Biomedical subjects

M Kikuchi

Publications and source records attributed to M Kikuchi.

At least 307 records · Page 17Linked to original sources

PubMed1998

Cloning of complementary deoxyribonucleic acid for the follicle-stimulating hormone-beta subunit in the Japanese quail.

Complementary DNA clones encoding the luteinizing hormone-beta and common alpha subunits have been isolated not only in mammals but also in some nonmammalian tetrapod vertebrates. However, cloning of cDNA encoding the follicle-stimulating hormone (FSH)-beta subunit is limited to mammals, and no clone for the FSH-beta subunit has been isolated for nonmammalian vertebrates. We report here the isolation and characterization of cDNA encoding the FSH-beta subunit precursor molecule in the Japanese quail from a cDNA library of the pituitary gland of this bird. As the hybridization probe for the screening, we used a cDNA clone prepared by the polymerase chain reaction (PCR) with primers designed from known nucleotide sequence data of cDNA for FSH-beta subunit precursors of mammals. The FSH-beta subunit precursor cDNA we isolated was longer than any other FSH-beta subunit precursor cDNAs previously reported. This is due to the extraordinarily long 3'-untranslated region (2135 bp). This region is extremely rich in consensus sequences reported to cause instability of mRNA, suggesting that mRNA for the FSH-beta subunit, especially that of the Japanese quail, is unstable. Northern blot analysis of mRNA for the FSH-beta subunit revealed that the pituitary content of mRNA in the nonbreeding season was about 1/5 to 1/10 that in the breeding season in male Japanese quail kept in an outdoor cage. The profile of the seasonal change in FSH-beta subunit mRNA in the quail pituitary gland was similar to that of the seasonal change in the concentration of FSH in plasma reported previously. The deduced amino acid sequence of the mature protein region showed that the quail FSH-beta subunit is more similar to the opossum FSH-beta subunit than to the other mammalian FSH-beta subunits.

Amino Acid Sequence↗

PubMed1998

Molecular evolution of the domain structures of protein disulfide isomerases.

Protein disulfide isomerase (PDI) is an enzyme that promotes protein folding by catalyzing disulfide bridge isomerization. PDI and its relatives form a diverse protein family whose members are characterized by thioredoxin-like (TX) domains in the primary structures. The family was classified into four classes by the number and the relative positions of the TX domains. To investigate the evolution of the domain structures, we aligned the amino acid sequences of the TX domains, and the molecular phylogeny was examined by the NJ and ML methods. We found that all of the current members of the PDI family have evolved from an ancestral enzyme, which has two TX domains in the primary structure. The diverse domain structures of the members have been generated through domain duplications and deletions.

Evolution, Molecular↗

PubMed1998

Repetitive mitochondrial Ca2+ signals synchronize with cytosolic Ca2+ oscillations in the pancreatic beta-cell line, MIN6.

We examined the relationship between cytosolic Ca2+ concentration ([Ca2+]c) and mitochondrial matrix Ca2+ concentration ([Ca2+]m) in the pancreatic beta-cell line, MIN6. [Ca2+]c was monitored in a single or a group (30 cells) of fura-2-loaded MIN6 cells, and [Ca2+]m was measured in a group (1 x 10[6] cells) of MIN6 cells stably transfected with aequorin targeted at the mitochondria. Exogenous ATP (0.25 mmol/l) produced a single transient increase in [Ca2+]c whereas 22 mmol/l KCl produced a sustained plateau increase. ATP and KCl evoked transient increases in [Ca2+]m but with distinct time courses of [Ca2+]m decline: the [Ca2+]m increase induced by ATP decreased more rapidly than that induced by KCl. Nitrendipine (3 micromol/l), a blocker of L-type Ca2+ channels, inhibited both [Ca2+]c and [Ca2+]m signals in response to KCl and tolbutamide, but not those to ATP. Peak levels of [Ca2+]m increase (around 2 micromol/ l) exceeded those of [Ca2+]c increase (around 500 nmol/l). A rise in glucose concentration from 3 to 30 mmol/l induced oscillations of [Ca2+]c that overlay the sustained increases in [Ca2+]c in single cells. An oscillatory increase in [Ca2+]m was similarly observed in response to glucose. Addition of 10 mmol/l 2-ketoisocaproic acid at 20 mmol/l glucose further increased the plateau level of [Ca2+]c and the frequency of [Ca2+]c oscillations, which were correlated with a further increase in [Ca2+]m. In response to pulsatile exposure to KCl, [Ca2+]c and [Ca2+]m increased synchronously. These data suggest that an oscillatory increase in [Ca2+]m in beta cells, the signal which is thought to be necessary for continuous stimulation of mitochondrial metabolism, is produced synchronously with the [Ca2+]c oscillations.

Adenosine Triphosphate↗

PubMed1998

Role of JTT-501, a new insulin sensitiser, in restoring impaired GLUT4 translocation in adipocytes of rats fed a high fat diet.

JTT-501 is an insulin-sensitising compound with an isoxazolidinedione rather than a thiazolidionedione structure. Sprague-Dawley rats fed a high fat diet for 2 weeks were used as an animal model of insulin resistance, and JTT-501 was administered for the final week of the diet. An euglycaemic glucose clamp study showed that the glucose infusion rate (GIR) required to maintain euglycaemia was 57% lower in rats fed a high fat diet than in control rats, and that JTT-501 treatment restored the reduction in GIR produced by the high fat diet. To explain the mechanisms underlying the effects of a high fat diet and JTT-501 treatment, epididymal fat pads were excised and used in the analysis of insulin action. The high fat diet caused: (1) a 58% decrease in insulin receptor substrate-1 (IRS-1) content with a 58% decrease in IRS-1 tyrosine phosphorylation; (2) reductions of 56% and 73% respectively in insulin-induced maximal PI 3-kinase activation in anti-phosphotyrosine and anti-IRS-1 antibody immunoprecipitates; (3) a 46% reduction in the glucose transporter protein, GLUT4 content and, consequently, (4) severely impaired insulin-induced GLUT4 translocation to the plasma membrane and glucose uptake in adipocytes. JTT-501 treatment restored appreciably the protein content and tyrosine phosphorylation level of IRS-1. Insulin-stimulated PI 3-kinase activation was also restored in anti-phosphotyrosine and anti-IRS-1 antibody immunoprecipitates. As reflected by these improvements in insulin signalling, JTT-501 treatment improved considerably insulin-induced GLUT4 translocation to the plasma membrane as well as insulin-induced glucose uptake. However, JTT-501 had no effect on the decrease in GLUT4 content produced by the high fat diet. These observations suggest that JTT-501 enhances insulin signalling and may be effective in reducing insulin resistance.

Adipocytes↗

PubMed1998

Abnormal functional connectivity in Alzheimer's disease: intrahemispheric EEG coherence during rest and photic stimulation.

Electroencephalography (EEG) coherence provides a measure of functional correlations between two EEG signals. The present study was conducted to examine intrahemispheric EEG coherence at rest and during photic stimulation (PS; 5, 10 and 15 Hz) in ten unmedicated patients with presenile dementia of the Alzheimer type (AD; mean age at onset 56 years). In the resting EEG, the AD patients had significantly lower coherence than gender- and age-matched control subjects in the alpha-1, alpha-2 and beta-1 frequency bands. The EEG analysis during PS also showed that the patients had significantly lower coherence in the frequency corresponding to PS at 10 and 15 Hz. In this study, the changes in coherence from the resting state to the stimulus condition (i.e. PS-related coherence reactivity) were examined. The patients were found to show significantly smaller coherence reactivity to PS at 5 and 15 Hz. These findings suggest that, in addition to the resting state, AD patients have an impairment of intrahemispheric functional connectivity during PS. They also suggest that AD shows a failure of PS-related functional reorganization.

Aged↗

PubMed1998

Supernumerary ring chromosomes and nuclear blebs in some low-grade malignant soft tissue tumours: atypical lipomatous tumours and dermatofibrosarcoma protuberans.

We investigated the diagnostic significance of supernumerary ring chromosomes in low-grade soft-tissue neoplasms. Chromosome slides were prepared from 123 samples of soft-tissue tumours using the standard trypsin-Giemsa banding technique. Supernumerary ring chromosomes were found in 6 cases of soft tissue tumours: 5 cases of atypical lipomatous tumour (ALT) and 1 case of dermatofibrosarcoma protuberans (DFSP). By chromosome painting with fluorescence in situ hybridization (FISH), the ring chromosome in 1 ALT was painted over its entire length with the chromosome 12 probe. Nuclear blebs and micronuclei, which were observed in each case of ALT, also contained chromosome 12 material; and these structures may represent a topological distribution of ring or giant marker chromosomes in the interphase nuclei. Our findings suggest that supernumerary ring chromosomes are characteristic of some low-grade soft tissue neoplasms including ALT and DFSP.

Adult↗

PubMed1998

Apoptosis of cytotoxic T-cells in histiocytic necrotizing lymphadenitis.

Cell death is necrotic or apoptotic. In histiocytic necrotizing lymphadenitis (HNL), apoptosis is the main form of cell death, resulting in the creation of nuclear debris that is one of the characteristic features of HNL. To investigate the cell type of apoptotic cells, 12 cases of HNL were analyzed using the immunohistochemical staining for TIA-1, a cytotoxic granule of either cytotoxic T or NK cells. One quarter to over half of all apoptotic cells were positive for TIA-1, and some of the nuclear debris was also positive. The necrotic lesions of HNL were found to consist of nuclear debris, apoptotic cells, histiocytes and lymphocytes. The lymphocytes were mainly CD8-positive T-cells or CD4-positive cells, while B- and NK cells were only rarely observed. The number of TIA-1-positive lymphocytes was more closely related to the number of CD8-positive cells than to the number of CD4 cells. In double staining, the TIA-1 positive lymphocytes were mainly CD8 positive, but rarely CD4 positive. In HNL, then, CD8-positive cytotoxic T-cells are likely to undergo apoptosis.

Adolescent↗

PubMed1998

Characterization of neutralizing monoclonal antibody escape mutants of Hantaan virus 76118.

Neutralizing monoclonal antibody (MAb) escape mutants of Hantaan virus were generated using MAbs to envelope protein G1 (16D2) and G2 (11E10). The mutant viruses (mu16D2 and mu11E10), lacked reactivity only to the selecting MAb, or a MAb belonging to the same antigenic site. Both mutants had a single amino acid (a.a.) substitution. The a.a. substitution, found in mu16D2, was different from that found in another mutant selected with the same MAb (16D2). Although MAb 11E10 immunoprecipitated G2 protein, a deduced a.a. substitution was located in the G1 region. These results suggest that antigenic sites defined by neutralizing MAbs are composed of discontinuous epitopes over the G1 and G2 proteins. Mutant 11E10 showed a significant decrease in virulence in suckling mice. A virulence revertant of mu11E10, selected through passages in suckling mice brain, showed exactly the same deduced a.a. sequence as mu11E10 and still was not neutralized by MAb 11E10. Since mutant 16D2 was virulent for suckling mice, neutralization related epitopes found with MAbs 11E10 and 16D2 were independent of pathogenicity in BALB/c mice.

Amino Acid Substitution↗

PubMed1998

Characterisation of Trypanosoma cruzi populations by DNA polymorphism of the cruzipain gene detected by single-stranded DNA conformation polymorphism (SSCP) and direct sequencing.

Fifty fresh isolates of Trypanosoma cruzi from Triatoma dimidiata vectors and 31 from patients with Chagas' disease were analysed for DNA polymorphisms within the 432-bp core region of the cruzipain gene which encodes the active site of cathepsin L-like cystein proteinase. The cruzipain gene showed signs of polymorphism consisting of four different DNA sequences in Central and South American isolates of T. cruzi. The PCR fragments of Guatemalan isolates could be divided into three groups, Groups 1, 2 and 3, based on different patterns of single-stranded DNA conformation polymorphism. All of the strains isolated from Brazil, Chile, and Paraguay, except for the CL strain, showed a Group 4 pattern. Two to four isolates from each group were analysed by cloning and sequencing. A silent mutation occurred between Groups 1 and 2, and five nucleotides and two aa substitutions were detected between Groups 1 and 3. The DNA sequence of Group 4 contained five nucleotides and one aa substitution from Group 1. All of the DNA sequences corresponded well with the single-stranded DNA conformation polymorphism. The Group 1 isolates, the majority in the Guatemalan population (70/81, 86.4%), were isolated from both triatomines and humans, but Group 3 were isolated only from humans. Moreover, the Group 2 isolates were detected only in triatomine vectors (9/50; 18%), but never in humans (0/32, P<0.05) suggesting that this group has an independent life-cycle in sylvatic animals and is maintained by reservoir hosts other than humans.

Amino Acid Sequence↗

PubMed1998

Retinoic acids induce growth inhibition and apoptosis in adult T-cell leukemia (ATL) cell lines.

Adult T-cell leukemia (ATL) is a peripheral T-cell neoplasm caused by human T-cell leukemia virus type I (HTLV-I). Despite the administration of combined intensive chemotherapy, the reported survival time of patients with acute and lymphoma types of ATL is less than 10 months. We therefore examine the effects of all-trans-retinoic acid (ATRA), 9-cis-RA and 13-cis-RA and tried to elucidate the mechanisms of inducing growth inhibition and apoptosis by these RAs using four ATL cell lines established in our laboratory. All the investigated RAs inhibited cell growth and the cells were arrested at the G1 phase. Apoptosis was induced in three out of four cell lines. Among the growth regulatory proteins examined, the level of p21Waf1/Cip1 protein was found to increase after RA treatment, thus resulting in pRb hypophosphorylation which also induced the arrest of the cells at the G1 phase. In addition, the p53 level decreased at the same time. Fas-FasL system and the downregulation of CD25 (IL-2R/alpha) expression did not seem to be involved. Based on these findings, the ability of RAs to induce a remission of ATL is thus strongly suggested.

Adult↗

PubMed1998

Delayed gastric emptying by Helicobacter pylori lipopolysaccharide in conscious rats.

The present study was carried out to investigate the possibility that lipopolysaccharide deprived from Helicobacter pylori may alter gastric motility. To address the question, we examined the effect of H. pylori lipopolysaccharide on gastric emptying in conscious rats. Gastric emptying was evaluated by the phenol red method. Time-course and dose-related effects of intraperitoneal administration of H. pylori lipopolysaccharide were investigated. Intraperitoneal injection of H. pylori lipopolysaccharide significantly suppressed gastric emptying of a liquid meal in a dose-dependent manner. The inhibitory action of H. pylori lipopolysaccharide was observed 2, 4, 8, or 12 hr after the injection. These results suggest for the first time that H. pylori lipopolysaccharide may suppress gastric emptying in a long-lasting fashion. It is also suggested that H. pylori may influence gastric function through its cell wall structure named lipopolysaccharide.

Animals↗

PubMed1998

Nodal T-cell lymphoma in an HTLV-I-endemic area: proviral HTLV-I DNA, histological classification and clinical evaluation.

Adult T-cell leukaemia/lymphoma (ATLL) is a human malignancy associated with human T-cell leukaemia virus type I (HTLV-I). The histology usually indicates a pleomorphic type, but is not consistent. To clarify the relationship between the histological classification and prognosis in ATLL, and to confirm the significance of clonal HTLV-I integration, we reclassified 572 cases with nodal T-cell lymphoma in which the T-cell phenotype and/or genotype was confirmed. In all cases the clonal integration of HTLV-I proviral DNA in the lymph nodes was examined by Southern blot analysis. In addition, anti-ATL antigen (ATLA) determination in the serum or PCR analysis of HTLV-I pX amplification in lymph nodes was also performed. 66/313 (21%) cases with ATLA had no evidence of clonal HTLV-I integration. 572 cases were classified into three groups: (A) cases with clonal integration (247 cases), (B) cases with ATLA without clonal integration of HTLV-I proviral DNA (66 cases), (C) cases without ATLA (259 cases). Histologically, groups B and C frequently demonstrated large cell type and angioimmunoblastic lymphadenopathy with dysproteinaemia (AILD) type; however, group A tended to show a pleomorphic type. Clinically, group A showed a poorer prognosis than groups B and C. In conclusion, group A cases were defined as ATLL (HTLV-I-associated T-cell lymphoma), whereas group B was classified as T-cell lymphoma, which had coincidently occurred in HTLV-I infected carriers. The simplified classification of REAL indicated clinical outcome: the prognosis of ATLL was poor, the unspecified type was intermediate, whereas the other types of lymphoblastic, AILD and anaplastic large cell type were all relatively favourable.

Blotting, Southern↗

PubMed1998

Elevated serum soluble Fas ligand in natural killer cell proliferative disorders.

We evaluated the serum level of soluble Fas ligand (sFasL) in patients with natural killer lymphocyte proliferative disorders (NK- LPD). The serum sFasL level was elevated in neoplastic groups of aggressive NK leukaemia, indolent NK leukaemia and NK lymphoma, all of which contained clonal EBV-DNA. In NK leukaemia the serum sFasL level was significantly higher than that found in others. However, it was not elevated in the patients with reactive NK-LPD and in one patient with NK leukaemia in remission. These findings indicate that the serum sFasL level is a useful indicator in evaluating disease activity.

Adolescent↗

PubMed1998

Beta2-glycoprotein I-dependent and -independent anticardiolipin antibody in non-obese diabetic (NOD) mice.

In the present study we investigate whether or not anticardiolipin antibody (aCL) is produced in NOD mice, which is a representative animal model of insulin-dependent diabetes mellitus (IDDM). We found that IgG class aCL appeared in 6.9% of non-diabetic NOD mice at weeks 5-15. The rates increased with age to 31.6% at weeks 16-25 and 71.9% at weeks 26-35. In addition, the titre and incidence of aCL were higher in diabetic mice than in non-diabetic mice. It was also found that aCL in NOD mice involved beta2-glycoprotein I (beta2-GPI)-dependent and -independent aCL, when beta2-GPI was added to the aCL assay system. The IgG subclass of both beta2-GPI-dependent and -independent aCL belonged exclusively to IgG2a. In addition, immunohistochemical studies revealed the predominant accumulation of IgG2a- or IgG3-positive B lymphocytes within insulitis. These results suggest that the autoimmunity in NOD mice may thus be associated with Th1 predominant immunological response. In conclusion, aCL with multiple antigenic specificity were produced in NOD mice along with the development of insulitis and diabetes. NOD mice should thus be added to the list of animal models possessing antiphospholipid antibody.

Animals↗

PubMed1998

Perforin and Fas pathways of cytotoxic T-cells in histiocytic necrotizing lymphadenitis.

AIMS: Cell death can be divided into necrosis and apoptosis. In histiocytic necrotizing lymphadenitis (HNL), apoptosis is the main form of cell death. Two molecular mechanisms of T-cell-mediated cytotoxicity, one perforin-based and the other Fas-based, have been demonstrated, and both systems induce apoptosis of the target cells. This study was designed to investigate the Fas and perforin pathways in HNL. METHODS AND RESULTS: Twelve cases of HNL were analysed using immunohistochemical staining. The Fas and/or FasL-positive cells, including lymphocytes and histiocytes, were frequently encountered in the necrotizing lesions, however, they were rare in the nonpathological regions. The perforin and/or granzyme-positive cells were also confined in the necrotizing lesions. In double staining, CD8-positive lymphocytes occasionally expressed Fas and/or FasL, and histiocytes also expressed FasL and/or Fas. However, CD4-positive lymphocytes rarely expressed FasL and/or Fas. In a flow cytometric analysis, most of the cytotoxic T-cells, which were recognized by cytolytic granules of TIA-1 and considered to be CD8-positive lymphocytes, expressed FasL and Fas. The perforin-positive lymphocytes also expressed FasL and Fas. CONCLUSION: In our previous study, the apoptotic cells were T-cells, especially CD8-positive cells rather than CD4-positive cells. Based on these findings, in Fas and perforin pathways, the CD8-positive cells were considered to be effector and target cells, while histiocytes could possibly be enhancers. As a result, both pathways seemed to induce an abundance of apoptosis and thus induce necrotizing lesions.

Adolescent↗

PubMed1998

Insulin secretion from isolated rat islets induced by the novel hypoglycemic agent A-4166, a derivative of D-phenylalanine.

A derivative of D-phenylalanine, A-4166, reportedly evokes a more rapid and short-lived hypoglycemic action in vivo than any of the currently available sulfonylureas. This novel oral hypoglycemic agent is structurally different from sulfonylureas. Therefore, studies were designed to elucidate the mechanisms by which A-4166 stimulates insulin secretion. Insulin release from incubated or perifused rat islets was dose-dependently stimulated by 10 to 200 mumol/l A-4166, in the presence of 2.8 mmol/l glucose. Both A-4166 and tolbutamide evoke a prompt rise in insulin secretion followed by a sustained gradually decreasing release from perfused islets in the presence of low glucose, although A-4166 appeared to be more sensitive than tolbutamide to subthreshold glucose concentration. Diazoxide abolished the initial release and blunted sustained release. Removing calcium from the perifusate abolished insulin release within 15 minutes. A-4166 inhibited [3H]-glibenclamide binding to HIT cell membranes and 86Rb efflux from ATP-depleted or diazoxide-treated cells. These results suggest that the insulin release induced by A-4166 is relevant to this agent occupying the tolbutamide binding sites. Therefore, one possible mechanism accounting for the more rapid and short-lived hypoglycemic action of A-4166 in vivo, as compared with tolbutamide, may involve the reported differences in the bioavailability of A-4166.

ATP-Binding Cassette Transporters↗

PubMed1998

Glycine-valine dimorphism at the 86th amino acid of HLA-DRB1 influenced the prognosis of postschistosomal hepatic fibrosis.

Chinese patients (n = 113) with schistosomal hepatic fibrosis diagnosed by ultrasonography (grade I, II, or III) and 184 age- and sex-matched persons with no clinical information of schistosomal infection were typed for their HLA-DRB1 alleles by DNA typing. There was no single allele that conferred susceptibility or resistance to fibrosis. However, there were three groups of alleles that showed decreased (resistant), increased (susceptible), or neutral frequency in the patients with fibrosis. The susceptible alleles, DRBI *1202, DRB1 *1404, and DRBI *1405, shared a valine at amino acid residue 86, whereas the resistant alleles, DRB1*11011, DRB1*0409, and DRB1*0701, all had glycine at position 86. Therefore, this study focused on the glycine-valine dimorphism at aa 86, which influences the depth of the P1 pocket in the antigen binding groove, and found that the 86th valine allele was significantly increased in the patients with fibrosis (odds ratio = 2.2; 95% CI = 1.34-3.61, corrected P < .05).

Alleles↗

PubMed1998

Isolation of anti-glutathione antibodies from a phage display library.

We have isolated anti-glutathione antibodies from a human synthetic phage antibody scFv library (Nissim,A., Hoogenboom,H.R., Tomlinson,I.M., Flynn,G., Midgley,C., Lane,D. and Winter,G., 1994, EMBO J., 13, 692-698). Glutathione (GSH) conjugates with carrier proteins, such as bovine serum albumin (BSA), keyhole limpet hemocyanin (KLH) and human lysozyme (LZM), were used as antigens. After four cycles of panning and affinity chromatography, clones that recognized GSH-conjugated proteins, but not BSA, KLH or LZM, were isolated. The isolated phage antibodies and the soluble scFv fragments were characterized by immunoblotting, and the nucleotide sequences of the VH segments of selected clones were determined. The binding of several isolates to GSH-BSA was competitively inhibited by GSH in an ELISA. These observations have demonstrated that antibodies against GSH, a tripeptide, can be isolated from the library. We constructed the tertiary models of several scFv fragments and discussed the mechanism of antigen binding sites.

Antibodies, Viral↗