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S Fucharoen

Publications and source records attributed to S Fucharoen.

At least 91 records · Page 5Linked to original sources
PubMed1995

Role of alternatively spliced beta E-globin mRNA on clinical severity of beta-thalassemia/hemoglobin E disease.

In spite of seemingly identical genotypes, severity of beta-thalassemia/hemoglobin (Hb) E patients can vary greatly. Some may have a severe clinical disorder approaching that seen in homozygous beta-thalassemia. Since mutation in codon 26 of the beta E-globin gene can lead to an alternative splicing, Hb E acts like a mild beta(+)-thalassemia. Variation in the amount of beta E-globin mRNA may also govern the difference in severity of anemia in beta-thalassemia/Hb E patients who otherwise have the same genetic determinants. We have determined the percentage of the alternatively spliced beta E-globin mRNA by the RT-PCR technique in 14 patients and found that the amount of abnormal spliced beta E-globin mRNA in those patients with severe symptoms ranged between 2.9 to 6.1%, whereas those with milder symptoms had the values which ranged between 1.6 to 2.6%. The extent of beta E-globin mRNA cryptic splicing was better associated with clinical severity of the patients than did the patterns of the Xmn I polymorphism at position -158 of the G gamma-globin gene or levels of Hb F.

Adolescent↗
PubMed1995

Expression of hemoglobin E in newborn.

Hemoglobin E(HbE) is an abnormal hemoglobin resulted from a point mutation in codon 26 (GAG-AAG) of beta-globin gene. The mutation causes an amino acid substitution (Glu-Lys) and abnormal splicing in exon 1 that reduce the amount of beta E chain synthesis. While in adult, the HbE can easily be diagnosed, its level in newborn is usually underrepresent. In this study, we examined a relationship between genotype of HbE and the amount of HbE in cord blood. 145 Cord blood specimens were analysed by starch gel electrophoresis and the amounts of HbE were determined by microcolumn chromatography. The zygosity of beta E globin gene was determined by the polymerase chain reaction. The levels of HbE were 3.17 +/- 1.79% for 59 heterozygotes, 8.55 +/- 2.52% for 3 homozygotes and 0.48 +/- 0.22% in 83 normal newborns. This result provides useful data for a neonatal screening program and implies that expression of HbE in newborn dependent on a copy number of beta E globin gene in each individual.

Adult↗
PubMed1995

Molecular basis of alpha (0)-thalassemia in northeast of Thailand.

The molecular basis of alpha(0) thalassemia were studied in 95 patients who attended at Srinagarind Hospital, Khon Kaen University during September 1993-January 1994. From these 95 patients, hemoglobin electrophoresis indicated that there were 14 cases with A2AH, 21 cases with A2ABart'sH, 6 cases with ConSpA2AH, 31 cases with ConSpA2ABart'sH, 6 cases with EABart's, 3 case with EFBart's, 4 cases with ConSpEABart's, 5 cases with Portland Bart's and 5 cases with A2A. White blood cell lysate was prepared from peripheral blood leukocytes of these patients and was subjected to the polymerase chain reactions for detection of the SEA type alpha thalassemia gene deletion. Altogether 100 chromosomes with the SEA deletion were detected from all patients examined, the result indicated that this type of alpha(0) thalassemia gene deletion is the most common among these Thai population. These data will be useful for a carrier screening and a prenatal diagnosis program of homozygous alpha (0) thalassemia which is a lethal condition in northeast of Thailand.

DNA↗
PubMed1995

Alterations and pathology of thalassemic red cells: comparison between alpha- and beta-thalassemia.

Two main types of thalassemia have been categorized according to defective production of the globin gene ie alpha-thalassemia and beta-thalassemia. We report different red cell abnormalities between these two types. The study included 139 thalassemic patients including 91 patients with hemoglobin (Hb) H disease (52 cases with the classical genotype and 39 cases with Hb Constant Spring) and 48 were beta-thalassemia/Hb E disease. The deformability index of thalassemic red cells measured by laser diffractometer was significantly lower than that of normal red cells. Increased susceptibility of the thalassemic red cells to monocyte phagocytosis was markedly noted. Few sialic acid molecules were scattered on red cell surface of thalassemic red cells. Reticulocytes with delayed maturation stage were also observed in thalassemia indicating enhanced release from the bone marrow. The alpha-thalassemic red cells had relatively better deformability, increased susceptibility to phagocytosis, reduced sialic acid content and greater degree irregular distribution of sialic acid on red cell surface as compared to beta-thalassemic red cells. The alpha-type with hemoglobin Constant Spring (Hb CS) had increased percentage of reticulocyte and young reticulocyte (high fluorescent intensity) as compared to beta-thalassemic red cells. The different abnormalities between alpha- and beta-thalassemic red cells may lead to different mechanism of red cell destruction and different severity of the disease.

Erythrocyte Deformability↗
PubMed1995

A simple non radioactive method for detecting beta-thalassemia/hbe disease: application to prenatal diagnosis.

We have developed allele specific polymerase chain reaction (ASPCR) that allows rapid screening of the beta E-globin and common beta-thalassemia genes in Thailand. These non-radioactive methods are based on the amplification by the polymerase chain reaction of the beta E and beta-thalassemia specific DNA fragments using specific primers. With this approach, both heterozygote and homozygote for the disease could readily be identified on agarose gel electrophoresis of the amplified DNA. We have applied the method for a prenatal diagnosis of beta-thalassemia/HbE disease in a Thai family at the second trimester of pregnancy. The result obtained was comparable to that of conventional dot blot hybridization using radioactive probes. The simplicity, accuracy and non isotopic of the approach make it a highly promising method for a carrier screening and a prenatal diagnosis of this common disorder.

Alleles↗
PubMed1995

Prenatal diagnosis for beta-thalassemia syndromes using HRP-labeled oligonucleotide probes at Siriraj Hospital.

Characterization of the molecular defect of beta-thalassemia in Thais has enabled us to establish prenatal diagnosis for homozygous beta-thalassemia and beta-thalassemia/Hb E. The nature of the beta-thalassemia mutation of each high risk couple or of the previous affected child was firstly identified after counseling. Detection of beta-thalassemia mutations was performed by dot-blot hybridization of the amplified DNA with a set of HRP-labeled ASO-probes specific for the common mutations. If the mutation could be characterized, prenatal diagnosis (PND) would be performed by using DNA extracted either from the chorionic villi (CVS) or amniotic fluid fibroblast in the first trimester of pregnancy or from fetal blood in the second trimester. DNA analysis was carried out in 23 couples at risk of having homozygous beta-thalassemia and 88 couples at risk for beta-thalassemia/Hb E. However, PND was performed by this technique in 22 pregnancies from 21 couples at risk of having homozygous beta-thalassemia children and 86 pregnancies from 71 couples at risk for beta-thalassemia/Hb E; 9 couples underwent more than one prenatal diagnosis. The results showed that, although there are more than 20 beta-thalassemia mutations in the Thai population, PND by DNA analysis could be carried out in more than 95% of the risk couples by using beta(E) and 10 different HRP-labeled ASO probes. This technique was simple, economic and avoided the use of radioactive isotope.

Amniocentesis↗
PubMed1994

Flow cytometric immunophenotyping of lymphocyte subsets in samples that contain a high proportion of non-lymphoid cells.

Flow cytometric (FCM) immunophenotyping of peripheral blood from thalassemia patients presents technical difficulties because of the high proportion of immature red cells. The combination of forward scatter (FSC) and side scatter (SSC) with fluorescence associated with human leukocyte antigen/monocyte antigen (CD45/CD14) was unable to identify the lymphocyte population in thalassemia patients; therefore, it was necessary to exclude immature red cells to analyze lymphocyte subsets in these patients. A simultaneous three-color FCM method was developed, with the basis that transferrin receptor (CD71) or glycophorin A (glyco A) is present on all immature red cells, but is not expressed on CD45 positive leukocytes. In this study, the lymphocyte population was identified by gating out unwanted cell populations using the FSC/CD71-fluorescein isothiocyanate (FITC), FSC/glyco A-FITC, or FSC/CD45-peridinin chlorophyll protein (PerCP) profiles. The CD71-FITC negative cells, glyco A-FITC negative cells, or CD45-PerCP positive cells were identified, then analyzed on the basis of FSC/SSC to allow any remaining non-lymphocyte cells in FSC/SSC gate to be excluded. The cells in FSC/SSC gate were then analyzed using other irrelevant two-color antibodies. Of the three gating strategies, CD45-PerCP and glyco A-FITC methods are better than the CD71-FITC gating method. Both methods markedly increase the purity of lymphocytes in the analysis gate. Either method is easy, straightforward, requires a six-tube set of reagent tubes, and provides a reliable method for immunophenotyping lymphocyte subsets in preparations containing a large percentage of non-lymphoid cells.

Adult↗
PubMed1994

Hydration of red cells in alpha and beta thalassemias differs. A useful approach to distinguish between these red cell phenotypes.

Differences in the pathobiologic features of alpha and beta thalassemic red cells include differing derangements in the ability of these cells to regulate their volume. To explore differences in cell hydration in these two red cell phenotypes, heterogeneity in hemoglobin concentration of individual red cells was quantitated. Red cells from 91 patients with alpha thalassemia (hemoglobin H disease) and 69 patients with beta thalassemia of the genotype beta-thal zero/Hb E were analyzed. Marked differences between these thalassemias were noted in the heterogeneity of hemoglobin concentration among individual cells. Decreased cell hemoglobin concentration and increased cell hydration were features of alpha-thalassemic blood samples, whereas both decreased and increased cell hemoglobin concentration as a result of cell hydration and dehydration, respectively, were features of beta-thalassemic blood samples. The documented differences in the hemoglobin concentration distributions should prove useful in distinguishing between the two thalassemic phenotypes.

Erythrocyte Indices↗
PubMed1994

Nondeletional type of hereditary persistence of fetal haemoglobin: molecular characterization of three unrelated Thai HPFH.

The beta-globin gene clusters of three unrelated Thai families with a nondeletional type of hereditary persistence of fetal haemoglobin (HPFH) were studied using polymerase chain reaction-related techniques. All appeared to have normal nucleotide sequences from the Cap site to position -400 of both the G gamma- and A gamma-globin genes. Two individuals suspected of having a beta-thalassaemia gene linked to the high HbF condition also had a normal beta-globin gene sequence, spanning from position -108 from the Cap site to the polyadenylation site. Deletion of four nucleotides, AGCA, at positions -225 to -222 of one A gamma-globin allele was detected in one subject and was confirmed by dot-blot hybridization. Restriction fragment length polymorphisms in the beta-globin gene cluster showed that the 5' haplotype (-+-++) and the presence (+) of an Xmm 1 polymorphic site at -158 of the G gamma-globin gene are associated with the high F phenotype in these families. Direct sequencing of the 5' hypersensitive-2 (5' HS-2) site of the locus control region (LCR) showed that this Xmn 1 (+) site is also linked to a specific rearrangement of TA repeats (TA)9CACATATACG(TA)10, in HS-2 segment.

Adult↗
PubMed1994

Number and maturation of reticulocytes in various genotypes of thalassaemia as assessed by flow cytometry.

Ineffective erythropoiesis is a prominent defect leading to anaemic status in thalassaemic patients. Reticulocyte enumeration in the peripheral blood is a non-aggressive method of measuring bone marrow erythropoietic activity. We used an automated reticulocyte counter (Sysmex R-3000) to determine the number and maturation level of circulating reticulocytes among various types of thalassaemia: non-splenectomized beta-thalassaemia/haemoglobin E (beta E) and splenectomized cases (beta E-S), classical haemoglobin H disease (H), haemoglobin H disease with haemoglobin Constant Spring (H/CS), homozygous haemoglobin Constant Spring (CS/CS), homozygous haemoglobin E (EE), heterozygous thalassaemics and other rare combinations. Haemoglobin H disease has a higher absolute count than beta-thalassaemia (beta E), indicating relatively better compensatory erythropoiesis in haemoglobin H disease. Those with CS genes (H/CS and CS/CS) have poorer reticulocyte maturation than any other type of thalassaemia with rather high absolute numbers, especially in H/CS. This indicates a severer degree of ineffective erythropoiesis in beta-thalassaemia (beta E), which reflects an insufficient rise in reticulocyte number in comparison with alpha-thalassaemia (H). The presence of haemoglobin Constant Spring is associated with abnormally low reticulocyte maturation due to enhanced erythrocyte production or direct effect of Constant Spring globin itself, both still unexplained with the current information. The splenectomized beta E has increased reticulocyte number and cells with high DNA content, probably nucleated red cells, designated as the upper particle count parameter. However, there is the same degree of reticulocyte maturation in non-splenectomized and splenectomized beta E patients, suggesting a role for splenic pooling of reticulocytes.

Erythrocyte Count↗
PubMed1994

Molecular basis of beta-thalassemia in Japan: heterogeneity and origins of mutations.

Characterization of beta-thalassemia mutations was attempted for 13 unrelated Japanese patients heterozygous for beta-thalassemia. We have systematically analyzed beta-thalassemia genes using polymerase-chain-reaction-related techniques; dot blot hybridization with oligonucleotide probes complementary to known mutations, restriction endonuclease assay and direct sequencing of amplified genomic DNA. Seven different mutations were detected. Six of them are an amber mutation in codon 90 (GAG to TAG), a four-base-pair deletion in codons 41 and 42 causing premature termination due to frameshift, a C-T substitution at position 654 of IVS-2, a G-A substitution at position 1 of IVS-2 and a C-G substitution at position 848 of IVS-2, leading to splicing defects, and an ocher mutation (GAA-TAA) in codon 121 causing a thalassemia intermedia phenotype with inclusion body formation in erythrocytes. A silent mutation (CTG-TTG) was also detected in codon 91 of the allele with the IVS-2 position 1 mutation. These mutations have been reported previously in the Japanese population. The other mutation is a novel one in the Japanese, an amber mutation (TGG-TAG) in codon 15, causing a beta zero-thalassemia phenotype by premature termination of the beta-globin chain synthesis. We analyzed haplotypes of chromosomes bearing each beta-thalassemia mutation. Origins and a spectrum of mutations in comparison with those detected in malaria-endemic regions are discussed.

Adolescent↗
PubMed1993

Activation of monocytes for the immune clearance of red cells in beta zero-thalassaemia/HbE.

We have recently provided evidence that IgG antibodies play a role in the destruction of red cells in thalassaemia syndromes. In order further to delineate factors involved in the clearance of thalassaemic cells, monocytes of 30 Thai patients with beta zero-thal/HbE (17 non-splenectomized and 13 splenectomized) and 16 normal controls were examined for their ability to bind and phagocytose normal red cells coated with IgG anti-Rh(D). In beta zero-thal/HbE, the mean number of red cells attached to the monocytes was approximately 3-fold greater than in normal controls and the number ingested 30% higher. Among the non-splenectomized patients, the number of red cells attached to and ingested by the monocytes, correlated inversely with mean basal Hb levels, suggesting that activation of mononuclear phagocytes for the immune clearance of red cells is a factor in determining the severity of the anaemia. As Fc-gamma-RI is of primary importance in the recognition of IgG-coated red cells by monocytes, leucocytes from 10 beta zero-thal/HbE patients (four non-splenectomized and six splenectomized) and five normal controls were investigated for their expression of Fc-gamma-RI by flow cytometry. In beta zero-thal/HbE there was an approximately 3-fold increase in the percentage of leucocytes expressing this receptor; the receptor was up-regulated on monocytes and induced on granulocytes. The up-regulation of Fc-gamma-RI in beta zero-thal/HbE is likely to be an important component in the activation of monocytes and in mediating their enhanced effector function towards antibody-coated cells.

Erythrocytes↗
PubMed1993

Severity differences in beta-thalassaemia/haemoglobin E syndromes: implication of genetic factors.

Genetic factors determining the difference in severity of anaemia in beta-thalassaemia/HbE disease were studied in 90 patients who had haemoglobin levels, at steady state, ranging from 4.2 to 12.6 g/dl. Co-inheritance of alpha-thalassaemia 2 and haemoglobin Constant Spring could significantly decrease the severity of the disease. Inheritance of a beta-thalassaemia chromosome with Xmn I cleavage site at position -158 of the G gamma-globin gene which was linked to the haplotype -+-++ or ++-++, was associated with a milder anaemia. Two copies of these alleles were necessary to produce a significant clinical effect. Increased expression of the G gamma-globin gene and higher production of haemoglobin F, which could reduce the overall globin chain imbalance, were also associated with homozygosity for the Xmn I cleavage site and thus with less severe anaemia. However, this effect was not seen in Xmn I site heterozygotes. Whether the effects of the Xmn I polymorphism, HbF concentration and G gamma/A gamma ratio act separately or through common mechanisms in reducing anaemia remains to be ascertained.

Adolescent↗
PubMed1993

Identification of Hb C [beta 6(A3)Glu-->Lys] in a Thai male.

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 beta chain indicated that the abnormal hemoglobin was Hb C [beta 6(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 beta gene, in agreement with heterozygosity for Hb C and Hb A. Furthermore, the beta gene framework (Ava II-, Bam HI+) of the propositus suggested that the beta C gene may have arisen from an independent mutation. Since Hb C and Hb E have the same mutation (Glu-->Lys) in the beta 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.

Adult↗