Expression of Hb β-T and Hb β-E genes in Eastern India—Family studies

The distribution patterns of different haemoglobins were observed amongst the family members of β-thalassaemia homozygous and HbE-β-thalassaemia patients with the aid of gel electrophoretic and alkali denaturation techniques. Of the 18 families studied, four belonged to β-thalassaemia homozygous and 14 to HbE-β-thalassaemia patients. Interaction of HbE and β-thalassaemia genes resulted in major clinical abnormalities with increase in the percentages of haemoglobins F and E. The percentages of HbA₂ in homozygous β-thalassaemia were within the normal range. Although in Southeast Asia the β° type of HbE-thalassaemia is more prevalent, only one individual with this type of thalassaemia was observed during this survey. In the rest of the patients examined the percentages of adult haemoglobin ranged from 5.2 to 42.5 indicating the presence of a β⁺ type gene.     

Twelve families with Hb 0 Arab in the Burgas district of Bulgaria observations on sixteen examples of Hb 0 β° thalassaemia

Summary 12 families with Hb 0 Arab from the Burgas district of Bulgaria were examined. Of the 262 members of the 12 families, 44 persons were simple Hb 0 Arab heterozygotes, 18 had -thalassaemia, and 16 were double heterozygotes for Hb 0 Arab and ^° thalassaemia.     

Organization of α-chain genes among Hb G-Philadelphia heterozygotes in association with Hb S, β-thalassemia,and α-thalassemia-2

The percentages of the -chain variant Hb G-Philadelphia (Hb G) or ₂ 68 AsnLys₂ were evaluated in 84 adult and 18 newborn heterozygotes. These included members of three families who were studied in more detail by nucleic acid hybridization techniques. The adult heterozygotes fell in two categories, one with a higher proportion of Hb G [46.5±1.0% (SD), N=21] and another with lower values (33.9±3.4%, N=63). Among the newborn heterozygotes, two babies fell in the category with the higher proportion of Hb G while 16 babies gave values between 25 and 34%. Studies of -chain gene organization on the parents of one neonate with a Hb G level of 27% at birth and 37% at 8 months excluded the presence of chromosomes with triplicated -chain genes which could lead to the ^0G/ genotype. Rather, these studies on five Hb G heterozygotes from three families confirmed the linkage between Hb G and a specific type of -thalassemia-2 associated with the presence of a 16-kbp Bgl II fragment which most probably carries the ^G locus since it has been found in 19 Hb G heterozygotes studied to date. The presence of an -thal-2 heterozygosity and three -chain genes (^0G/) was confirmed among Hb G heterozygotes with lower proportions of this variant. It is likely that the even lower values found in some newborn could arise through defective assembly of ^G- dimers. The presence of an -thal-2 homozygosity and two active -chain genes, one on each chromosome (^0G/⁰), was confirmed among heterozygotes with the higher proportion of Hb G. One of each of these categories was present in each of the three families investigated. This type of variability in the number of active -chain genes due to a heterozygosity or a homozygosity for -thalassemia-2 explains the trimodality of Hb S percentages among heterozygotes and the atypical hematological or biosynthetic features among patients with -thalassemia and sickle-cell syndromes.This research was supported by USPHS Research Grants HLB-05168 and HLB-15158 and by designated research funds of the Veterans Administration. This is Contribution No. 0693 of the Department of Cell and Molecular Biology, Medical College of Georgia, Augusta.     

Structure dynamics of the hemoglobin mutants Hb H?tel Dieu, HbG Philadelphia, HbJ Mexico, Hb St. Mandé and Hb San Diego, studied by nanosecond-laser-flash photolysis

The kinetics of the change from the carboxy to the deoxy conformation of the mutated hemoglobins mentioned in the title and of normal human adult hemoglobin were determined from measurements of light absorption changes occurring up to 50 microseconds after nanosecond-laser photodissociation of the corresponding CO complexes. The spectral evolution of the mutated hemoglobins was found to be similar in its main features to that of normal hemoglobin. The kinetics could be decomposed into two phases with rates 1.1-1.8 x 10(6) s-1 and 0.17-0.34 x 10(6) s-1 (except Hb St. Mandé which displayed only the faster phase). Study of the mutated subunits of HbJ Mexico (alpha subunit) and Hb H?tel Dieu (beta subunit) showed that they convert exponentially to the stable deoxy state after photodeligation at the same rates as the corresponding subunits of normal Hb: 1.1 x 10(6) s-1 (alpha) and 0.3 x 10(6) s-1 (beta). The results indicate that there is no direct correlation between the kinetics of spectral relaxation in the time range studied and the oxygenation properties for these hemoglobins. However, there is some indication that the kinetics are dependent upon the region of mutation.     

Clinical severity of β-thalassaemia/Hb E disease is associated with differential activities of the calpain-calpastatin proteolytic system

Earlier observations in the literature suggest that proteolytic degradation of excess unmatched α-globin chains reduces their accumulation and precipitation in β-thalassaemia erythroid precursor cells and have linked this proteolytic degradation to the activity of calpain protease. The aim of this study was to correlate the activity of calpain and its inhibitor, calpastatin, with different degrees of disease severity in β-thalassaemia. CD34(+) cells were enriched from peripheral blood of healthy individuals (control group) and patients with mild and severe clinical presentations of β(0)-thalassaemia/Hb E disease. By ex vivo cultivation promoting erythroid cell differentiation for 7 days, proerythroblasts, were employed for the functional characterization of the calpain-calpastatin proteolytic system. In comparison to the control group, enzymatic activity and protein amounts of μ-calpain were found to be more than 3-fold increased in proerythroblasts from patients with mild clinical symptoms, whereas no significant difference was observed in patients with severe clinical symptoms. Furthermore, a 1.6-fold decrease of calpastatin activity and 3.2-fold accumulation of a 34 kDa calpain-mediated degradation product of calpastatin were observed in patients with mild clinical symptoms. The increased activity of calpain may be involved in the removal of excess α-globin chains contributing to a lower degree of disease severity in patients with mild clinical symptoms.     

Hb M Milwaukee: Direct detection of the β-globin gene mutation in three generations of an afflicted family

Chromosomal DNA from three individuals with familial hemoglobin M (Hb M) Milwaukee was studied by restriction endonuclease analysis. The segregation of the mutant beta-globin gene could be followed through three generations by direct Sst I analysis at the gene level. Various restriction endonucleases were used to confirm the positions of Sst I sites in the delta-beta A- and delta-beta Mi-globin gene regions.     

Genetic and biosynthetic studies of families carrying hemoglobin J α Mexico: Association of α-thalassemia with Hb J

Summary Hemoglobin J Mexico, an chain mutant, was studied in eight unrelated Algerian families. The quantities of the abnormal hemoglobin in 116 subjects are trimodally distributed: 55% in homozygotes, 31% and 38% in heterozygotes. Both hematological data and the / chain biosynthetic ratio are normal in heterozygotes with 31% Hb J and in homozygotes. In contrast, the MCV and MCH as well as the / biosynthetic ratio are slightly reduced in heterozygotes with 38% Hb J and in their relatives carrying Hb A. The elevated expression of ^J chains in heterozygotes with 38% Hb J may be due to an thalassemia gene trans to the ^>J locus.     

Experimental analysis of Hb oxy–deoxy transition in single optically stretched red blood cells

Raman confocal microscopy, combined with an optical stretcher, is used to study the spatial distribution and the oxidation state of hemoglobin in erythrocytes under stretching condition. In particular, a near infrared laser (λ = 1064 nm) is used to generate multiple time-sharing Optical Tweezers to trap and stretch a single erythrocyte, while a second laser (λ = 532 nm) acts as Raman probe. Our study demonstrates that stretching induces hemoglobin transition to the deoxygenated state. Moreover, by using Principal Component Analysis we prove the reversibility of the $\text{oxy}?\text{deoxy}$ hemoglobin transition after application of the optically induced mechanical stress.     

Role of curcuminoids in ameliorating oxidative modification in β-thalassemia/Hb E plasma proteome

Thalassemic patients often exhibit high levels of oxidative stress and iron overload, which can lead to hazardous complications. Curcuminoids, extracted from the spice turmeric, are known to have antioxidant and iron-chelating properties and have been proposed as a potential upstream therapy of thalassemia. Here we have applied proteomic techniques to study the protein profile and oxidative damage in the plasma of β-thalassemia/Hb E patients before and after treatment with curcuminoids. In this study, 10 β-thalassemia/Hb E patients were treated with 500 mg curcuminoids daily for 12 months. The plasma protein profile and protein carbonyl content were determined at baseline, 6 and 12 months using two-dimensional fluorescence difference gel electrophoresis and carbonyl immunoblotting, respectively. Other hematological, clinical, and biochemical parameters were also analyzed. Twenty-six spots, identified as coagulation factors and proteins involved in iron homeostasis, showed significantly decreased intensity in thalassemic plasma, compared to those of normal subjects. Treatment with curcuminoids up-regulated the plasma levels of these proteins and reduced their oxidative damage. Serum non-transferrin bound iron, platelet factor-3 like activity, oxidative stress parameters and antioxidant enzymes were also improved after curcuminoids treatment. This study is the first proteomic study of plasma in the thalassemic state and also shows the ameliorating role of curcuminoids towards oxidative stress and iron overload in the plasma proteome.     

α-Thalassemia Associated with Hb Instability: A Tale of Two Features. The Case of Hb Rogliano or α1 Cod 108(G15)Thr→Asn and Hb Policoro or α2 Cod 124(H7)Ser→Pro.

We identified two new variants in the third exon of the α-globin gene in families from southern Italy: the Hb Rogliano, α1 cod108 ACC>AAC or α1[α108(G15)Thr→Asn] and the Hb Policoro, α2 cod124 TCC>CCC or α2[α124(H7)Ser→Pro]. The carriers showed mild α-thalassemia phenotype and abnormal hemoglobin stability features. These mutations occurred in the G and H helices of the α-globin both involved in the specific recognition of AHSP and β1 chain. Molecular characterization of mRNA, globin chain analyses and molecular modelling studies were carried out to highlight the mechanisms causing the α-thalassemia phenotype. The results demonstrated that the α-thalassemia defect associated with the two Hb variants originated by different defects. Hb Rogliano showed an intrinsic instability of the tetramer due to anomalous intra- and inter-chain interactions suggesting that the variant chain is normally synthesized and complexed with AHSP but rapidly degraded because it is unable to form the α1β1 dimers. On the contrary in the case of Hb Policoro two different molecular mechanisms were shown: the reduction of the variant mRNA level by an unclear mechanism and the protein instability due to impairment of AHSP interaction. These data highlighted that multiple approaches, including mRNA quantification, are needed to properly identify the mechanisms leading to the α-thalassemia defect. Elucidation of the specific mechanism leads to the definition of a given phenotype providing important guidance for the diagnosis of unstable variants.     

Hb Q-H病的生化遗传研究

本文报道在江西萍乡发现的我国第一个Hb Q-H病家系的研究结果,并对Hb Q的化学结构和Hb Q-H病的遗传学进行讨论分析。     

Evaluation of HPFH and δβ-thalassemia mutations in a Brazilian group with high Hb F levels

Fetal hemoglobin (Hb F) is characteristic of the fetal development period. However, in some genetic conditions, such as hereditary persistence of fetal hemoglobin (HPFH) and delta-beta thalassemia (δβ-thalassemia), Hb F continues to be produced in adulthood. We evaluated the frequency of two mutations of HPFH, HPFH-1 and HPFH-2 African, and two mutations in δβ-thalassemia, Sicilian and Spanish, in a Brazilian population. Peripheral blood samples were collected from adults from hospitals and blood centers in southeast and northeast Brazil. These individuals were healthy and without complaints of anemia, but had increased Hb F. Samples were submitted to electrophoretic and chromatographic analyses to quantify Hb F values and, subsequently, to molecular analyses to verify the mutations. In the molecular analysis, 16 of the 60 samples showed a heterozygous profile for the HPFH mutations, two for HPFH-1 and 14 for HPFH-2. In the same sample set, three were heterozygous for Spanish δβ-thalassemia and none were heterozygous for Sicilian δβ- thalassemia. The Hb F values in the HPFH-2 heterozygotes differed from those previously reported for this mutation. In this group, the HPFH mutations were more frequent than the δβ-thalassemia mutations. The finding of these mutations in this Brazilian population reflects the mixing process that occurred during its formation.     

The interaction of hemoglobin O Arab with Hb S and β+ thalassemia among Israeli Arabs

Summary We have studied 105 individuals in the village of Jasser El Zarka in the Northern Coast of Israel of whom 59% had at least one abnormal hemoglobin. Of the individuals studied 41% were AA, 13.3% AS, 28.6% AO_Arab, 10.5% SO_Arab, 0.9% SS, 38% Arab-⁺ Thal, and 1.9% Thal trait. The SO_Arab double heterozygotes were characterized by a normal mean corpuscular volume (MCV) and mean corpuscular hemoglobin concentration (MCHC), and an increase of Hb F (11.7±4.3%) and 2,3-diphosphoglycerate levels (27.8 m/g Hb). The increase of Hb F is higher than the one seen among O_Arabs of other ethnic backgrounds. Their clinical course was moderately severe and osteoporosis was quite frequent. The interactions of Hb O_Arab and Hb S were studied in vitro and it was confirmed the Hb O_Arab lowers the minimal gelling concentration of mixtures with Hb S (as compared to mixtures of Hb S and Hb A), but that this effect is ionic-strength dependent. Our data are in conflict with previous claims that Hb O_Arab mixtures with Hb S polymerized almost as much as pure S. Oxygen association curves show a significant displacement of the p50 to the right, but the effect of oxygen dissociation is less apparent. The displacement was not nearly as significant as with SS cells, confirming our gelation data. Blood group determinations establish that these Arab populations had black African admixture.The Hb O_Arab/⁺ Thal double heterozygotes exhibit moderate anemia (10.3g% of Hb) and the percentage of Hb A was 17.2±1.8%. The fetal Hb was 5.4±2.1% and the 2,3-diphosphoglycerate level in two cases was 17.4 mol/g Hb. The only case of a homozygote SS had moderate anemia (10.3 g Hb%), 25.7% of Hb F, and a very benign course.     

The levels of ζ, γ, and δ chains in patients with Hb H disease

Summary Details are given of a study of blood samples from 24 patients with Hb H disease from different Mediterranean countries and from the Far East. Four different types of -thal-1 (--) were observed, namely-() ( 20.5-kb deletion);--MED-I ( 17.5-kb deletion);--MED-II (>26.5-kb deletion); and--SEA ( 18-kb deletion, in Orientals only). The -thal-2 was mainly of the deletion type (16 with the 3.7-kb deletion; 1 with the 4.2-kb deletion), while 4 of the 7 patients with a nondeletional type had the five-nucleotide deletion at the donor splice site of the first intron of the 2 gene. All patients had a mild-to-moderate hemolytic anemia; no significant differences in hematology were observed between the groups. Hb A₂ was decreased to about one-third of the normal level. The Hb H formation varied considerably and its quantitation was not always satisfactory. Patients with Hb H disease due to any -thal-1 combined with a nondeletional -thal-2 had the highest Hb H levels and a more marked anemia. The chain production was small and absent in patients with the MED-II type of -thal-1 because this deletion included the and genes. The highest chain levels were present in the four patients with the SEA type of -thal-1. The chain production was increased, particularly in patients with a mutation of C T at position-158 to the ^G globin gene. This chain was primarily present as Hb Bart's (or ₄) and only about 15% was recovered as Hb F or ₂₂. The evaluation of the rate of chains produced in these patients was greatly facilitated by data from one patient who had Hb H disease and a heterozygosity for the ^A-⁺. The low levels of Hb A₂ and of Hb F (relative to Hb Bart's) can be explained by a decreased affinity of chains for and chains as compared with chains in conditions of severe chain deficiency.     

Does Gγ/Aγ ratio and Hb F level influence the severity of sickle cell anaemia

Sickle cell anaemia (SCA) exhibits significant variations in clinical presentation in different populations for which several genetic factors including SCA-associated -and -thalassaemias, G-6-PD deficiency and elevated Hb F level have been implicated as possible ameliorating factors. Saudi Arabia is unique in that mild and severe forms of the disease occur at a high frequency. We investigated the G/A ratio and Hb F level and correlated these values with the severity of SCA. The results showed that Hb F level varies significantly in both groups of patients with no evident correlation with the mild clinical manifestations. However, G/A ratio correlated significantly with the disease severity where a high ratio was observed in patients with the mild and a low ratio in patients with the severe disease. The results are evaluated and discussed in the light of correlation studies and regression analysis.     

巴西橡胶树HbγGCS基因克隆及表达分析

γ-谷氨酰半胱氨酸合成酶（TGCS）是细胞内谷胱甘肽（GSH）生物合成的限速酶,GSH在植物许多生理过程中发挥重要作用。本研究采用简并PCR和RACE技术获得巴西橡胶树γGCS基因全长cDNA序列,命名为HbTGCS（GenBank登录号：GU997638）。该基因全长2187bp,最长开放阅读框为1572bp,编码523个氨基酸。进化分析结果表明HbγGCS属双子叶植物γGCS亚类,同葡萄γGCS分为一组,与单子叶植物的亲缘关系较远。半定量RT-PCR结果表明HbγGCS基因在胶乳、叶片、树皮、花中均有表达,以花中表达量最大。健康橡胶树胶乳中HbγGCS达量高于死皮树。HbγGCS表达受乙烯、茉莉酸、过氧化氢、机械伤害、干旱、低温和高盐调控。     

Hb a 2 gluß2 (Hb I) in a Caucasian family: Independent mutation or common origin?

Summary Hemoglobin 16glu (Hb I-Skamania) was confirmed in six persons of a Caucasian family by amino acid analysis of the abnormal tryptic peptide, T3,4. The confirmation of Hb 16glu in Caucasians and the apparent absence of Hb I in racially unmixed Negroes or in American Indians suggest that Hb 16gly may be of European origin. However, an independent European and African origin can currently not be ruled out. The origin of seven other rare hemoglobin mutants is also uncertain. Independent genetic origin of a rare mutant is confirmed only when a new mutation can be proved.     

Hb疫苗无反应者的免疫效应研究

<正>乙型肝炎(HB)疫苗70年代后期引入美国,1982年正式批准使用。预料使用该疫苗将显著减少乙肝病毒(HBV)感染的发病率。慢性乙肝与原发性肝癌之关系目前已经证实,因而,广泛使用HB疫苗,尤其在亚洲和非洲地区,将显著减少这种恶性肿瘤的发生。