Biomedical subjects

T Kudo

Publications and source records attributed to T Kudo.

At least 181 records · Page 10Linked to original sources

PubMed1999

Cloning and expression of a human gene encoding an N-acetylgalactosamine-alpha2,6-sialyltransferase (ST6GalNAc I): a candidate for synthesis of cancer-associated sialyl-Tn antigens.

The sialyl-Tn (sTn) antigen is a well known cancer-associated antigen, the expression of which is related to the prognosis of cancer patients. We aimed to isolate a human gene encoding an N -acetylgalactosamine alpha2,6-sialyltransferase which synthesizes sTn antigen, and to characterize the enzyme. Degenerate primers encoding sialyl motifs were used for the polymerase chain reaction to amplify complementary DNAs prepared from RNAs of human pyloric mucosae with intestinal metaplasia, which abundantly expressed sTn antigen, followed by screening of full-length cDNAs using the amplified DNA fragment as a probe. We isolated two human cDNA clones, long-form (2.46 kb) and short-form (2.23 kb) cDNAs. The former encodes an active enzyme with a predicted 600 amino acid sequence. The latter, a splice-variant of the long-form, encodes an inactive enzyme. HCT15 human colorectal cancer cells stably expressing the long-form cDNA expressed sTn epitopes on O -glycans. The long form cDNA was considered to encode a human homologue of chick ST6GalNAc I for the following reasons: (1) the putative amino acid sequence showed greater homology to that of chick ST6GalNAc I (55%) compared to other sialyltransferases, (2) it encodes the extraordinarily long stem region that is a typical feature of chick ST6GalNAc I, and (3) the substrate specificity was very similar to that of chick ST6GalNAc I. In situ hybridization demonstrated that the localization of transcripts correlated well with that of sTn antigen in gastric cancer cells and Goblet cells in intestinal metaplastic glands. Thus, we determined that the long-form cDNA of the human ST6GalNAc I gene encodes the probable candidate for the human sTn synthase(s).

Acetylglucosamine↗

PubMed1999

Molecular behavior of mutant Lewis enzymes in vivo.

The expression of type-1 Lewis antigens on erythrocytes and in digestive organs is determined by a Lewis type alpha(1,3/1, 4)-fucosyltransferase (Lewis enzyme) encoded by the Fuc-TIII gene ( FUT3 gene; Lewis gene). We have classified the Lewis alleles in the Japanese population into four types, the wild-type allele ( Le ) and three mutated alleles, i.e., le1, which has missense mutations T59G and G508A, le2, which has T59G and T1067A, and le3, which has only T59G. Here we carried out an extensive study on the biological properties of the three mutant Lewis enzymes, the le1, le2, and le3 enzymes, using native tissues and obtained the following results. (1) In in vivo and in vitro experiments, the le1 and le2 enzymes were found to be susceptible to protease digestion probably because the one missense mutation in the catalytic domains, i.e., Gly170 to Ser in the le1 enzyme and Ile356 to Lys in the le2 enzyme, makes the three-dimensional structures of the enzymesunstable, while the le3 and wild-type Lewis enzymes wereresistant to protease digestion. (2) The le1 and le2 enzymes cannot synthesize type 1 Lewis antigens on either glycolipids or mucins. The le3 enzyme cannot synthesize Lewis-active glycolipids, which result in the Lewis antigen-negative phenotype of erythrocytes, while it can synthesize Lewis antigens on mucins in normal and cancerous colon tissues. The missense mutation, Leu20 to Arg, in the transmembrane domain reduces retention of the le3 enzyme in the Golgi membrane resulting in an apparent reduction of enzyme activity as revealed by the lack of Lewis antigen synthesis. (3) The Lewis gene dosage actually has effects in vivo on the amount of the Lewis enzyme, its activity, and finally the amounts of Lewis carbohydrate antigens. This is the first article that clearly demonstrates the gene dosage effects on the amount of the glycosyltransferase protein, its activity, and the amounts of carbohydrate products in vivo.

Alleles↗

PubMed1999

Molecular mechanisms of expression of Lewis b antigen and other type I Lewis antigens in human colorectal cancer.

Lewis b (Leb) antigens are gradiently expressed from the proximal to the distal colon, i.e., they are abundantly expressed in the proximal colon, but only faintly in the distal colon. In the distal colon, they begin to increase at the adenoma stage of cancer development and then increase with cancer progression. We aimed to clarify the molecular basis of Leb antigen expression in correlation with the expression of other type I Lewis antigens, such as Lewis a (Lea) and sialylated Lewis a (sLea), in colon cancer cells. Considering the Se genotype and the relative activities of the H and Se enzymes, the amounts of Leb antigens were proved to be determined by both the H and Se enzymes in noncancerous and cancerous colon tissues. But the Se enzyme made a much greater contribution to determining the Lebamounts than the H enzyme. In noncancerous colons, the Se enzyme were gradiently expressed in good correlation with the Leb expression, while the H enzyme was constantly expressed throughout the whole colon. In distal colon cancers, the H and Se enzymes were both significantly upregulated in comparison with in adjacent noncancerous tissues. In proximal colon cancers, expression of the H enzyme alone was highly augmented. The augmented expression of Leb antigens in distal colon cancers is caused mainly by upregulation of the Se enzyme and partly by the H enzymes, while it is caused by upregulation of the H enzyme alone in proximal colon cancers. The Se gene dosage profoundly influences the amounts of the Leb, Lea, and sLea antigens in whole colon tissues, regardless of whether they are noncancerous or cancerous tissues. It suggests that the Se enzyme competes with alpha2,3 sialyltransferase(s) and the Le enzyme for the type I acceptor substrates.

Antibodies, Monoclonal↗

PubMed1999

Induction of macrophage migration inhibitory factor in human ovary by human chorionic gonadotrophin.

The role of macrophage migration inhibitory factor (MIF) in human ovarian function remains obscure. The aim of this study was to investigate how MIF was related to ovulation by quantitative analysis of serum, follicular fluid and culture medium of granulosa cells obtained from in-vitro fertilization (IVF) and embryo transfer patients. Serum MIF concentrations in ovarian stimulation cycles for IVF-embryo transfer were higher at day 1 (median 92.6 ng/ml), which took place 35 h after human chorionic gonadotrophin (HCG) administration and just before the retrieval of oocytes, than those before day -6 (12.1 ng/ml), at day -5 to about day 0 (17.5 ng/ml) or at day 2 to about day 14 (8.2 ng/ml). MIF concentrations in the follicular fluid (113.4 ng/ml) obtained in ovarian stimulation cycles for IVF-embryo transfer were significantly higher than in serum (72.0 ng/ml) collected at the same time. MIF concentrations in the follicular fluid in natural cycles were higher in the ovulatory phase (51.6 ng/ml) than in the late follicular phase (13.8 ng/ml). MIF concentrations in the culture media of granulosa cells increased from 3.2 ng/ml to 7.2 ng/ml with HCG stimulation, and decreased from 2.4 ng/ml to 1.2 ng/ml when stimulation was withheld. These results indicate that HCG can induce the elevation of serum and follicular fluid MIF concentrations through the stimulation of ovarian cells, and that MIF is probably involved in the mechanism of ovulation.

Adult↗

PubMed1999

Generation of peroxynitrite and apoptosis in placenta of patients with chorioamnionitis: possible implications in placental abruption.

The reaction of nitric oxide (NO) and superoxide results in the formation of peroxynitrite, a potent and relatively long-lived oxidant. In infectious diseases, these molecules are not only bactericidal but also toxic to host cells. Chorioamnionitis is often complicated by premature rupture of membranes and can be associated with placental abruption. These diseases are significant causes of premature low-birth-weight deliveries and consequently the morbidity and mortality of neonates. Lipopolysaccharide, bacterial endotoxin, is known to be elevated in the amniotic fluid of patients with chorioamnionitis. Lipopolysaccharide is known to induce the formation of NO and superoxide. We report here that nitrite/nitrate, stable metabolites of NO, were increased in serum from patients with chorioamnionitis. Immunohistochemical studies demonstrated enhanced expression of inducible NO synthase and formation of nitrotyrosine, a footprint of peroxynitrite, in the placentae from patients with chorioamnionitis and also in patients with placental abruption. Furthermore, apoptotic cell death was also increased in the placentae from patients with both diseases. These results suggest that chorioamnionitis and a portion of placental abruption may share a common cascade of placental injury. Nitric oxide and its metabolities may play an important role in this cascade.

Adult↗

PubMed1999

Classification of 'Nocardioides fulvus' IFO 14399 and Nocardioides sp. ATCC 39419 in Kribbella gen. nov., as Kribbella flavida sp. nov. and Kribbella sandramycini sp. nov.

By 16S rDNA sequence analysis, two strains identified previously as members of the genus Nocardioides, namely 'Nocardioides fulvus' IFO 14399 and Nocardioides sp. ATCC 39419, were found to form lines of descent distinct from the genus Nocardioides. Therefore, the two strains were studied taxonomically in detail by phenotypic, particularly chemotaxonomic, characterization, and by phylogenetic analysis including other related taxa. The genomic DNA G + C contents of 'N. fulvus' IFO 14399 and Nocardioides sp. ATCC 39419 are 70 and 68 mol%, respectively. The two strains contain LL-diaminopimelic acid as the diamino acid in their peptidoglycan, together with L-alanine at position 1 of the peptide subunit. The predominant menaquinone found in the two strains is MK-9(H4). The major cellular fatty acid found in the two strains is anteiso-C15:0' which is a minor component of members of the genus Nocardioides. The diagnostic polar lipid of the two strains is phosphatidylcholine. A phylogenetic tree was constructed which showed that the two strains from evolutionary lineages distinct from other LL-diaminopimelic-acid-containing taxa such as, for example, the genus Nocardioides. On the basis of our data, a new genus, Kribbella gen. nov., and two new species, Kribbella flavida gen. nov., sp. nov. and Kribbellla sandramycini gen. nov., sp. nov., are proposed. It is also proposed that the new genus be included in the family Nocardioidaceae.

Actinomycetales↗

PubMed1999

Microbispora corallina sp. nov., a new species of the genus Microbispora isolated from Thai soil.

Two actinomycete strains, DF-28 and DF-32T, were isolated from soil samples collected in a deciduous dipterocarp forest in Thailand. They produced longitudinally paired spores on the tips of short sporophores alternately branched from aerial hyphae, and the chemotaxonomic properties of the isolates were the same as those of members of the family Streptosporangiaceae. These phenotypic properties, together with the results of a phylogenetic analysis based on 16S rRNA gene sequences, indicated that these isolates should be assigned to the genus Microbispora. The two isolates showed more than 93% DNA relatedness to each other, but their relatedness to any previously described species of the genus Microbispora was only 45% or less. They were distinguishable from previously described Microbispora spp. by a combination of physiological and biochemical properties. Therefore, a new species is proposed for these strains, under the name Microbispora corallina sp. nov. The type strain is strain DF-32T (= JCM 10267T).

Actinomycetales↗

PubMed1999

Catenuloplanes crispus (Petrolini et al. 1993) comb. nov.: incorporation of the genus Planopolyspora Petrolini 1993 into the genus Catenuloplanes Yokota et al. 1993 with an amended description of the genus Catenuloplanes.

The taxonomic position of the genus Planopolyspora comprising a single species, Planopolyspora crispa, was reviewed. This genus was originally characterized by formation of long, curly and sometimes branching sporangia containing numerous zoospores arranged in a single row and by the presence of meso-diaminopimelic acid and madurose (3-O-methyl-D-galactose) in whole-cell hydrolysates. However, our chemotaxonomic analyses of the type strain of P. crispa did not agree with the original description. The peptidoglycan contained L-lysine but not meso-diaminopimelic acid, and the whole-cell hydrolysate contained xylose as the characteristic sugar but not madurose. These characteristics and other chemotaxonomic profiles (e.g. menaquinone, phospholipid and cellular fatty acid compositions) of the genus Planopolyspora coincided with those of the genus Catenuloplanes. These two genera also had very similar morphological characteristics, but in the original description of the genus Catenuloplanes the presence of sporangia was not referred to. This difference is considered to originate from a divergence of views owing to the ambiguity of the definition of the term 'sporangium' in actinomycete morphology. Phylogenetic analysis based on 16S rDNA sequences also supported the proposal that the genera Planopolyspora and Catenuloplanes should be combined into one genus. Levels of DNA relatedness among the type strains of P. crispa and six Catenuloplanes species and their cultural, physiological and biochemical characteristics indicated that P. crispa should be classified as an independent species of the genus Catenuloplanes, which has priority over the genus Planopolyspora. Therefore, it is proposed that Planopolyspora crispa be transferred to the genus Catenuloplanes as Catenuloplanes crispus comb. nov.

Bacterial Typing Techniques↗

PubMed1999

Genetic organization and characteristics of the 3-(3-hydroxyphenyl)propionic acid degradation pathway of Comamonas testosteroni TA441.

Comamonas testosteroni TA441 degrades 3-(3-hydroxyphenyl)propionate (3HPP) via the meta pathway. A gene cluster required for degradation of 3HPP was cloned from strain TA441 and sequenced. The genes encoding six catabolic enzymes, a flavin-type hydroxylase (mhpA), extradiol dioxygenase (mhpB), 2-keto-4-pentenoate hydratase (mhpD), acetaldehyde dehydrogenase (acylating) (mhpF), 4-hydroxy-2-ketovalerate aldolase (mhpE) and the meta cleavage compound hydrolase (mhpC), were found in this cluster, encoded in this order. mhpD and mhpF were separated by two genes, orf4 and orf5, which were not necessary for growth on 3HPP. The gene mhpR, encoding a putative transcriptional activator of the IcIR family, was located adjacent to mhpA in the opposite orientation. Disruption of the mhpB or mhpR genes affected growth on 3HPP or trans-3-hydroxycinnamate. The mhpB and mhpC gene products showed high specificity for 3-(2,3-dihydroxyphenyl)propionate (DHPP) and the meta cleavage compound produced from DHPP, respectively.

Aldehyde Oxidoreductases↗

PubMed1999

Crystallization and preliminary x-ray diffraction studies of hyperthermostable glutamate dehydrogenase from Thermococcus profundus.

Recombinant glutamate dehydrogenase from a hyperthermophilic archaeon, Thermococcus profundus, was crystallized in the presence of both polyethylene glycol 8000 and lithium sulfate. Four types of crystals having different morphologies appeared in the crystallization trials; however, only one type was suitable for X-ray crystal structure analysis. The crystal belonged to the monoclinic space group P2(1) and the unit-cell parameters were a = 112.99, b = 163.70, c = 133.07 A, beta = 113.46 degrees at 110 K. The calculated V(M) value of 3.42 A(3) Da(-1) was acceptable when one hexamer of the enzyme, which was the physiological functional unit, occupied a crystallographic asymmetric unit. X-ray diffraction intensity data were collected to a resolution of 2.25 A with good statistics at the BL44B2 beamline of SPring-8.

Crystallization↗

PubMed1999

Receptor for hyaluronan-mediated motility and CD44 expressions in colon cancer assessed by quantitative analysis using real-time reverse transcriptase-polymerase chain reaction.

Receptor for hyaluronan (HA)-mediated motility (RHAMM) is a receptor for HA-mediated motility and its expression is correlated with malignancy of ras-transformed cells in that binding of HA to this receptor activates their migratory ability. CD44, a cell surface receptor for HA is also implicated in metastatic behavior of some cancer cells. In this study we examined the relationships of cancer progression with mRNA levels of RHAMM, CD44 (all forms), and exon 6 of CD44 using the real-time reverse transcriptase-polymerase chain reaction method in specimens of colon cancers at different diagnostic stages from 30 patients. Increased mRNA levels of RHAMM were observed in 29 specimens (97%), CD44s (all forms) in 21 specimens (70%), and its exon 6 in 19 specimens (63%) in comparison with those in the corresponding noncancerous tissue specimens. A statistically significant correlation between RHAMM expression and cancerous specimens at any of Dukes' stages A, B, and C was found, and the overexpression of CD44 mRNAs was confirmed in specimens at Dukes' stage C. Thus, our present study for the first time suggests that RHAMM expression may be a clinically useful indicator of colon cancer.

Adult↗

PubMed1999

Assessment of myometrial invasion at the invasion site of an endometrial carcinoma by ultrasonography along with an intrauterine catheter.

OBJECTIVES: To evaluate by transvaginal ultrasonography (TVU) the thickness of the intact myometrium at the presumed tumor-origin site and to establish criteria for a half myometrial invasion. METHODS: A total of 19 successive patients with endometrial cancer who were treated between January 1, 1997, and January 31, 1998, participated in this study. TVU mode B with and without the use of an intrauterine silicon catheter was performed. RESULTS: Using a catheter, the origin site was correctly detected in 15 cases (79%). The best criterion for half myometrial invasion was a 6-mm thickness of the intact myometrium at the origin site. The sensitivity/specificity/accuracy of TVU with the use of a catheter in cases with the correct estimated origin site, were 1.00/0.67/0.86 for myometrial invasion < 1/2, and 0.67/1.00/0.86 for myometrial invasion > or = 1/2. CONCLUSION: It is of value to use 6-mm as the criterion for the thickness of the intact myometrium at the estimated tumor-origin site in connection with TVU with the use of a catheter for preoperatively assessing half myometrial invasion.

Adult↗

PubMed1999

Distribution of beta-lactamases in actinomycetes.

The distribution of beta-lactamase activities in a collection of actinomycete strains was surveyed. Six of 127 strains were found to produce beta-lactamase. This low frequency was in contrast to the case with Streptomyces species. The producing strains were not related phylogenetically. MICs of benzylpenicillin did not correlate with beta-lactamase production.

Actinomycetales↗

PubMed1999

Phylogenetic diversity of nitrogen fixation genes in the symbiotic microbial community in the gut of diverse termites.

Nitrogen fixation by the microorganisms in the gut of termites is one of the crucial aspects of symbiosis, since termites usually thrive on a nitrogen-poor diet. The phylogenetic diversity of the nitrogen-fixing organisms within the symbiotic community in the guts of various termite species was investigated without culturing the resident microorganisms. A portion of the dinitrogenase reductase gene (nifH) was directly amplified from DNA extracted from the mixed population in the termite gut. Analysis of deduced amino acid sequences of the products of the clonally isolated nifH genes revealed the presence of diverse nifH sequences in most of the individual termite species, and their constituents were considerably different among termite species. A majority of the nifH sequences from six lower termites, which showed significant levels of nitrogen fixation activity, could be assigned to either the anaerobic nif group (consisting of clostridia and sulfur reducers) or the alternative nif methanogen group among the nifH phylogenetic groups. In the case of three higher termites, which showed only low levels of nitrogen fixation activity, a large number of the sequences were assigned to the most divergent nif group, probably functioning in some process other than nitrogen fixation and being derived from methanogenic archaea. The nifH groups detected were similar within each termite family but different among the termite families, suggesting an evolutionary trend reflecting the diazotrophic habitats in the symbiotic community. Within these phylogenetic groups, the sequences from the termites formed lineages distinct from those previously recognized in studies using classical microbiological techniques, and several sequence clusters unique to termites were found. The results indicate the presence of diverse potentially nitrogen-fixing microbial assemblages in the guts of termites, and the majority of them are as yet uncharacterized.

Animals↗

PubMed1999

Culture-independent characterization of a gene responsible for nitrogen fixation in the symbiotic microbial community in the gut of the termite Neotermes koshunensis.

Expression of the nitrogen fixation gene, nifH, in the gut of the termite Neotermes koshunensis was characterized without cultivation. nifH cDNA was directly amplified from mRNA of the mixed microbial population in the gut by reverse transcription (RT)-PCR. Analyses of the RT-PCR products revealed that, among the diverse nifH sequences, only a few corresponding to an alternative nitrogenase (encoded by the anf gene) were preferentially transcribed in the termite gut. Expression of the anf gene was further investigated quantitatively under several termite feeding conditions by competitive PCR. The levels of expression of the anf gene were largely congruent with the nitrogen fixation activity displayed by the termite. The amounts of the genomic anf gene in the population showed no significant change, indicating that the level of expression was critical for nitrogen fixation activity. Interestingly, no significant decrease in the expression level was observed when the diet contained molybdenum (Mo), which represses ordinary anf genes. A 3.6-kb DNA region downstream of the anf gene was isolated and found to contain reading frames homologous to anfH, anfD, and anfG of the Bacteria domain which encode subunits of an alternative nitrogenase having no Mo as a cofactor. This DNA region also contained reading frames encoding glnB-like proteins, which is a common feature of the nitrogenase genes of the Archaea domain. These results indicate that the anf group of nitrogenase genes is the most important group of genes responsible for nitrogen fixation in N. koshunensis and that the anf gene possesses novel features with respect to the regulation of its expression and its gene organization.

Animals↗

PubMed1999

Two distinct mechanisms cause heterogeneity of 16S rRNA.

To investigate the frequency of heterogeneity among the multiple 16S rRNA genes within a single microorganism, we determined directly the 120-bp nucleotide sequences containing the hypervariable alpha region of the 16S rRNA gene from 475 Streptomyces strains. Display of the direct sequencing patterns revealed the existence of 136 heterogeneous loci among a total of 33 strains. The heterogeneous loci were detected only in the stem region designated helix 10. All of the substitutions conserved the relevant secondary structure. The 33 strains were divided into two groups: one group, including 22 strains, had less than two heterogeneous bases; the other group, including 11 strains, had five or more heterogeneous bases. The two groups were different in their combinations of heterogeneous bases. The former mainly contained transitional substitutions, and the latter was mainly composed of transversional substitutions, suggesting that at least two mechanisms, possibly misincorporation during DNA replication and horizontal gene transfer, cause rRNA heterogeneity.

Base Composition↗

PubMed1999

Analysis by multiplex PCR of the physical status of human papillomavirus type 16 DNA in cervical cancers.

Integration of human papillomavirus (HPV) DNA occurs early in cancer development and is an important event in malignant transformation of cervical cancer. Integration of HPVs preferentially disrupts or deletes the E2 open reading frame, which results in the loss of its expression. The preferential disruption of the E2 gene causes the absence of the E2 gene sequences in the PCR product following integration. Twenty-two carcinomas positive for HPV type 16 (HPV-16) DNA were first tested for the disruption of the E2 gene by PCR. A specific fragment of the E2 gene was not amplified in 10 cases, suggesting integration of HPV DNA into the host genome. Next, multiplex PCR for the HPV E2 and E6 genes was carried out in the remaining 12 cases. Copy numbers of both genes should be equivalent in episomal forms, while the E2 gene copy number will be smaller than that for E6 following the preferential disruption of the E2 gene in concomitant forms. Although relative ratios of HPV E2 to E6 PCR products (E2/E6 ratios) ranged from 1.40 to 2.34 in 10 of 12 cases, multiplex PCR products from 2 cases displayed extremely low ratios of 0.69 and 0.61. Southern blot hybridization with an HPV-16 probe revealed that only in these two cases was both episomal and integrated HPV DNA being carried simultaneously. Thus, multiplex PCR for the E2 and E6 genes of HPV-16 DNA following PCR for the E2 gene can distinguish the pure episomal form from a mixed form of episomal and integrated HPV DNA. Clinical application of this technique will help researchers to understand the implication of the integration of HPV DNA for cervical carcinogenesis and cervical cancer progression.

Base Sequence↗

PubMed1999

Clinical experience of long-term transdermal treatment with nitric oxide donor for women with preeclampsia.

Isosorbide dinitrate (ISDN), a nitric oxide donor, was applied transdermally for 4-16 days to 4 preeclamptic women with oligohydramnios, intrauterine fetal growth retardation (IUGR), and elevated resistance of blood flow in the uterine arteries. Pulsed Doppler ultrasonography revealed immediate and drastic improvement of pulsatility index (PI) of uterine arteries following treatment with ISDN. The average PI in uterine arteries of the 4 patients was reduced to approximately 67% of that of the untreated state. In 2 patients the amniotic fluid gradually increased over a few days which suggested improvement of fetoplacental circulation during administration of ISDN. This study suggests that long-term transdermal ISDN is an effective therapy, at least in a portion of preeclamptic women, to avoid maternal hypertension, fetal distress, oligohydramnios, and IUGR, and consequentially to prolong the gestational period.

Administration, Cutaneous↗