Search PubMed⌕ Search

Biomedical subjects

K Ikuta

Publications and source records attributed to K Ikuta.

At least 199 records · Page 11Linked to original sources
PubMed1996

Heterogeneity of N insertion capacity in fetal hematopoietic stem cells.

TCR gene rearrangement is strictly regulated during mouse ontogeny. The V-(D)-J junctions of alphabeta and gammadelta TCR transcripts expressed in the adult thymus are more highly diverse than those in the fetal thymus. We previously showed that adult hematopoietic stem cells (HSC) have a higher capacity to insert N nucleotides into Vgamma4 TCR transcripts than fetal HSC and that the level of N nucleotide insertion is determined, at least in part, at the level of HSC. To analyze this developmental change of HSC at the single cell level, we investigated N nucleotide insertions in three TCR transcripts (Vgamma4, Vgamma2 and Vbeta8) derived from limiting numbers of fetal liver HSC by fetal thymic organ culture. Eight day-14 fetal liver HSC clones showed various levels of N nucleotide insertions in Vgamma transcripts (0-78%). On the other hand, the level of N insertions was similarly regulated in Vgamma4, Vgamma2, and Vbeta8 TCR transcripts in a clone-specific way. These results suggested that the level of N insertion is programmed at the level of single HSC and that fetal liver contains a heterogeneous population of HSC in terms of N insertion capacity. After 3 weeks culture with a stromal cell line, fetal HSC showed higher levels of N insertion capacity than before culture. This result and the presence of HSC with intermediate N insertion capacity support the hypothesis that the developmental potential of individual HSC gradually changes from fetal to adult type in one stem cell lineage.

Animals↗
PubMed1996

Preferential requirement of CD3 zeta-mediated signals for development of immature rather than mature thymocytes.

Antigen recognition signals by the TCR are transduced through activation motifs present in the cytoplasmic region of CD3 chains. In vitro analysis has suggested that the CD3zeta chain mediates different signals from other CD3 chains. To analyze the in vivo function of CD3zeta-mediated signals for T cell development, mice expressing a mutant CD3zeta chain lacking all the activation motifs were generated by introducing the transgene into zeta-knockout mice. Mature CD4(+) single-positive (SP) thymocytes in these mice were greater in number than in zeta-deficient mice, and the promoted differentiation was indicated by the changes of CD69 and HSA phenotypes. We found that even in the absence of activation motifs in CD3zeta, these mature cells became functional, being able to induce Ca2+ mobilization and proliferation upon stimulation. On the other hand, CD4(-)CD8(-) double-negative (DN) thymocytes, most of which were arrested at the CD44(-)CD25(+) stage similarly to those in zeta-deficient mice, could not be promoted for differentiation into CD4(+)CD8(+) double-positive thymocytes in these mice in spite of the fact that the expression of the transgene in DN thymocytes was higher than that of zeta in wild-type mice. These results demonstrate the preferential dependence of the promotion of development and/or expansion of DN thymocytes rather than mature thymocytes upon the activation signals through the zeta chain and suggest differential requirements of TCR signaling for mature SP and immature DN thymocyte developments in vivo.

Animals↗
PubMed1996

Identification of a Gly862 to Ser substitution in the type I collagen gene from a single spermatozoon.

Individual spermatozoa from the father of two affected infants with osteogenesis imperfecta were separated by dilution and micromanipulation. A segment of the type I collagen gene containing the mutant region was amplified by nested polymerase chain reaction and sequenced. Among 40 individual spermatozoa, 15 specimens were identified as mutants with a substitution of guanine3208 to adenine (glycine862 to serine) while 18 and seven specimens were of the wild and mixed types respectively. Through this study, we have established a molecular procedure that can be considered as prerequisite for preimplantation diagnosis of genetic disorders with a single point mutation.

Collagen↗
PubMed1996

Possible correlation between Borna disease virus infection and Japanese patients with chronic fatigue syndrome.

Borna disease virus (BDV) is a neurotropic, as yet unclassified, non-segmented, negative-sense, single-strand RNA virus. Natural infection with this virus has been reported to occur in horses and sheep. In addition, antibodies to BDV in plasma or BDV RNA in peripheral blood mononuclear cells (PBMCs) were also found in patients with neuropsychiatric diseases. We describe here the possible link between the patients with chronic fatigue syndrome (CFS) and infection with BDV.

Adult↗
PubMed1996

Unusually high seroprevalence of Borna disease virus in clade E human immunodeficiency virus type 1-infected patients with sexually transmitted diseases in Thailand.

The seroprevalence of Borna disease virus (BDV) in human immunodeficiency virus type 1-infected individuals in Thailand was examined by using recombinant BDV p24. A high (38 to 48%) rate of seroprevalence of BDV was observed in clade E-infected patients with sexually transmitted diseases, compared with those in clade E-infected prostitutes (8.3%), pregnant women (0%), clade B-infected intravenous-drug users (0%), and human immunodeficiency virus type 1-negative blood donors (1.9%).

AIDS-Related Opportunistic Infections↗
PubMed1996

Demonstration of borna disease virus RNA in peripheral blood mononuclear cells derived from domestic cats in Japan.

Borna disease virus (BDV) naturally infects horses, sheep, and several other species, including humans, and it is believed to be related to neurological disorders. BDV infection in domestic cats has also been demonstrated by serological assays. We demonstrated for the first time BDV RNA in peripheral blood mononuclear cells from 11 of 83 (13.3%) randomly selected domestic cats in Japan by nested reverse transcriptase-PCR. The BDVs from cats were similar to but slightly different from those from horses and humans, as shown by sequencing the reverse transcriptase-PCR products. None of the cats was positive for both BDV RNA and anti-BDV antibodies.

Amino Acid Sequence↗
PubMed1996

Sequence variability of Borna disease virus open reading frame II found in human peripheral blood mononuclear cells.

A cDNA fragment of the Borna disease virus (BDV) open reading frame II (ORF-II), which encodes a 24-kDa phosphoprotein (p24 [P protein]), was amplified from total RNA of peripheral blood mononuclear cells (PBMC) from three psychiatric inpatients. The amplified cDNA fragments were cloned, sequenced, and analyzed. A total of 15 clones, 5 from each patient, were studied. Intrapatient divergencies of the BDV ORF-II nucleotide sequence were 4.2 to 7.3%, 4.8 to 7.3%, and 2.8 to 7.1% for the three patients, leading to differences of 7.7 to 14.5%, 10.3 to 17.1%, and 6.0 to 16.2%, respectively, in the deduced amino acid sequence for BDV p24. Interpatient divergencies among the 15 clones were 5.9 to 12.7% at the nucleotide level and 12.8 to 28.2% at the amino acid level. Thus, in p24, BDV in human PBMC of the patients undergoes mutation at high rates in vivo. Additionally, we found that the nucleotide sequence of the 15 human BDV ORF-II cDNA clones differed from those of the horse strains V and He/80-1 by 4.2 to 9.3%. However, comparison of the consensus amino acid sequence deduced from the 15 human clones with those of the horse strains revealed no human-specific amino acid residue, suggesting that the BDV infecting humans may be related to that infecting horses.

Amino Acid Sequence↗
PubMed1996

Discrimination of grip force for preschool children aged 5 to 6 years.

The ability to discriminate grip force was investigated in 30 preschool children aged 5 to 6 years in an experiment controlling for motivation and muscle fatigue. The subjects were required to maintain a relatively high force during the discrimination with forces of the standard stimulus being 1.0, 2.0, and 3.0 kgf. Comparison stimuli changed at an interval of 0.1 kgf. Discrimination was measured in terms of the lower threshold, upper threshold, and interval of uncertainty. Since the statistical analyses indicated that there were no significant gender differences, data for boys and girls were combined. Although the upper and lower thresholds and the interval of uncertainty increased with the force required by the standard stimuli, Weber fractions did not remain constant when the stimulus intensity changed, unlike the findings in previous studies for adults.

Adult↗
PubMed1996

[Safer trials of vaccine and gene therapy against HIV].

HIV, the etiological agent of AIDS, induces depletion of CD4+T cells. Upon primary infection, there is an initial viremia that is followed rapidly by a cell-mediated immune response and apparent viral clearance. Thereafter, the persistence of low levels of HIV in the blood for years before the onset of the disease is facilitated by the ability of the virus to establish persistent and latent infection. Notwithstanding the latent form in most population, the clinical stage of disease is significantly associated with all measures of virus load levels. This may be mainly due to the function of HIV to induce apoptosis in a patients' uninfected CD4+T cells, as a bystander effect. Thus, the mechanisms that latently infect in the cells after HIV infection and stimulate active replication of HIV from the latency are essential for an understanding of the pathogenicity of the disease. Therefore, prophylactic and therapeutic trials should be focused to their effect to reduce the HIV load level.

AIDS Vaccines↗
PubMed1995

Extracellular Nef protein regulates productive HIV-1 infection from latency.

In HIV-1-infected asymptomatic carriers, the vast majority of infected cells in PBMCs are believed to be latently or nonproductively infected. We have isolated a subclone (MOLT-20-2) from an infected T cell line that expressed HIV-1 Ags at a very low level. However, viral Ag expression was markedly up-regulated by stimulation with either TNF-alpha, A23187, or PMA, indicating that the subclone might provide a suitable model of HIV-1 latency. Our previous studies have shown that the carboxyl-terminal region of the extracellular form of HIV-1 Nef played an important role in the interaction of infected cells with uninfected T cells, and could induce the cytostatic state. This suggested that Nef might contribute to intracellular signal transduction through an interaction with latently infected cells. We show in this study that stimulation of MOLT-20-2 with soluble Nef leads to HIV-1 activation from latency in a dose-dependent manner. Moreover, using a total of 14 overlapping Nef-related synthetic peptides, stimulatory activity was mapped to a discrete peptide (amino acid residues 132-147) that had the potential to activate latent HIV-1. This novel Nef function was confirmed by activation of virus production from the PBMCs of asymptomatic carriers. In addition, Nef-dependent HIV-1 activation from latency was also observed in another independently derived, latently infected cell line, U1, though not in cell line ACH-2. These results extend the significance of the Nef activity in vivo to the regulation of productive HIV-1 infection from latency, and define the regions of the protein involved.

Amino Acid Sequence↗
PubMed1995

Inhibition of pancreatic beta-cell glucokinase by antisense RNA expression in transgenic mice: mouse strain-dependent alteration of glucose tolerance.

We have generated transgenic mice, in either C57BL/6 or C3H background, expressing antisense glucokinase mRNA in beta-cells. The glucose phosphorylating activity at 60 mM glucose in transgenic islets was significantly lower than that in controls, and the insulin secretory response to glucose was lower in transgenic islets than in those of controls in both strains. Following i.p. glucose challenge, higher blood glucose levels were observed in transgenic mice than in controls in the C57BL/6 but not the C3H background. These data suggest that a beta-cell secretory defect, in combination with other undefined genetic factors, causes impaired glucose homeostasis in mice.

Animals↗
PubMed1995

Poly(ADP-ribose) polymerase activity in various U937 cell subclones with different susceptibility to HIV-1 infection: its dramatic decrease following persistent virus infection.

Poly(ADP-ribose) polymerase, a nuclear enzyme, is suggested to be involved in apoptotic cell death. It is also known that apoptotic cell death following HIV-1 infection is the most important feature of AIDS pathogenesis. Thus, to evaluate the relations between the enzyme and HIV-1 infection, we examined the enzyme activity of several subclones of human promonocytic cell line U937, which showed different susceptibility to HIV-1 infection. The nuclear extracts of two "high type clones" (possessing high susceptibility to HIV-1 infection) contained approximately 4 to 7-fold less enzyme than two low type clones when assayed under a full activation of enzyme. Parent clone, possessing an intermediate susceptibility to HIV-1, showed an intermediate enzyme level, suggesting that low level of this enzyme in cells is important for an effective infection of HIV-1. Furthermore, when these U937 subclones persistently infected with HIV-1 were examined, a dramatic decrease of the enzyme activity, reaching 2 to 16% of uninfected cells, was observed in all of these clones. The levels of poly(ADP-ribose) glycohydrolase in these clones were relativity unchanged. Activity gel analysis and immunoblotting of the enzyme in the clones revealed that the low enzyme activities observed in uninfected "high type clones" and all HIV-1-infected clones were due to a marked decrease of the enzyme protein itself. All of these results suggest that HIV-1 infection involves some mechanism to downregulate cellular poly(ADP-ribose) polymerase and that a lower level of the enzyme may be essential for an effective production of the virus and/or for a stable virus/host interaction.

Acquired Immunodeficiency Syndrome↗
PubMed1995

Demonstration of human Borna disease virus RNA in human peripheral blood mononuclear cells.

BDV naturally infects horses and sheep, and causes sporadic neurological disease. Serological evidence suggests an association of BDV, or a related virus, with specific psychiatric diseases in humans. Here, by using a nested RT-PCR technique, we demonstrate that human BDV RNA is present in the PBMC of psychiatric patients. In an examination of a total of 60 patients from 5 wards of a hospital in Japan, the detection rate differed within each ward, ranging from 8% to > 50% (37% on the average). Of particular note was the finding that the human derived BDV sequences, which included deleted forms in about 23% of the positive samples, were slightly different from those derived from horse BDV. These results suggest urgent consideration of the measures to be taken to cope with the effects of blood transfusion. In addition, the detection of a high level of BDV in the PBMC of patients will help our understanding of the pathogenesis in the disease.

Antibodies, Viral↗
PubMed1995

Existence of branched side chains in the cell wall mannan of pathogenic yeast, Candida albicans. Structure-antigenicity relationship between the cell wall mannans of Candida albicans and Candida parapsilosis.

Isolation of side chain oligosaccharides from mannans of Candida albicans NIH B-792 (serotype B) and Candida parapsilosis IFO 1396 strains has been conducted by acetolysis under mild conditions. Structural study of these oligosaccharides by 1H and 13C NMR and methylation analyses indicated the presence of novel branched side chains with the following structures in C. albicans mannan. [sequence: see text] It was observed that the H-1 proton chemical shifts of the second and the third mannose units from the reducing terminus in each oligosaccharide are shifted upfield by substitution with an alpha-linked mannose unit at position 6 of the 3-O-substituted mannose unit. An agglutination inhibition assay between factor 4 serum and cells of Candida stellatoidea IFO 1397 lacking the beta-1,2-linked mannose unit, with oligosaccharides obtained from these mannans, indicated that only the branched oligosaccharides were active. This finding suggests that the branched oligosaccharides correspond to the epitope of antigenic factor 4. The presence of the branched structure in other mannans was detected by the characteristic H-1-H-2-correlated cross-peak of the alpha-1,2-linked mannose unit connected with the 3,6-di-O-substituted one by two-dimensional homonuclear Hartmann-Hahn spectroscopy.

Acids↗
PubMed1995

Interaction stabilizing tertiary structure of bacteriorhodopsin studied by denaturation experiments.

The structural stability of bacteriorhodopsin was studied by denaturation experiments, using aliphatic alcohol as denaturants. The disappearance of a positive peak at 285 nm of the circular dichroism spectra, the change in the intrinsic fluorescence decay time, and the decrease of the regeneration activity bacteriorhodopsin indicated the denaturation of the tertiary structure of this protein at a methanol concentration of about 3 M. The circular dichroism band at 222 nm was unchanged by the denaturation. It was concluded that the alcohol-denatured state in water was similar to the molten globule state of soluble proteins, in which only the tertiary structure was destroyed. Solvent substitution from water to hexane did not cause denaturation of bacteriorhodopsin. However, further addition of alcohol destroyed the secondary as well as the tertiary structures. Comparing the alcohol effects of bacteriorhodopsin in water to that in hexane, the dominant interactions for the structure formation of this protein could be revealed: the hydrophobic interaction that arose from the structure of water is essential for the stability of membrane spanning helices, while the interaction which binds the helices is polar in nature.

Alcohols↗
PubMed1995

Salmon GnRH synthesis in the preoptic area and the ventral telencephalon is activated during gonadal maturation in female Masu Salmon.

Changes in salmon gonadotropin-releasing hormone (sGnRH) synthetic activity in the brain during gonadal maturation were examined by in situ hybridization in 2-year-old female masu salmon. Oncorhynchus masou. During gonadal maturation, the numbers of neurons expressing sGnRH mRNA increased in the preoptic area and the ventral telencephalon, but not in the olfactory bulbs and the terminal nerve ganglion. The numbers of silver grains per neuron also increased in the preoptic area and the ventral telencephalon. These results indicate that sGnRH has multiple physiological functions according to the location of the neurons in the brain; neurons in the preoptic area and the ventral telencephalon are involved in gonadal maturation possibly by stimulating gonadotropin synthesis and release, whereas neurons in the olfactory bulbs and the terminal nerve ganglion may have different roles.

Animals↗