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E Kajii

Publications and source records attributed to E Kajii.

At least 73 records · Page 4Linked to original sources
PubMed1995

Genomic organization of the glycoprotein D gene: Duffy blood group Fya/Fyb alloantigen system is associated with a polymorphism at the 44-amino acid residue.

The Duffy blood group antigen has been characterized by its roles on red blood cells: as a receptor for the malarial parasites and as a promiscuous receptor for chemokine superfamily. Recently, the Duffy blood group associated glycoprotein D (gpFy) cDNA has been cloned (Chaudhuri et al: Proc Natl Acad Sci USA 90:10793, 1993). In this report we describe the organization of genomic DNA coding for the gpFy and elucidate the molecular nature of Fya/b polymorphisms. By a Southern blotting analysis probed with gpFy cDNA, gpFy gene was shown to be composed of three DNA fragments; 1.1-kb Sac I, 1.9-kb EcoRI, and their intervening 47-bp fragments. We cloned the 1.1-kb Sac I and 1.9-kb EcoRI fragments by inverted polymerase chain reaction (IPCR) procedure. The promoter region of the gpFy gene was cloned by IPCR of 1.1-kb Sac I fragment and the 3' flanking sequence was cloned by IPCR of 1.9 kb EcoRI fragment. The both IPCR products contained on both side the known gpFy cDNA sequence without introns, as expected. Although no TATA or CCAAT boxes are present in the promoter sequence, several transcription factor binding site motifs are contained, including AP-1, HNF-5, TCF-1, ApoE B2, W-element, H-APF-1, and Sp-1. The 3' flanking region has two additional polyadenylation signals, other than that used in the cDNA, and also has an indirect and a direct repeat sequence clustered with the 5' flanking region. These facts indicate a possibility that the gpFy gene has been evolved by multiple retrotransposition events. By comparing the coding area of the gpFy gene in 28 Duffy-positive individuals, we elucidated that one base change that results in an amino acid substitution [GA-T(Asp44)-->GGT(Gly)] is in accordance with the Fya/Fyb polymorphism. This fact proves that the gpFy cDNA and its gene described in this report encode the Duffy blood group system.

Amino Acid Sequence↗
PubMed1995

Intricate combinatorial patterns of exon splicing generate multiple Rh-related isoforms in human erythroid cells.

The Rhesus (Rh) blood group system shows complex polymorphisms in the human. Some of the heterogeneity may be generated by alternative RNA splicing. For a systematic analysis of Rh-related mRNA isoforms expressed in reticulocytes, we isolated mRNA, which was then reverse transcribed and amplified by the polymerase chain reaction (PCR) to give Rh-related cDNAs of two segments of 704 bp and 975 bp. The PCR amplification of the 5'-region yielded a single PCR product, whereas a complex electrophoretic pattern of PCR bands was derived from the 3'-region. A highly reproducible ladder of multiple additional bands migrated below the PCR products corresponding to the full-size cDNAs for RhPI and RhPII and encoding two different Rh polypeptides. Eleven and five truncated isoforms of the RhPI and RhPII cDNAs, respectively, were identified in the PCR products. These isoforms appear to be generated by combinatorial splicing of six RhPI and three RhPII exons. Our results suggest that the Rh-related polypeptides consist of a mixture of RhPI and RhPII polypeptide isoforms differing at the C terminus. Multiple RNA splicing pathways are thus operative in the two Rh-related genes even within a single cell lineage of human erythroid cells.

Alternative Splicing↗
PubMed1994

Expression of Rh blood group gene transcripts in human leukocytes.

The Rh blood group system was recognized as one of the most complex polymorphisms in human. Whether or not the Rh antigens are present on human leukocytes is still an unresolved question. The expression of the Rh gene at the mRNA was analyzed in purified populations of human leukocytes by using reverse transcription and polymerase chain reaction (RT-PCR) followed by Southern blot analysis. The PCR products of the 5'-terminal region showed a single band as expected in the CD19+TCR-1 and CD14+ cells but doubtfully in the CD13+CD71- and CD190-TCR-1+ cells. On the other hand, in all these cells, the PCR products of the 3' region exhibited multiple additional bands migrating ahead of the band as expected, which showed distinctly different sets of bands in each cell. The additional bands appeared to consist of RhPI- and RhPII-cDNA splicing isoforms. These results indicated that the expression of the Rh gene is not restricted to human erythroid lineage. Additionally, it was suggested that different transcription initiation sites might be utilized preferentially for Rh gene expression.

Antigens, CD↗
PubMed1994

A new genomic polymorphism of Rh-polypeptide genes.

To determine which Rh-polypeptide genes are related to which Rh (rhesus) antigens, genomic DNAs prepared from donors of ten kinds of different Rh phenotypes were analysed by Southern hybridization method using two probes for the 5' moiety and 3' moiety of an Rh-polypeptide cDNA. A polymorphism for the Rh blood-group system was observed with BamHI, HindIII, PstI and EcoRI restriction endonucleases between genomic DNAs from the RhD-positive and RhD-negative individuals as has previously been reported. Further study showed an additional genomic polymorphism. The new polymorphism was detected between the RhC/c phenotypes unrelated to the RhD/d phenotypes with RsaI restriction endonuclease using the probe for the 5' moiety of Rh polypeptide cDNA. A 1.9 kb RasI restriction fragment was missing in the genomes from individuals with the RhC phenotypes. However, this fragment was present in the genomes of Rhc phenotypes and in its half dose in the genomes of RhCc phenotypes. These results clearly demonstrate a new polymorphism between the RhC/c phenotypes regardless of the RhD-positive and RhD-negative phenotypes, and it is predicted that the Rh polypeptide genes encoding RhD, RhC/c and RhE/e antigens are different.

DNA, Complementary↗
PubMed1994

Molecular analysis of Rh polypeptides in a family with RhD-positive and RhD-negative phenotypes.

To investigate the genetic basis of the Rh polypeptide gene, we attempted the isolation of cDNA clones for Rh polypeptide from a family with the RhD-positive and RhD-negative phenotypes using the reverse transcription (RT)-PCR method for each reticulocyte RNAs followed by subcloning. The isolated cDNAs showed the existence of another Rh-related clone (RhPII-1 cDNA, tentative designation) besides the RhPI and RhPII cDNA clones reported previously by us. The RhPII-1 cDNA had a single nucleotide substitution with one amino acid substitution compared with the RhPII cDNA:substitution C-->T in nucleotide 380, changing codon 127 from GCG to GTG (Ala-->Val). The RhPI, RhPII, and RhPII-1 cDNA clones were detected in all individuals by the PCR experiment. This suggests that the Rh polypeptide genes have been inherited from parents and might be highly polymorphic. The PCR amplification of an RhPII-specific region from reticulocyte RNA and genomic DNA in all the family proved that the RhPII gene exists in both RhD-positive and RhD-negative individuals. By Southern-blot analysis of the DNAs from the family, two independent polymorphisms concerning the RhC/c and RhD/d phenotypes were observed. These results demonstrate that the RhPI and RhPII genes are also present in the RhD-negative donors, and the RhPII-related cDNAs encode not the RhD, but the RhC/c and/or E/e, polypeptides.

Base Sequence↗
PubMed1994

Identification of two Rh mRNA isoforms expressed in immature erythroblasts.

Two isoforms of RhPI cDNA encoding an Rh polypeptide, RhPI-alpha and RhPI-beta, were isolated from mRNAs of immature erythroblasts cultured in a selective two-phase liquid culture system for human erythroid progenitors in peripheral blood. The RhPI-alpha and RhPI-beta cDNA clones encoded the hydrophobic membrane polypeptides of 242 and 244 amino acids which were missing in amino acid residues 227 to 383 and 212 to 384 of RhPI polypeptide, respectively. These peptide deletions produce unexpected structural properties resulting in the generation of a new open reading frame with a new stop codon caused by frameshift translations in RhPI-alpha cDNA and in the reverse orientation of topology in RhPI-beta cDNA. Both isoforms are abundantly present in immature erythroblasts, but not in mature erythroblasts. These results indicate that the two Rh-related mRNAs identified are expressed at the immature erythroblast stage on the development of erythroid series.

Amino Acid Sequence↗
PubMed1994

An exon 28 mutation resulting in alternative splicing of the glycoprotein IIb transcript and Glanzmann's thrombasthenia.

The alternatively spliced from mRNA of platelet glycoprotein IIb (GPIIb) with a deletion of exon 28 (GPIIb-28) has been isolated from the HEL cell cDNA library. The defective expression on the surface of DNA cotransfected COS-1 cells with GPIIb-28 and GPIIIa cDNAs was described in an earlier report. We studied siblings with Glanzmann's thrombasthenia who expressed only the GPIIb-28 mRNA in their platelets. Flow cytometry showed that the patients' platelets failed to bind GPIIb/IIIa complex-specific and GPIIb-specific monoclonal antibody. Western blotting showed that the patients' platelets had defective GPIIb and have trace amounts of GPIIIa. Sequence analysis was performed after polymerase chain amplification of the patients' GPIIb and GPIIIa mRNAs. The patients' GPIIb cDNA had a deletion of the exon 28 nucleotides. The polymerase chain reaction (PCR) from exon 27 to 29 showed that the GPIIb-28 mRNA was 3% +/- 1.6% of the normally, spliced form in control platelets, and 61% in the megakaryoblastic cell line UT-7. The patients' platelets showed only the GPIIb-28. Family study and quantitative PCR studies showed that these patients were compound heterozygotes of two GPIIb gene defects. The father's allele is described in this report and involves skipping exon 28 secondary to a base substitution at codon Gln948, CAG-->TAG. The mother's allele appears to involve decreasing GPIIb mRNA levels in platelets. Our results indicate that the GPIIb-28 is not expressed on platelet membranes as a stable GPIIb/IIIa heterodimer or left as a monomer in platelets. Our studies confirm the previous data observed in COS-1 cells expressing recombinant GPIIb-28.

Alternative Splicing↗
PubMed1994

The electrokinetic behavior of red blood cells from a patient with Tn syndrome by Doppler electrophoretic light scattering analysis.

The Tn syndrome is an acquired form of persistent mixed-field polyagglutination displaying two distinct populations of Tn positive (Tn) and Tn negative (tn) red blood cells (RBCs). We investigated the electrophoretic behavior of RBCs showing polyagglutination from a patient with Tn syndrome by the doppler electrophoretic light scattering (D.E.L.S.) analysis. The mean of zeta potential of normal RBCs from ten individuals was -13.07 +/- 0.61 mV (mean +/- S.D.). The content of membrane-associated sialic acid equated with the zeta potential of RBCs. Among the proteases ficin was most effective on the zeta potential of RBCs. The zeta potential of the patient Tn RBCs and tn RBCs were -4.73 mV and -13.32 mV, respectively. Tn RBCs reduced 64.5% of zeta potential compared with tn RBCs and formed 48.8%. These results may provide some useful information for classification of Tn syndrome.

Antigens, Tumor-Associated, Carbohydrate↗
PubMed1994

[A new approach for diagnosis of autoimmune hemolytic anemia].

Sixty four cases of autoimmune hemolytic anemia (AIHA) referred to our laboratory from 1985 to 1993 consisted of 51 warm type AIHA, 9 cold agglutinin disease (CAD), one paroxysmal cold hemoglobinuria, and 3 mixed type AIHA. There were 5 patients who had all clinical features of AIHA except for a positive direct-antiglobulin test (DAT). These patients were diagnosed as DAT-negative AIHA because of the elevation of red blood cell-associated IgG (RBC-IgG). The mean RBC-IgG of 100 healthy individuals was 33 +/- 13 (SD) molecules per one RBC. On the other hand, the RBC-IgG of patients with DAT-positive and with DAT-negative warm type AIHA were ranged from 257 to 12,421 and from 126 to 256 molecules per one RBC, respectively. One of 9 patients with CAD had a hemolytic anemia associated with only a cold agglutinin titer of 32 using saline-suspended RBCs, but a titer of 4,096 in the presence of bovine albumin. This case was diagnosed as low titer CAD. All of patients with DAT-negative AIHA or low titer CAD showed a good response to corticosteroid treatment.

Anemia, Hemolytic, Autoimmune↗
PubMed1993

Isolation of a new cDNA clone encoding an Rh polypeptide associated with the Rh blood group system.

The polymerase chain reaction (PCR) was used to amplify Rh-related cDNAs from erythroid cells cultured by the selective two-phase liquid culture system for human erythroid progenitors in peripheral blood. Direct sequencing based on PCR presents heterozygous bands. Two Rh polypeptide cDNAs have been isolated from the PCR products and tentatively designated RhPI cDNA and RhPII cDNA. Both cDNA clones have an open reading frame composed of 1251 nucleotides. The RhPI cDNA clone shows a single nucleotide substitution with no amino acid substitution compared with the published sequence. The RhPII cDNA clone, on the other hand, differs from the above by 41 nucleotide substitutions with the open reading frame, resulting in 31 amino acid substitutions.

Amino Acid Sequence↗
PubMed1993

Point mutation in the band 4.2 gene associated with autosomal recessively inherited erythrocyte band 4.2 deficiency.

A patient who represented acute hemolytic crisis was studied. Analysis of the erythrocyte membrane proteins by SDS-PAGE revealed a deficiency of band 4.2. In the family, the sister of the patient who had been clinically normal was also shown to be deficient in band 4.2. Binding studies showed that the propositus' membranes were able to bind normal band 4.2 protein as much as control. It was suggested that the binding sites for the protein were prepared on the membrane. We analyzed the band 4.2 cDNA of the propositus and detected a mutation that changes a codon for alanine to one for threonine at residue 142. Band 4.2 exon III of genomic DNA which included the mutation site was amplified and sequenced directly in the family members, and it was revealed that only the homozygotes of the mutation allele manifested band 4.2 deficiency and the parents, who were heterozygotes, showed normal amounts of band 4.2. Recently, the same mutation was reported as Protein 4.2NIPPON in another 4 cases (Bouhassira et al. Blood 1992: 79: 1846-1854). This study supports the hypothesis that this mutation is the pathogenetic cause of band 4.2 deficiency and not a polymorphism.

Adult↗
PubMed1993

A patient with Tn syndrome associated with myelodysplastic syndrome showed abnormal glycophorin B.

A Japanese male patient with myelodysplastic syndrome (MDS) was shown to have associated Tn syndrome; the first report of Tn syndrome with MDS. The Tn expression was demonstrated on erythrocytes, granulocytes, monocytes, platelets, and lymphocytes by flow cytometric analysis using a lectin and an antibody. Electrophoresis of erythrocyte membrane proteins revealed slower mobility of glycophorin B from the patient than that from normal individuals, suggesting a glycophorin B molecular abnormality.

Antigens, Tumor-Associated, Carbohydrate↗
PubMed1992

Expression of the Tn antigen on erythroid cells from a patient with Tn syndrome.

In order to examine expression of the Tn antigen on erythroid cells from a patient with Tn syndrome, we applied a selective two phase liquid culture system for human erythroid progenitors in peripheral blood. The cells were analyzed with flow cytometry employing an anti-Tn antibody and a lectin of Vicia villosa which recognizes only the Tn determinant. In the second phase, the Tn antigen was expressed on the cultured cells from the patient on day 3 and Tn-positive cells reached 62.7% on day 9. On the other hand, Tn-positive cells were not detected in the volunteer's cultured cells. When the patient's cells were co-cultured with the cells from a healthy volunteer, the percentage of Tn-positive cells was much lower than the expected value, suggesting that the normal cells suppressed the expression of Tn antigen on the patient's cells.

Acetylgalactosamine↗
PubMed1992

Characterization of a 28-kD polymorphic polypeptide detected by two-dimensional electrophoresis of human platelets.

A genetic polymorphism of a human platelet polypeptide with a molecular weight of 28 kD detected by two-dimensional electrophoresis was investigated in family and population studies, and cell distribution. The 28-kD polypeptide showed autosomal codominant inheritance of two alleles. The gene frequencies of the two alleles were 0.925 and 0.075, respectively. The 28-kD polypeptide was observed in lymphocytes, neutrophils, eosinophils and monocytes, in addition to platelets. This polypeptide showed good reproducibility in electrophoresis, and appears to be useful as a genetic marker of the human genome in gene mapping and pedigree analysis.

Blood Platelets↗
PubMed1991

Method for detecting the lectin activity of Momordica charantia transferred from micro two-dimensional electrophoretic gel on to nitrocellulose.

A method was devised for detecting both the molecular mass and the isoelectric point (pI) of the lectin in the seed extract of Momordica charantia on a nitrocellulose membrane. It was associated with the electrophoretic blotting technique that produced replicas of proteins separated on micro two-dimensional polyacrylamide gels. The red blood cell adherence procedure on the blotted membrane exhibited only one red spot with molecular mass 107.10(3) and pI 5.3, which indicated the lectin activity. Additionally, the lectin appeared to be a glycoprotein with mannose and/or glucose, because it was stained by concanavalin A-peroxidase staining.

Animals↗
PubMed1991

Characterization of autoantibodies in mixed-type autoimmune hemolytic anemia.

Three of 46 patients with autoimmune hemolytic anemia (AIHA) satisfied the diagnostic criteria for mixed-type AIHA in which both warm-type and cold-type autoantibodies against red blood cells (RBCs) are present. The specificities of these autoantibodies were analyzed. All of the warm-type autoantibodies were IgG-chi, and the specificities were not serologically classified. The autoantibody of patient 1 reacted to the 41- and 80-kD peptides on immunoblotting, and the epitope corresponding to it was papain sensitive. Two warm-type autoantibodies from patients 2 and 3 resembled each other in serological analyses and reacted with a protease- and neuraminidase-resistant antigen. However, the antigen corresponding to the autoantibody of patient 3 was located on the 37-kD peptide by immunoprecipitation. All of the cold-type autoantibodies were IgM-cha and showed high titer and high thermal amplitude. According to the reaction pattern with untreated and enzyme-treated RBCs, the cold-type autoantibodies of patients 2 and 3 were revealed to be anti-Om and anti-I, respectively. In patient 1, the cold-type autoantibody was characterized as having high affinity for autologous RBCs, but its specificity was unclassified. The antigens corresponding to the cold-type autoantibodies were not located by immunoblotting and immunoprecipitation. These serological and immunochemical approaches to autoantibodies in mixed-type AIHA revealed that the warm and cold components recognized the different antigens.

Aged↗