Impact of the detection of ζ-globin chains and hemoglobin Bart's using immunochromatographic strip tests for α0-thalassemia (--SEA) differential diagnosis.

α0-Thalassemia is an inherited hematological disorder caused by the deletion of α-globin genes. The Southeast Asian deletion (--SEA) is the most common type of α0-thalassemia observed in Southeast Asian countries. Regarding WHO health policy, an effective α0-thalassemia screening strategy is needed to control new severe α-thalassemia cases. In this study, a monoclonal antibody panel was used to develop immunochromatographic (IC) strip tests for detecting the Hb Bart's and ζ-globin chain. Among 195 samples, all α0-thalassemia traits (78 α0-thalassemia (--SEA) and 4 α0-thalassemia (--THAI)) had low MCV or MCH values. The sensitivity, specificity, PPV and NPV of the IC strip tests for ζ-globin and Hb Bart's for screening α0-thalassemia (--SEA) within the low MCV or MCH samples were 100%, 65.2%, 90.7%, 100% and 96.2%, 47.8%, 86.6%, 78.6%, respectively. All 4 α0-thalassemia (--THAI) traits were negative for ζ-globin chains but positive for Hb Bart's using the IC strip tests. These results led to a α0-thalassemia screening being proposed in which blood samples are first evaluated by MCV, MCH and Hb typing. Samples with high MCV and MCH values are excluded for the presence of the α0-thalassemia gene. Samples with low MCV or MCH values are assayed using the developed IC strip tests, where only samples testing positive are further assayed for α0-thalassemia by PCR. Patients with Hb H, EA Bart's or EF Bart's diseases do not need to use this IC strip assay. Thus, in this study, a simple and cost effective α0-thalassemia point of care test was developed.

Facile spectroscopy and atomic force microscopy for the discrimination of α and ß thalassemia traits and diseases: A photodiagnosis approach.

Thalassemia (Thal) is an inherited blood disorder endemic to the Mediterranean and Middle East (e.g., KSA and UAE). This disease is caused by defects in the synthesis of one or more hemoglobin chains in red blood cells (RBCs). Alpha (α) Thal is caused by a reduced or absent alpha globin segment. Similarly, beta (ß) Thal is caused by a defect in the beta globin segment. We divided the diseases into four groups: α Thal trait, α Thal disease, ß Thal trait, and ß Thal disease. The α or ß Thal traits are milder variants of these diseases and do not require treatment; but ß Thal disease (and to a lesser extent, α Thal) causes hemolytic anemia, splenomegaly, and bone deformities and requires repeated lifelong blood transfusions. This paper presents results regarding the identification of Thal variants using fluorescence spectroscopy of blood biomolecules and atomic force microscopy analysis of the morphologic features of red blood cells. The combined results provide new insights into the characteristics of these diseases. Furthermore, this study shows why ß Thal disease subjects are often transfusion-dependent, and α Thal disease subjects are only occasionally transfusion dependent.

An assessment of methods used in the investigation of iron status: findings in a population of young British South Asian children.

AIMS: To assess the value of laboratory tests available for the investigation of iron status in a population of young British South Asian children. METHODS: Blood count, red cell distribution width (RDW), percentage hypochromic red cells (%hypo), concentrations of C-reactive protein (CRP), zinc protoporphyrin (ZPP), ferritin, soluble transferrin receptor, plasma iron measurements and incidence of deletional forms of α-thalassaemia were determined. RESULTS: Haemoglobin, mean cell haemoglobin (MCH), ferritin and CRP values classified iron status in 151/205 (73.6%) consecutive children aged 4-43 months. Fifty-four could not be classified: 12 were anaemic with findings, other than normal CRP values, indistinguishable from those with anaemia of inflammation and 42 were non-anaemic with reduced MCH values. All 42 had normal ferritin concentration and 8 of 36 successfully tested had deletional α-thalassaemia trait. Despite apparent iron sufficiency the RDW, %hypo and ZPP values of these 42 were not significantly different from the 32 children classified with iron-deficient erythropoiesis. The gene frequency of deletional α-thalassaemia trait in the entire group was 8.6%. CONCLUSIONS: Among 205 British South Asian children aged 4-43 months with high incidences of anaemia, iron deficiency, infection and α-thalassaemia, 151 (73.6%) were classified using haemoglobin, MCH, ferritin and CRP values. In 42 non-anaemic, iron-sufficient children with subnormal MCH values, that is with a phenotype of α-thalassaemia trait, RDW, %hypo and ZPP values did not differ significantly from those with iron-deficient erythropoiesis. Raised RDW, %hypo and ZPP values should be interpreted with caution in non-anaemic young British South Asian children with microcytosis.

[Relationship between HLA-A, B alleles and red blood cell parameters of patients with --(SEA/αα) subtype of α(0)-thalassemia of Han ethnic population of Wuzhou city].

This study was purposed to investigate the relationship between HLA-A, B allele polymorphisms and red blood cell parameters of patients with --(SEA/αα) subtype of α(0)-thalassemia in Han ethnic population of Wuzhou city. The HLA genetic polymorphisms were determined by polymerase chain reaction-sequence-based typing (PCR-SBT) in 57 patients with --(SEA/αα) subtype of α(0)-thalassemia of Han ethnic population in Wuzhou city, Guangxi province, China. Mean corpuscular volume (MCV), hemoglobin (Hb), mean corpuscular hemoglobin (MCH) and mean corpuscular hemoglobin concentration (MCHC) were detected by automatic blood analyzer system. HbA2 were detected by electrophoretic method. The statistical analysis was performed by ordinal polytomous logistic regression. The results showed that Hb and HbA2 levels were significantly lower in patients positive for HLA-A*33:03, B*15:01 or B*55:02, and were significantly higher in patients positive for B*15:02 (P < 0.05). It is concluded that several HLA alleles may be associated with Hb level of --(SEA/αα) subtype of α(0)-thalassemia of Han ethnic population in Wuzhou city. This result has the value for understanding phenotype-genotype relationships in thalassemia.

Hemoglobin Q-Thailand related disorders: origin, molecular, hematological and diagnostic aspects.

We describe the molecular and hematological profiles of thalassemia syndromes caused by interactions of hemoglobin (Hb) Q-Thailand [α74(EF3) Asp-His] and various hemoglobinopathies found in 52 unrelated adult Thai subjects. Ten genotypes including several previously undescribed conditions were observed, which were classified into 4 groups. Group I included 26 Hb Q-Thailand heterozygotes and a homozygotous subject. Group II included subjects with Hb Q-Thailand and other α-thalassemia alleles in trans including 1 compound Hb Q-Thailand/α(+)-thalassemia (-α(3.7)), 2 Hb Q-Thailand/Hb Constant Spring disease and 6 Hb H/Q-Thailand disease. The average levels of Hb Q-Thailand were found to be 29.8%, 82.3%, 34.7%, 49.2-49.3% and 79.4%, respectively. Both Hbs Bart's and H were observed in addition to Hb Q-Thailand in all 6 cases with Hb Q-H disease but not in a homozygous Hb Q-Thailand. Group III included 7 double heterozygotes for Hb Q-Thailand/Hb E, 3 Hb Q-Thailand/Hb E/α(+)-thalassemia (-α(3.7)), 3 heterozygous Hb Q-Thailand/homozygous Hb E and 1 triple heterozygote for Hb Q-Thailand/Hb Constant Spring/Hb E. In this group, Hbs E (α(A)(2)ß(E)(2)), Q-Thailand (α(QT)(2)ß(A)(2)) and QE (α(QT)(2)ß(E)(2)) were observed on both HPLC and capillary electrophoresis. The Hb QE, rather than Hb Q-Thailand, was detected in all 3 cases with heterozygous Hb Q-Thailand and homozygous Hb E. The remaining two cases in group 4 were double heterozygotes for Hb Q-Thailand and ß(0)-thalassemia in which Hb Q-Thailand, elevated Hb A(2) (α(A)(2)δ(2)), and Hb QA(2) (α(QT)(2)δ(2)) were detected. DNA analysis identified the Hb Q-Thailand mutation (α74: GAC-CAC) and the linked (-α(4.2)) in all cases. Analysis of α-globin gene haplotype provided the first evidence of a single origin of this Hb variant in Thai population.

Variable clinical phenotypes of alpha-thalassemia syndromes.

Genetic mutations of the alpha genes are common worldwide. In Asia and particularly Southeast Asia, they can result in clinically significant types of alpha-thalassemia, namely hemoglobin (Hb) H disease and Hb Bart's hydrops fetalis. The latter is generally a fatal intrauterine condition, while Hb H disease results in clinical complications that are frequently overlooked. The high prevalence of the carrier state and the burden of these diseases (and other alpha-thalassemia variants) call for more attention for improved screening methods and better care.

Genetic etiologies for phenotypic diversity in sickle cell anemia.

The clinical course of patients with sickle cell anemia, a Mendelian trait, is characteristically highly variable. HbF concentration and the presence of á thalassemia are established modulators of the disease, but cannot account for all of its clinical heterogeneity. To find additional genetic modulators of disease, genotype-phenotype association studies, where single nucleotide polymorphisms (SNPs) in candidate genes are linked with a particular phenotype, have been informative. SNPs in several genes of the TGF-beta/BMP superfamily, and some other genes linked to the endothelial function, and nitric oxide biology are associated with the subphenotypes of stroke, osteonecrosis, priapism, leg ulcers, pulmonary hypertension, and a more general measure of overall disease severity. Genome-wide association studies should help to confirm these observations and also to find hitherto unsuspected genetic modulators. Genetic association studies can have immediate prognostic value; they might also help to identify new pathophysiological pathways that could be susceptible to modulation.

Rapid diagnosis of alpha(o)-thalassemia using the relative quantitative PCR and the dissociation curve analysis.

A method for diagnosis of alpha(o)-thalassemia was developed based on detection of accumulated PCR product using SYBR Green I, a double-stranded DNA binding dye, and a fluorescence-detecting thermocycler. Primers were designed to specifically amplify - -SEA and - -Thai deletions of alpha(o)-thalassemia. Albumin was selected as the reference gene. The comparative CT method was used to quantitate the target gene relative to the albumin gene. Dissociation curve analysis was used as a qualitative tool to distinguish different types of alpha(o)-thalassemia. In this study, the melting temperatures of the - -Thai and - -SEA deletions were 83 and 86 degrees C, respectively. Application of the assay for the diagnosis of alpha(o)-thalassemia heterozygotes and homozygotes is reported. The assay is highly robust and amenable to high throughput. It is thus a useful tool for the rapid detection of alpha(o)-thalassemia in populations with a high frequency of alpha(o)-thalassemia, - -SEA and - -Thai deletions.

Genetic and clinical features of hemoglobin H disease in Chinese patients.

BACKGROUND: Normally, one pair of each of the two alpha-globin genes, alpha1 and alpha2, resides on each copy of chromosome 16. In hemoglobin H disease, three of these four alpha-globin genes are affected by a deletion, a mutation, or both. We studied the alpha1-globin gene abnormalities and the clinical and hematologic features of Chinese patients with hemoglobin H disease in Hong Kong. METHODS: We assessed the clinical features, hematologic values, serum ferritin levels, and liver function of 114 patients with hemoglobin H disease. We also performed echocardiography and magnetic resonance imaging of the liver and examined the two pairs of alpha-globin genes. RESULTS: Hemoglobin H disease in 87 of the 114 patients (76 percent) was due to the deletion of three of the four alpha-globin genes (--/-alpha), a combination termed the deletional type of hemoglobin H. The remaining 27 patients (24 percent) had the nondeletional type of hemoglobin H disease, in which two alpha-globin genes are deleted and a third is mutated (--/alphaalphaT). All 87 patients with the deletional type of hemoglobin H were double heterozygotes in whom there was a deletion of both alpha-globin genes from one chromosome, plus a deletion of the alpha1 or alpha2 gene from the other chromosome (--/alpha- or --/-alpha). A variety of mutated alpha-globin genes was found in the patients with nondeletional type of hemoglobin H disease. Patients with the nondeletional type of the H disease had more symptoms at a younger age, more severe hemolytic anemia, and larger spleens and were more likely to require transfusions than patients with deletional hemoglobin H disease. The severity of iron overload was not related to the genotype. CONCLUSIONS: Chinese patients in Hong Kong with the nondeletional type of hemoglobin H disease have more severe disease than those with the deletional type of the disease. Iron overload is a major cause of disability in both forms of the disease.

Red blood cell phenotypes in alpha-thalassemias in the Spanish population.

BACKGROUND AND OBJECTIVE: alpha-thalassemia is very common on all thalassemic geographical regions. The present work aimed at analyzing the relationship between the degree of microcytosis and hematological parameters and the type of alpha-thalassemic mutation. DESIGN AND METHODS: Five hundred and thirty-six subjects with 4 kinds of alpha-thalassemia were examined using established techniques that determined all hematological parameters, and globin synthesis and molecular biological techniques to study the DNA of globin genes by Southern blotting. RESULTS: Adult carriers of alpha (+)-thalassemia (-alpha/alpha alpha) present very few hematological alterations. In a statistical comparison with normal individuals (alpha alpha/alpha alpha), significant differences were found between the hemocytometric data and the MCV and MCH of heterozygous alpha + thalassemia and the heterozygous alpha zero or homozygous alpha + genotype. Hb H disease was detected in 15 patients, presenting a severe degree of anemia, a significant increase in RDW and globin chain synthesis with an alpha/beta ratio of 0.5 +/- 0.1. INTERPRETATION AND CONCLUSIONS: These data provide reference values for geographical areas where alpha + thalassemia is common. These hematocytometric data, together with hemoglobin analysis, could be useful as a future reference data for new patients diagnosed with alpha-thalassemia.

[A rapid method for diagnosing alpha-thalassemia-1 of Southeast Asia type].

OBJECTIVE: To explore a rapid method for diagnosing alpha-thalassemia-1 of Southeast Asia type. METHODS: Seventy-six patients were detected by polymerase chain reaction (PCR). RESULTS: Thirty-one patients were diagnosed as alpha-thalassemia-1 of Southeast Asia type. PCR was compared with polyacrylamide gel electropheresis for detecting zeta-globin chain in 56 patients and the correspondance rate was 83.93%. Ten cases of prenatal diagnosis were performed by PCR, and the results showed that one was Hb bart's hydrop fetalis, four were carriers of alpha-thalassemia-1 of Southeast Asia type, and the others were normal fetalis or carrier of alpha-thalassemia-2. CONCLUSION: The PCR method is simple and accurate for diagnosing alpha-thalassemia of Southeast Asia type, and it provides a new approach to the prenatal diagnosis of alpha-thalassemia fetalis and the detection of alpha-globin gene cluster.

Alpha thalassemia and its impact on other clinical conditions.

Mild alpha thalassemia is the most prevalent genetic trait worldwide. We have recently developed a multiplex polymerase chain reaction method to screen for the most common deletions which give rise to alpha thalassemia. The authors have used this method to investigate a potential association of alpha thalassemia with hypertension. Our results show that the prevalence of hypertension in hospitalized blacks who have the -alpha 3.7 deletion is 71 percent higher than the prevalence in hospitalized blacks who do not have the deletion. The authors present a potential mechanism to explain this association.

Frequency of alpha-thalassemia-1 of the Southeast Asian-type among pregnant women in northern Thailand determined by PCR technique.

Five hundred pregnant women were analyzed for the presence of alpha-thalassemia-1 of the Southeast Asian (SEA)-type by polymerase chain reaction (PCR) technique at the Maharaj Nakhon Chiang Mai University Hospital in Chiang Mai during the period from April to June 1995. Forty-four of them (8.8%) were recognized as carriers, corresponding to a frequency of 0.044. Homozygous alpha-thalassemia-1 of the SEA-type, the fatal condition of hemoglobin Bart's hydrops fetalis, has an expected frequency of 0.00194, or about 2 hydrops fetalis cases per 1,000 births in this population.

The alpha / beta and alpha 2 / alpha 1-globin mRNA ratios in different forms of alpha-thalassemia.

The present study provides information about the alpha / beta and alpha 2 / alpha 1-mRNA ratios in reticulocytes of normal adults and individuals with different alpha-globin gene deficiencies; it found its origin in analytical data of blood samples from a Laotian couple and their newborn baby. The father carried the 4.2 kb deletion on one chromosome and a TAA --> CAA mutation at the terminating codon of the alpha 2 gene (Hb Constant Spring or CS) on the other chromosome. The mother had the 3.7 kb deletion on one chromosome and a TA A --> TAT mutation at the terminating codon of the alpha 2-globin gene (Hb Paksé) of the second chromosome. The baby was a compound heterozygote for the two termination codon mutations. The mRNA data for this family were compared to those for persons with several well-defined alpha-globin gene deficiencies. The results confirm the importance of the alpha 2 alpha 1-mRNA for the synthesis of alpha chains in alpha-thalassemia-2 homozygotes (-alpha/-alpha) and in patients with Hb H disease due to the deletion of three alpha-globin genes (-alpha/--). Furthermore, the MRNA production of the alpha 1-globin gene on the chromosome with the alpha CS mutation (alpha CS alpha) is only one-half of that by the alpha 2 alpha 1-globin gene of a chromosome with a 3.7 or 4.2 kb deletion, explaining the greater severity of, and higher Hb H level in Hb H patients with the alpha CS alpha condition (alpha CS alpha/--) as compared to those with the three gene deletion (-alpha/--). The methodology could be useful as a preliminary screening for the presence of point mutations leading to the functional loss of a single alpha-globin gene, provided common deletional alleles have been excluded.

Thalassaemia and other haemoglobinopathies in general practice.

Haemoglobinopathies are now common in Australia following the migration of people from areas of high prevalence. This article gives practical advice on the management of patients incidentally found to be heterozygotes for the more common forms and also outlines complications of major haemoglobinopathies that are seen in family practice.

[Alpha-thalassemia]. / Les alpha-thalassémies.

alpha-Thalassaemias are probably the most common genetic disorder worldwide. alpha-Thalassaemias are haemolytic anaemias resulting from inherited deficient synthesis of alpha-globin chains. In this paper, the classification and nomenclature of alpha-thalassaemias are developed. Procedures to ensure the laboratory diagnosis are explained. The heterozygous carrier states for these disorders are, in most cases, not associated with any easily discernible change in the haemoglobin pattern, except, sometimes, a reduced MCV in the blood picture. Heterozygotes for alpha-thalassaemia deletions are now detectable by the accurate PCR method. Because of the high prevalence of these disorders in large segments of the world population, alpha-thalassemia and haemoglobinopathies often occur in the same individual. The laboratory features of these interactions and, particularly, the role of alpha-thalassaemia as a potential modulator of sickling haemoglobinopathies are discussed.

Thalassemia: pathophysiology of red cell changes.

The thalassemias are extremely heterogeneous in terms of their clinical severity, and their underlying pathophysiology relates directly to the extent of accumulation of excess unmatched globin chains: alpha in beta thalassemia and beta in the alpha thalassemias. However, the accumulation of each separate globin chain affects red cell membrane material properties and the state of red cell hydration very differently. These observations presumably account for the varying extent of ineffective erythropoiesis and peripheral blood hemolysis in the major variants of thalassemia. The thalassemias are a worldwide group of inherited disorders of globin-chain synthesis that developed in multiple geographic regions, probably because they provided partial protection against malaria. In normal assembly of adult hemoglobin (HbA-alpha 2 beta 2), alpha and beta globin are synthesized by genes on different chromosomes, whereas heme is synthesized primarily on mitochondria. The synthesis of these chains is very tightly coordinated so that the ratio of alpha globin to beta globin (beta in this case including the beta-like globins delta and gamma) is normally 1 +/- 0.05. Furthermore, specific erythroid proteases are designed to attack and destroy excess alpha or beta globin chains, demonstrating the deleterious impact of the accumulation of excess unmatched globin chains. In beta thalassemia, production of beta globin decreases and excess alpha globin accumulates. In alpha thalassemia, on the other hand, this process occurs in reverse. Perhaps in these disorders more than any others, molecular biologists have documented the deletional and transcriptional events leading to diminished synthesis of specific classes of globin chains.(ABSTRACT TRUNCATED AT 250 WORDS)

[Diagnosis of hemoglobinopathies and thalassemias]. / Diagnostikk av hemoglobinopatier og talassemier.

During recents the number of immigrants to Norway from Africa and South East Asia has risen considerably. These persons come from countries with a high prevalence of haemoglobinopathies. Most of the immigrants with haemoglobinopathies are silent carriers, but some have a serious disease or have offspring with serious disease. This situation calls for increasing awareness and knowledge of haemoglobinopathies, especially the thalassemias and sickle cell trait. Genetic counselling is particularly important, since the majority of these immigrants marry within small ethnic groups. This paper gives a survey of the most frequent haemoglobinopathies likely to be encountered in Norway.