gene therapy. It is expected that under multinational collaboration the prevention of thalassemia will happen worldwide.
Preview · Article · Jul 2012 · International journal of laboratory hematology
[Show abstract][Hide abstract] ABSTRACT: Non-transferrin bound iron (NTBI) is found in plasma of β-thalassemia patients and causes oxidative tissue damage. Cardiac siderosis and complications are the secondary cause of death in β-thalassemia major patients. Desferrioxamine (DFO), deferiprone (DFP) and deferasirox (DFX) are promising chelators used to get negative iron balance and improve life quality. DFP has been shown to remove myocardial iron effectively. Curcuminoids (CUR) can chelate plasma NTBI, inhibit lipid peroxidation and alleviate cardiac autonomic imbalance. Effects of CUR on cardiac iron deposition and function were investigated in iron-loaded mice. Wild type ((mu)β(+/+) WT) and heterozygous β-knockout ((mu)β(th-3/+) BKO) mice (C57BL/6) were fed with ferrocene-supplemented diet (Fe diet) and coincidently intervened with CUR and DFP for 2 months. Concentrations of plasma NTBI and malondialdehyde (MDA) were measured using HPLC techniques. Heart iron concentration was determined based on atomic absorption spectrophotometry and Perl's staining methods. Short-term electrocardiogram (ECG) was recorded with AD Instruments Power Lab, and heart rate variability (HRV) was evaluated using MATLAB 7.0 program. Fe diet increased levels of NTBI and MDA in plasma, nonheme iron and iron deposit in heart tissue significantly, and depressed the HRV, which the levels were higher in the BKO mice than the WT mice. CUR and DFP treatments lowered plasma NTBI as well as MDA concentrations (p <0.05), heart iron accumulation effectively, and also improved the HRV in the treated mice. The results imply that CUR would be effective in decreasing plasma NTBI and myocardial iron, alleviating lipid peroxidation and improving cardiac function in iron-loaded thalassemic mice.
No preview · Article · Jan 2011 · Medicinal chemistry (Shāriqah (United Arab Emirates))
[Show abstract][Hide abstract] ABSTRACT: DNA from members of 2 Thai families with conditions considered to be δβ-thalassaemia were studied by using restriction endonuclease DNA mapping. The propositus in family A is a double heterozygote for β-thalassaemia and δβ-thalassaemia. DNA analysis reveals a deletion of the β-globin gene cluster starting at the area between the Sac I and Eco RI sites near the 3′ end of the Gγ-gene and extending through the Aγ-, δ- and β-genes to an unknown extent downstream. In family B, the propositus is δβ-thalassaemia/Hb E. Deletion of the β-globin gene cluster begins in the large intervening sequence of the Aγ-gene and removes both δ- and β-genes downstream.
No preview · Article · Aug 2009 · European Journal Of Haematology
[Show abstract][Hide abstract] ABSTRACT: β-Thalassemia and Hb E patients, with seemingly identical genotypes, have a remarkable variability in severity. Reduction in red cell survival in β-thalas-semia is correlated with the amount of intracellular unmatched α-globin chains. However, it was only recently realized that mRNA, whose translation is prematurely terminated, is also unstable. No systematic attempts have been made to investigate mRNA stability in β-thalassemia arising from nonsense mutations located upstream from the normal termination codon. In this study, one-step real-time polymerase chain reaction has been employed to compare the levels of α- and β-globin mRNA in reticulocytes from β-thalassemia/Hb E subjects. The results showed the highest α/β-globin mRNA ratio (median = 5.70, n = 13) in frameshift codons 41/42 (-TTCT)/Hb E individuals compared to normal subjects (median = 1.02, n = 6), or those with Hb E trait (median = 2.15, n = 8). In addition, there was a concomitant increase in the α/β-globin mRNA ratio with decrease in hemoglobin level, ie., increase in severity. The difference in the ratio among β-thalassemia/Hb E patients with the same genotype may be attributed to individual variations of efficiency in βE-globin mRNA splicing and in the destruction of prematurely terminated mRNA.
[Show abstract][Hide abstract] ABSTRACT: The propositus was a 29-year-old Thai male, whose electrophoretic pattern showed Hb A (58%) plus an abnormal hemoglobin (42%) with mobility identical to Hb A2 and Hb E. Protein sequencer analysis and tryptic peptide mapping of the ß chain indicated that the abnormal hemoglobin was Hb C [ß6(A3)Glu→Lys], rather than Hb E which is more commonly found in South East Asia. This conclusion was confirmed by direct sequence analysis of the propositus' DNA, which showed AAG as well as GAG at codon 6 of the ß gene, in agreement with heterozygosity for Hb C and Hb A. Furthermore, the ß gene framework (Ava II -, Bam HI +) of the propositus suggested that the ßc gene may have arisen from an independent mutation. Since Hb C and Hb E have the same mutation (Glu→Lys) in the ß chain, although at different positions, and behave similarly in electrophoresis, cases of Hb C and Hb E may sometimes have been mistakenly identified for each other, based on whichever variant is most prevalent in the particular population.
[Show abstract][Hide abstract] ABSTRACT: Clinical manifestations and hematologic data of thalassemia intermedia were observed in three siblings of a Thai family. Analyses of the hemoglobin of their parents and other siblings indicated that they inherited a δβ-thalassemia gene from the father and a β-thalassemia gene from the mother. Globin gene mapping confirmed that they carry two abnormal β-globin gene complexes. On one chromosome more than 70 kb of DNA was removed which resulted in Gγ(Aγδβ)-thalassemia. The deletion started at the Hind III site located just 3' to the Gγ gene, and extended downstream to a region recognized by the p3'N 2.8R probe which is located more than 45 kb from the 3' end of the B gene. The other chromosome carried a β-thalassemia gene, and a 5 kb deletion between the Gγand Aγ genes which produced a hybrid -GAγ_ gene. A synthetic oligonucleotide probe showed that this β°-thalassemia arose from a CT mutation at position 654 of IVS-II in the β-globin gene.
[Show abstract][Hide abstract] ABSTRACT: In Thailand α-thalassemia (thal), β-thal, hemoglobin (Hb) E and Hb Constant Spring (Hb CS) are prevalent. The incidences are 20–30% for α-thal (3.5% for α-thal-1 and 16% for α-thal-2), 3–9% for β-thal, up to 54% for Hb E and nearly 8% for Hb CS (1). Different combinations of these genes result in a spectrum of thalassemia syndromes ranging in severity from asymptomatic to intrauterine death. From the known gene frequencies the numbers of thalassemic patients per generation (total population of 50 million) are as follows: Hb Bart's hydrops fetalis 20,000; homozygous β-thal 31,250; β-thallHb E disease 162,500; Hb H disease (two genotypes) 200,000, making a total of 413,750. In addition, individuals may carry more than two of the abnormal genes leading to complex thalassemia syndromes such as αβ-thal, AE-Bart's and EF-Bart's diseases.
[Show abstract][Hide abstract] ABSTRACT: EF Bart's disease is an uncommon form of thalassaemia intermedia resulting from the co-inheritance of α-thalassaemia and haemoglobin E in the same subject. Starch-gel electrophoresis revealed two phenotypes in 19 patients with EF Bart's. 16 patients had Hbs CS + E + F + Bart's and the remainder had Hbs E + F + Bart's. DNA mapping and haemoglobin electrophoresis indicated that there are four genotypes, involving 5 abnormal globin genes, responsible for this thalassaemia syndrome.
No preview · Article · Apr 2009 · European Journal Of Haematology
[Show abstract][Hide abstract] ABSTRACT: We evaluated the contribution of 67 single nucleotide polymorphisms (SNPs) within the beta-globin gene cluster to disease severity in groups of 207 mild- and 305 severe unrelated patients from Thailand with Hemoglobin E (HbE)/beta(0)-thalassemia and normal alpha-globin genes. Our analysis showed that these SNPs comprise two distinct linkage disequilibrium blocks, one containing the beta-globin gene and the other extending from the locus control region (LCR) to the delta gene, which are separated by a recombination hotspot in the narrow region of the beta-globin gene promoter. Forty-five SNPs within the interval including the LCR region and the delta gene showed strong association with disease severity. The strongest association was observed with the XmnI polymorphism located 158-bp upstream to the G gamma gene (p = 4.6E-12). Carriers of the T allele of XmnI were more likely to have a milder disease course and higher level of fetal hemoglobin (HbF) in both the mild (p = 0.005) and severe (p = 8.7E-06) patient groups. Haplotype analysis revealed that the T allele of XmnI was nearly always in cis with the HbE allele. The high frequency of this haplotype may be favored by positive selection against malarial infection. Further studies are needed to validate this hypothesis and determine whether XmnI or another closely linked variant modulates severity and HbF levels in patients with beta(0)-thalassemia/HbE disease.
Full-text · Article · Jan 2008 · Clinical Genetics
[Show abstract][Hide abstract] ABSTRACT: This study described the expression of beta(E)-globin in newborns heterozygous for HbE. Despite the lower level of HbE, the pattern of beta(E)-globin gene expression was similar to beta(A)-globinn because the increase in HbE and HbA reached the peak level at the same time. A delayed decline of HbF was observed.
[Show abstract][Hide abstract] ABSTRACT: Erythropoietin (EPO) and interferon-gamma (IFN-gamma) added to human erythroid progenitor cells purified from peripheral blood (erythroid colony-forming cells; ECFC) significantly reduces apoptosis as assessed by flow cytometry (FCM) using annexin V. To clarify the role of NF-kappaB in the regulation of the apoptosis of erythroid progenitor cells, cyclosporin A (CsA), which blocks dissociation of the NF-kappaB complex, was added to serum-free cultures of ECFC. CsA induced the apoptosis of ECFCs in the presence of EPO or IFN-gamma, but at different magnitudes. In the presence of a relatively low concentration of CsA (10 microm), apoptosis was induced only in cultures with EPO. The direct involvement of NF-kappaB was then assessed by Western blotting and confocal microscopy. In the presence of EPO, NF-kappaB was abundant both in the cytoplasm and in the nucleus, and nuclear expression was diminished after adding CsA. In contrast, NF-kappaB was undetectable in the nucleus in the presence of IFN-gamma. The effect of CsA on mitochondrial function was investigated by determining the DeltaPsim and reactive oxygen species production. CsA disturbed the transmembrane potential in the presence of either EPO or IFN-gamma, although the viability of the cells was maintained in the presence of IFN-gamma plus CsA. These results indicate that IFN-gamma reduced the apoptosis of erythroid progenitor cells through a unique signaling pathway that is independent of NF-kappaB translocation, and which is not mediated by modulating mitochondrial function, whereas EPO reduced apoptosis through NF-kappaB translocation to the nucleus.
No preview · Article · May 2005 · European Journal Of Haematology
[Show abstract][Hide abstract] ABSTRACT: Thalassemia and abnormal hemoglobins are common genetic disorders in Southeast Asia. Thalassemia is not only an important public health problem but also a socio-economic problem of many countries in the region. The approach to deal with the thalassemic problem is to prevent and control births of the new cases. This requires an accurate identification of couple at high risk to have a thalassemic child. The diagnosis of thalassemia carriers need several tests that are not practical for screening the population at large. In this study we used two simple laboratory tests to screen for potential thalassemia carriers and hemoglobin E individuals. Three-hundred pregnant women and 40 spouses were recruited in this study. The methods were the red cell osmotic fragility (OF) screening test with 0.36% NaCl and the dichlorophenolindophenol (DCIP) precipitation test to detect Hb E and unstable hemoglobins. Standard methods for red cell indices, hemoglobin analysis and detection of alpha-thalassemia by immunological method were also performed to confirm genotypes of thalassemia. The results showed that 98 women (32.7%) were carriers of thalassemias and hemoglobin E. The number of false positive by OF test was 3.2% and by DCIP test was 0.6%. Sensitivity and specificity of OF test were 89.5% and 93.3%, respectively whereas those of DCIP test were 100%. Of the 40 couples investigated, one was found to be at risk of having beta-thalassemia/Hb E fetus. Screening techniques including one tube osmotic fragility and DCIP precipitation tests are sensitive and specific for the detection of thalassemia and unstable hemoglobins such as Hb E. The techniques are also simple, economic, rapid, and give minimal false negative result.
No preview · Article · Feb 2002 · The Southeast Asian journal of tropical medicine and public health
[Show abstract][Hide abstract] ABSTRACT: Thalassemia syndromes and abnormal hemoglobins (Hbs), a group of autosomal recessive inherited disorders, are prevalent in Thailand. Measures for prevention and control of thalassemia include screening for the carriers, genetic counseling, and prenatal diagnosis (PND) in high-risk couples. PND may be carried out by analysis of either fetal blood or fetal tissue samples such as chorionic villi or amniotic fluid fibroblasts. Hb analysis of fetal blood by automated high-performance liquid chromatography (HPLC) has been recently applied for PND of many thalassemia syndromes. In this study, we used automated HPLC (VARIANT-Beta-Thalassemia Short Program; Bio-Rad, Hercules, CA) to carry out PND using fetal blood samples obtained by cordocentesis from fetuses at risk for severe thalassemic diseases including homozygous β-thalassemia, β-thalassemia/Hb E, and Hb Bart's hydrops fetalis. Hb types determined using HPLC were compared with those obtained using DNA analyses. The Hb molecules of fetuses at risk for β-thalassemic diseases in Thailand were divided into 4 groups: Hb F, Hb FA, Hb A2(E) F, and Hb A2(E) FA. Homozygous β°-thalassemia and β°-thalassemia/Hb E have Hb F and Hb E and Hb F, respectively. These 2 disorders can be easily diagnosed by HPLC because they are characterized by the absence of Hb A in the fetal cord blood. However, cases with β+-thalassemia produce small amounts of Hb A (0.5%-1%) and can be misdiagnosed as thalassemia carriers. Moreover, the HPLC system may not differentiate β°-thalassemia/Hb E from homozygous Hb E because both conditions have Hb type A2(E)+F without Hb A, and the amount of Hb A2(E) may overlap. Cases in which the presence of Hb A is detected in cord blood (Hb type FA) at 16 to 24 weeks gestation can be normal fetuses, β-thalassemia heterozygotes, or have β+-thalassemia syndrome. The pattern of the chromatogram obtained by this automated HPLC is also very specific for Hb Bart's hydrops fetalis, which makes the diagnosis simple, reliable, and inexpensive.
[Show abstract][Hide abstract] ABSTRACT: The spectrum of the beta-thalassemia mutations of Thailand, Pakistan, India, Sri Lanka, Mauritius and Syria has been further characterized by a multi-center study of 1,235 transfusion-dependent patients, and the mutations discovered used to assess the fidelity of a simple diagnostic strategy. A total of 44 beta-thalassemia mutations were identified either by allele-specific oligonucleotide hybridization, amplification with allele-specific primers, or DNA sequencing of amplified product. The results confirm and extend earlier findings for Thailand, Pakistan, India, Mauritius and Syria. This is the first detailed report of the spectrum of mutations for Sri Lanka. Two novel mutations were identified, codon 55 (-A) and IVS-I-129 (A-->C), both found in Sri Lankan patients. Two beta-thalassemia mutations were found to coexist in one beta-globin gene: Sri Lankan patients homozygous for the beta0 codon 16 (-C) frameshift were also homozygous for the beta+ codon 10 (C-->A) mutation. Studies of Sri Lankan, Pakistani, and Indian carriers suggest the codon 10 (C-->A) mutation is just a rare polymorphism on an ancestral allele, on which the beta0 codon 16 (-C) mutation has arisen. Each country was found to have only a few common mutations accounting for 70% or more of the beta-thalassemia alleles. A panel of primers to diagnose the majority of the mutations by the amplification refractory mutation system was developed, enabling a simple molecular diagnostic strategy to be introduced for each country participating in the multi-center study.