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T Okazaki

Publications and source records attributed to T Okazaki.

At least 19 recordsLinked to original sources
PubMed1992

Cloning and nucleotide sequence of the Pseudomonas aeruginosa nfxB gene, conferring resistance to new quinolones.

The gene nfxB is one of the genes which affect the cell membrane permeability of quinolones in Pseudomonas aeruginosa PAO. Both wild-type nfxB and a mutant nfxB (nfx13E) were cloned and the DNA sequences were determined. The wild-type gene was dominant in PAO strains. The nfxB mutation was a point mutation (cytosine----guanine) which generates an amino acid exchange (arginine----glycine) in the putative nfxB product. The amino acid sequence of the wild-type NfxB protein revealed that it has a helix-turn-helix motif which may be responsible for the ability to bind in a sequence-specific manner to DNA. This finding indicated that the NfxB protein may regulate the expression of genes that are associated with cell permeability of drugs in P. aeruginosa. The position of the amino acid substitution between the NfxB protein and the Nfx13E protein was located within a possible DNA-binding domain, suggesting that the mutant protein (Nfx13E) may have lost DNA binding ability and regulator activity.

Amino Acid Sequence↗
PubMed1992

Expression of the Qa-2k phenotype encoded by the Q5k gene on the surface of tumor cells derived from H-2k mice.

Immunological and biochemical characteristics of the Qa-2 murine nonclassical histocompatibility class 1 antigen expressed on tumor cells derived from H-2k (Qa-2-) mice were studied. It was found that the Qa-2 antigen on normal H-2b lymphocytes reacted with Qa-2-specific monoclonal antibodies (mAbs) 34-1.2, 59 (both specific to the alpha 1/alpha 2 region) and 141-15.8 (specific to the alpha 3 domain), and the Qa-2 antigen on H-2k tumor cells (Qa-2k antigen) reacted with mAbs 59 and 141-15.8, but not with 34-1.2. The normal Qa-2 antigen was susceptible to treatment with phosphatidylinositol-specific phospholipase C, but the Qa-2k antigen was insensitive to it. By Northern hybridization, polymerase chain reaction (PCR) studies on cDNA, Southern hybridization, Western blotting, and nucleotide sequence analysis, the Q5k gene was identified as the gene encoding the Qa-2k antigen expressed on BW5147 lymphoma cells derived from a mouse of AKR strain (H-2k, Qa-2-). The nucleotide sequence of PCR-amplified BW5147 Q5k cDNA showed complete agreement with the reported sequence of exons 1-5 of the Q5k gene of C3H/He. It also showed complete deletion of the region corresponding to exons 6 and 7, and a very short coding region in exon 8, resulting in very short cytoplasmic domain of the product compared with regular class 1 antigens. These characteristics were expected from the reported Q5k genomic sequence. These results revealed that the Qa-2k antigen was distinct from the normal Qa-2 antigen expressed on H-2b lymphocytes although it cross-reacted with some Qa-2-specific mAbs.

Amino Acid Sequence↗
PubMed1992

Detection of allogeneic Qa/TL and Ly specificities on murine tumor cells with IgD in tumor-regressor serum.

Serum from C3H/He mice, which show regression of MM2 tumor cells after transplantation and removal (regressor serum, RS) contains non-gammaglobulin components that cross-react with various tumor cells of mice [22, 23]. In addition to tumor cells, various allogeneic lymphocytes are also susceptible to an RS-dependent lymphocyte-mediated cytotoxic reaction. To identify tumor cell surface antigens that cause the cross-reactive host response, the serum components were analyzed by absorption of RS with allogeneic lymphocytes. RS components were found to recognize allogeneic lymphocyte antigens including Qa-2 and Ly6.2. Specificity for the Qa-2 antigen was further tested using Qa-2-congenic mice. The expression of Qa-2 antigen was detected on the surfaces of MM2 and other tumor cells derived from H-2k mice (seven among nine cell lines tested) by a membrane immunofluorescence method using a Qa-2-specific mAb. Physical characteristics of the Qa-2-specific component in RS were determined and found to differ from those of regular IgGs but to be similar to those of IgDs. Using an enzyme-linked immunosorbent assay with an IgD-specific mAb and Qa-2-lacZ fusion protein, the existence of IgD in RS with specificity for Qa-2 was confirmed. These results suggest that the RS component with Qa-2 specificity is an IgD, the specificity and physiological role of which are unknown.

Animals↗
PubMed1992

Correlation between extrinsic nerve fibers and synapses in the muscle layers of bowels affected by Hirschsprung's disease.

The aim of this study is to clarify any correlation between nerve terminals (synapses) and proliferating extrinsic nerve fibers in the muscle layers of bowels affected by Hirschsprung's disease (HD). Synapses and extrinsic nerve fibers in the muscle layers of bowels of 10 patients with HD and 8 comparable controls were labeled with monoclonal antibody (MAb) 171B5 and acetylcholinesterase (AChE), respectively. In the control a rich and even distribution of synapses, representing neuromuscular junctions, was seen in the muscle layers, together with dense clusters of synapses in the adjacent myenteric plexuses; proliferation of extrinsic nerve fibers was not seen. In the transitional oligoganglionic segment of HD, many synapses were present in the myenteric plexus, but a few synapses in the muscle layers; there was a gradual increase of extrinsic nerve fibers from proximal to distal. In the narrow aganglionic segment very few synapses were seen in the muscle layers; proliferating nerve fibers and bundles were prominent. We conclude that the muscle layers of bowels affected by HD were almost denervated despite presence of intrinsic nervous elements in the oligoganglionic segment and proliferating extrinsic nerve fibers in the aganglionic segment.

Hirschsprung Disease↗
PubMed1992

Phenotypic and genotypic characterization of 14 leukemia and lymphoma cell lines with 11q23 translocations.

11q23 translocation is the most popular chromosomal abnormality in infant leukemia. In adults, it is often encountered in non-Hodgkin's lymphoma (NHL). In this study, we analyzed the phenotypic and genotypic characteristics of 9 acute leukemic cell lines with 11q23 translocations and one with deletion of the 11q23 locus, nine of which were established by researchers in this group, together with 4 NHL cell lines with 11q23 translocations. All lines were considered to belong to the B-cell lineage at different stages. All 10 leukemic lines showed clonal rearrangement of the immunoglobulin heavy chain (IgH) gene: two corresponded to the B-precursor stage (CD19+, cytoplasmic mu-), while the other 8 corresponded to the pre-B stage (cytoplasmic mu+). All 4 NHL lines showed rearrangements of both the IgH and Ig kappa genes with three expressing surface Ig; specifically, mature B-cell phenotype. As for myelocytic-monocytic markers, at least one out of 4 antigens examined were positive in 8 of the 10 leukemic cell lines, while only one of the 4 NHL lines was reactive. There were essentially no clear phenotypic or genotypic differences between t(4;11) and t(11;19) cell lines, supporting the view that both diseases have similar clinicopathological characteristics. These cell lines are also valuable for cloning genes at the chromosomal breakpoints.

Acute Disease↗
PubMed1992

Transcriptional activity of a mutant thyroid hormone receptor beta in a family with generalized resistance to thyroid hormone.

We previously reported a family with generalized resistance to thyroid hormone (GRTH) which had a point mutation with codon 448 CCT (proline) being converted to ACT (threonine) in the thyroid hormone receptor (TR) beta. To characterize functional properties of the mutant TR beta, transient expression studies were performed in COS cells. A double stranded oligonucleotide encompassing thyroid hormone response element (TRE) derived from the rat GH gene was synthesized. We constructed chloramphenicol acetyl transferase (CAT) plasmid containing the thymidine kinase promoter under the control of the rat GH TRE. T3 induction of CAT activity by the mutant TR beta was significantly reduced as compared with that of the normal TR beta. This was observed in the presence of 0.5-50 nM T3, but not at 500 nM T3. When the normal and mutant TR beta were cotransfected, the mutant TR beta inhibited gene activation regulated by the normal TR beta. However, a high molar excess was necessary to significantly inhibit the function of the normal receptor. Additionally, the binding of in vitro synthesized mutant TR beta to TRE was preserved.

Animals↗
PubMed1992

Centromere protein B assembles human centromeric alpha-satellite DNA at the 17-bp sequence, CENP-B box.

We purified 15,000-fold from HeLa cell nuclear extract the centromere antigen that reacts specifically with the 17-bp sequence, designated previously as CENP-B box, in human centromeric alpha-satellite (alphoid) DNA by a two-step procedure including an oligonucleotide affinity column. The purified protein was identified as the centromere protein B (CENP-B) by its mobility on SDS-PAGE (80 kD), and reactivities to a monoclonal antibody raised to CENP-B (bacterial fusion protein) and to anticentromere sera from patients with autoimmune diseases. Direct binding by CENP-B of the CENP-B box sequence in the alphoid DNA has been proved using the purified CENP-B by DNA mobility-shift assay, Southwestern blotting, and DNase I protection analysis. The binding constant of the antigen to the CENP-B box sequence is 6 x 10(8) M-1. DNA mobility-shift assays indicated that the major complex formed between the CENP-B and the DNA contains two DNA molecules, suggesting the importance of the CENP-B/CENP-B box interaction in organization of higher ordered chromatin structures in the centromere and/or kinetochore. Location of DNA binding and dimerization domains in CENP-B was discussed based on the DNA mobility-shift assays performed with a protein fraction containing intact and partial cleavage products of CENP-B.

Autoantigens↗
PubMed1992

A human centromere protein, CENP-B, has a DNA binding domain containing four potential alpha helices at the NH2 terminus, which is separable from dimerizing activity.

The alphoid DNA-CENP-B (centromere protein B) complex is the first sequence-specific DNA/protein complex detected in the centromeric region of human chromosomes. In the reaction, CENP-B recognizes a 17-bp sequence (CENP-B box) and assembles two alphoid DNA molecules into a complex, which is designated complex A (Muro, Y., H. Masumoto, K. Yoda, N. Nozaki, M. Ohashi, and T. Okazaki. 1992. J. Cell Biol. 116:585-596). Since CENP-B gene is conserved in mammalian species and CENP-B boxes are found also in mouse centromere satellite DNA (minor satellite), this sequence-specific DNA-protein interaction may be important for some kind of common centromere function. In this study we have characterized the structure of CENP-B and CENP-B-alphoid DNA complex. We have shown by chemical cross-linking that CENP-B formed a dimer, and have estimated by molecular weight determination the composition of complex A to be a CENP-B dimer and two molecules of alphoid DNA. The DNA binding domain has been delimited within the NH2-terminal 125-amino acid region containing four potential alpha-helices using truncated CENP-B made in Escherichia coli cells. We have shown that CENP-B had sites highly sensitive to proteases and that the DNA binding domain was separable from the dimerizing activity by the proteolytic cleavage at 20 kD from the COOH terminus of the molecule. Thus, CENP-B may organize a higher order structure in the centromere by juxtaposing two CENP-B boxes in the alphoid DNA repeat through both the DNA-protein and protein-protein interactions.

Amino Acid Sequence↗
PubMed1992

The role of the seminal vesicles and coagulating glands in fertilization in the rat.

To investigate the role of the seminal vesicles (SV) and coagulating glands (CG) in fertilization in the rat, partial resections of the SV (25%--group B), 50%--group C, and 75%--group D) or bilateral resection of the SV alone (100%--group E), or together with the CG (group F) were undertaken. In other groups, bilateral resection of the CG only (group G), or bilateral resection of the CG with ipsilateral SV resection (group H) were performed. A group of sham-operated rats served as controls (group A). There were no significant differences among groups A, B, C and D in the number of fertile rats post-operatively but significant differences were evident in groups E, F, G, H when compared with the control group (P less than 0.01 for all groups). Only the fertile rats were able to form copulatory plugs (CP) post-operatively. Bilateral ligation of the SV ducts was performed in ten fertile rats. Post-operatively, the proportion of fertile rats was decreased significantly (P less than 0.01). Removal of the ligatures resulted in a significant increase (P less than 0.01) in fertility. It was also demonstrated that the addition of rat seminal vesicular secretions (SVS) to epididymal sperm suspensions resulted in a significant decrease (P less than 0.001) in sperm motility. These results suggest that the SV and CG are necessary for fertility in rats, and that one role of SVS is the formation of a CP and not the maintenance of sperm motility.

Animals↗
PubMed1992

Surgical repair versus medical treatment of varicocele in the rat: pharmacological manipulation of the varicocelized testicle.

Human chorionic gonadotropin, kallikrein, indomethacin, and hydralazine were administered to different groups of varicocelized rats, while surgical repair of the varicocele was performed in another group of rats. The effects of conservative and surgical treatment on epididymal sperm content and motility, the weights of the testes, epididymis, and male accessory genital glands, and fertility were compared between each group and a sham-treated group of rats. Surgical repair significantly improved all the evaluated parameters and all the conservative regimens, except hydralazine, resulted in a significant improvement in most parameters. Our results indicate that stimulation of the Leydig or/and Sertoli cells of a varicocelized testicle can counteract some of the detrimental consequences of the varicocele itself.

Animals↗
PubMed1992

Conserved mechanism of negative gene regulation by extracellular calcium. Parathyroid hormone gene versus atrial natriuretic polypeptide gene.

We found that a negative calcium-responsive element (nCaRE) originally reported in the human parathyroid hormone gene is conserved among several genes. The results of the present study show that expression of one of the genes, the rat atrial natriuretic polypeptide gene, was negatively regulated in the heart by extracellular calcium by using an in vivo infusion system. Moreover, transfection of the cultured cells revealed that this DNA element conferred negative regulation by extracellular calcium on the reporter gene. It is suggested that there is a gene family whose expression is negatively regulated by extracellular calcium through this conserved DNA motif, nCaRE.

Animals↗
PubMed1992

A point mutation in the 3,5,3'-triiodothyronine-binding domain of thyroid hormone receptor-beta associated with a family with generalized resistance to thyroid hormone.

A tight linkage between generalized resistance to thyroid hormone (GRTH) and the thyroid hormone receptor-beta (TR beta) gene is indicated. We evaluated a family with GRTH for the TR beta gene. We found that a new point mutation, consisting of a cytosine to adenine replacement at nucleotide position 1642, resulted in substitution in codon 448 in the T3-binding domain of TR beta. This base substitution was found in only one allele of affected members, but not in unaffected members of the family. The in vitro translation products of this mutant TR beta gene demonstrated significantly reduced T3-binding affinity. Previously, others have reported a kindred with GRTH, in that the same codon was subjected to proline to histidine replacement due to a mutation consisting of a cytosine to adenine replacement at nucleotide position 1643. There appeared to be a significant phenotypic difference between our kindred and that described by others.

Alleles↗
PubMed1992

Matlystatins, new inhibitors of typeIV collagenases from Actinomadura atramentaria. I. Taxonomy, fermentation, isolation, and physico-chemical properties of matlystatin-group compounds.

During the course of a screening for inhibitors of typeIV collagenases, new metabolites, designated matlystatins, have been isolated from an actinomycete strain, which was identified as a strain of Actinomadura atramentaria. Matlystatins were composed of five congeners, which were separated and purified by n-butanol extraction and chromatography.

Acetylcysteine↗
PubMed1992

[Clinical and cytological features of CD7 positive biphenotypic leukemias].

Clinical and cytological features of CD7 positive acute leukemias with biphenotypic characteristics in childhood were documented. From 87 patients with CD7+ acute leukemias, nine patients were selected on the basis of the biphenotypic expression of T-lymphoid and myelomonocytic antigens. The blasts of these patients expressed cell surface CD7, cytoplasmic CD3 and cytoplasmic CD13. In addition to these antigens, surface CD13 was also expressed after short term culture without any mitogens or stimulators. The double PAP method for detecting cytoplasmic antigens (CD3, CD13, beta F1, delta TCS1) was employed in this study. The average age of these patients was higher (10.2 y/o) than patients with common ALL. Mediastinal masses were observed in 4 of 9 patients. They were treated according to the diagnoses based on conventional hematological methods and surface antigen expression. In all 9 patients, complete remission was achieved, however, early relapse was noticed in 7. This study suggests that the leukemia cells of these patients may be derived at an early stage during the differentiation of multipotential hematopoietic stem cells. New therapeutic approaches are necessary to improve the outcome of such T/M biphenotypic leukemias.

Acute Disease↗
PubMed1992

[Urinary 11-dehydrothromboxane B2 and 6-keto-prostaglandin F1 alpha in normal pregnant women and in women complicated with pregnancy-induced hypertension].

In normal pregnancy, increased production of platelet thromboxane A2(TXA2) parallels increased biosynthesis of vascular prostacyclin (PGI2). An imbalance in the formation of these prostaglandins is believed to be associated with the pathogenesis of pregnancy-induced hypertension (PIH). Recent evidence suggested that aspirin in low doses was effective in reducing the incidence of PIH, by selective inhibition of platelet-derived TXA2 biosynthesis. In this communication, we determined the urinary 11-dehydro TXB2 and 6-keto-PGF1 alpha, which are major metabolites of TXA2 and PGI2, respectively, from early to late pregnancy of normal pregnant women and of women complicated with PIH. The ratio of 11-dehydro TXB2 to 6-keto-PGF1 alpha decreased significantly from as early as 10wks of gestation when compared with that in non-pregnant controls (1.43 +/- 0.15 vs 1.99 +/- 0.13: Mean +/- SEM, p less than 0.05), and increased in later pregnancy to the control values at term. No significant difference was found in the excretion of 11-dehydro TXB2 between normal pregnant women and women with PIH. In contrast, urinary excretion of 6-keto-PGF1 alpha decreased in women with PIH. The ratio of 11-dehydro TXB2 to 6-keto-PGF1 alpha increased significantly as compared with that of pregnant controls. These results demonstrated that disturbed production of vascular PGI2 may be the primary cause of PIH, and affect the vascular responsiveness to pressor inducers such as angiotensin II.

6-Ketoprostaglandin F1 alpha↗
PubMed1992

[Correction of haptoglobin contents in hemolytic specimen with addition of saturated hemoglobin].

Measurement of serum haptoglobin by immunoturbidimetry was markedly interfered with hemolysis because of haptoglobin-hemoglobin complex formation. Turbidity formed with haptoglobin and immunoglobulin complexes was gradually decreased by the addition of hemoglobin in the reaction mixture, and reached to a certain fixed levels when serum haptoglobin were saturated with hemoglobin. The turbidity derived from saturated haptoglobin and hemoglobin complex closely correlated with the haptoglobin level without hemoglobin. Thus, we could measure the corrected haptoglobin level in hemolytic specimen by the saturation of hemoglobin in reaction mixture.

Haptoglobins↗
PubMed1992

Serum erythropoietin concentrations and iron status in patients on chronic hemodialysis.

We studied the relationship between serum erythropoietin (EPO) concentration and iron status in 67 patients undergoing chronic hemodialysis. Serum concentrations of EPO were measured by RIA with recombinant human EPO. The geometric mean of the serum EPO concentration was 10.9 int. units/L (mean +/- SD range = 7.8 - 15.3 int. units/L) in hemodialysis patients, considerably lower than that in normal subjects (12.9 int. units/L). We found no significant correlation between concentrations of serum EPO and hemoglobin in hemodialysis patients, but found a significant negative correlation between serum concentrations of EPO and iron in hemodialysis patients. Moreover, we also found a significant positive correlation between the EPO concentration and the unsaturated iron-binding capacity (UIBC) in serum, and a significant negative correlation between the serum concentrations of EPO and ferritin in hemodialysis patients. Several patients who had relatively high EPO concentrations for hemodialysis patients also had low iron concentrations, high UIBC values, and low ferritin concentrations. These findings suggest that iron was utilized even at these EPO concentrations, which were very low for the degree of anemia observed in the hemodialysis patients.

Adult↗