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Biomedical subjects

K Ikuta

Publications and source records attributed to K Ikuta.

At least 19 recordsLinked to original sources
PubMed1992

Development of gamma delta T-cell subsets from fetal hematopoietic stem cells.

Hematopoietic stem cells (HSC) were isolated from mouse fetus, and their developmental potential was compared with adult HSC. Donor-derived V gamma 3+T cells were detected in fetal thymic lobes, repopulated in vitro with fetal liver HSC, but not in those with adult bone marrow HSC. Single clonogenic fetal HSC gave rise to thymic progeny that include V gamma 3+, other gamma delta+, and alpha beta+ T cells. No V gamma 3+ T cells were detected in adult thymus injected intrathymically with either fetal or adult HSC. These results support a hypothesis that only fetal HSC have the capacity to differentiate into V gamma 3+ T cells in the fetal thymic microenvironment, and that the developmental potential of HSC may change during ontogeny.

Aging↗
PubMed1992

Evidence that hematopoietic stem cells express mouse c-kit but do not depend on steel factor for their generation.

The interaction of the mouse c-kit receptor, designated Kit receptor, and steel factor promotes the proliferation and differentiation of hematopoietic progenitor cells. Monoclonal antibodies against the extracellular portion of the mouse Kit receptor were established. Five percent to 10% of total bone marrow cells expressed the Kit receptor, and half of them lack the expression of lineage markers. The Kit receptor was expressed on 70-80% of Thy-1.1lo Lin-Sca-1+ cells, which express Thy-1.1 antigen at a low level and constitute approximately 0.05% of adult bone marrow and fetal liver; by previous studies, these cells have been shown to be highly enriched for multipotent hematopoietic stem cells (HSCs) and are the only hematopoietic cell subset with this activity. Spleen colony formation and long-term multilineage reconstitution activities were contained in the Kit+ but not in the Kit- subpopulations of Thy-1lo Lin-Sca-1+ cells from adult bone marrow, suggesting that the Kit receptor is expressed on HSCs from the earliest stage-i.e., pluripotent HSCs. The role of steel factor in the development and self-renewal of HSCs was tested with Sl/Sl homozygote fetuses, which lack genes to encode functional steel factor. They were shown to have 30-40% of the number of HSCs on days 13-15 when compared with normal litermates. However, the absolute number of HSCs increased during fetal development in the Sl/Sl mice. The results suggest that the Kit receptor-steel factor interaction may not be essential for the initiation of hematopoiesis and the self-renewal of (at least) fetal HSCs.

Animals↗
PubMed1992

Determination and validity of critical velocity as an index of swimming performance in the competitive swimmer.

The purpose of this investigation was to test whether the concept of critical power used in previous studies could be applied to the field of competitive swimming as critical swimming velocity (vcrit). The vcrit, defined as the swimming velocity over a very long period of time without exhaustion, was expressed as the slope of a straight line between swimming distance (dlim) at each speed (with six predetermined speeds) and the duration (tlim). Nine trained college swimmers underwent tests in a swimming flume to measure vcrit at those velocities until the onset of fatigue. A regression analysis of dlim on tlim calculated for each swimmer showed linear relationships (r2 greater than 0.998, P less than 0.01), and the slope coefficient signifying vcrit ranged from 1.062 to 1.262 m.s-1 with a mean of 1.166 (SD 0.052) m.s-1. Maximal oxygen consumption (VO2max), oxygen consumption (VO2) at anaerobic threshold, and the swimming also velocity at the onset of blood lactate accumulation (vOBLA) were also determined during the incremental swimming test. The vcrit showed significant positive correlations with VO2 at anaerobic threshold (r = 0.818, P less than 0.01), vOBLA (r = 0.949, P less than 0.01) and mean velocity of 400 m freestyle (r = 0.864, P less than 0.01). These data suggested that vcrit could be adopted as an index of endurance performance in competitive swimmers.

Adolescent↗
PubMed1992

Relationship between superoxide dismutase (SOD) and viral liver diseases.

Immunohistochemical evaluation of Cu, Zn- and Mn-superoxide dismutase (SOD) activity in various viral liver diseases was performed by the peroxidase-conjugated antibody indirect method. Anti-human Cu, Zn-SOD (rabbit) and anti-human Mn-SOD (guinea-pig) derived and purified from SOD of human erythrocytes and placentas were used to determine SOD distribution in liver tissues. SOD in the liver tissues was detected in 68 inpatients of our unit. They consisted of 23 cases with chronic hepatitis caused by hepatitis B virus (13) and hepatitis C virus (10), 24 with liver cirrhosis caused by hepatitis B virus (5) and hepatitis C virus (19) (15: compensatory, 9: decompensatory) and 21 with hepatocellular carcinoma caused by hepatitis B virus (2) and hepatitis C virus (18) complicated of liver cirrhosis. In viral liver diseases, SODs in the liver tissues were distributed to hepatocytes mainly in the pattern of cytoplasmic diffusion. The incidence of immunohistochemical Cu, Zn-SOD and Mn-SOD were 47.8% and 56.5% in chronic hepatitis, 93.3% and 86.7% in compensated liver cirrhosis, 11.1% and 22.2% in decompensated liver cirrhosis, respectively. The aggression of viral liver disease was accompanied with the decrease of SOD concentration in the liver tissues. Hepatocellular carcinoma cells were negative for Mn-SOD in all cases, and weakly positive for Cu, Zn-SOD in 2 out of 21 cases. Comparatively strongly positive SOD findings were obtained from normal regions neighboring carcinomas. A close relationship between the depletion of SOD in liver tissues and carcinogenesis in viral liver diseases was observed.

Carcinoma, Hepatocellular↗
PubMed1992

Monoclonal antibodies to a CD4 peptide derivative which includes the region corresponding to an immunoglobulin CDR3: evidence of the involvement of pre-CDR3-related region in HIV-1 and host cell interaction.

A CD4 peptide of amino acid residues 68-130 [CD4(68-130)], which had the capacities to inhibit HIV-1 replication and HIV-1-induced syncytium formation, was used as an immunogen for the preparation of mAb. The mAbs prepared were classified into at least five types (I-V) in terms of their recognition sites by ELISA using various kinds of smaller CD4 peptides. Among them, the type I mAb no. 35 recognizing amino acid residues 72-84, which lies just before the region corresponding to an immunoglobulin third complementarity-determining region (CDR3), showed the strongest effects in reducing both HIV-1 infection and HIV-1-induced syncytium formation, although a large amount of no. 35 mAb was necessary to reduce such HIV-1 activities compared with those of anti-Leu-3a and OKT4A mAbs which recognize CD4 epitopes near a portion corresponding to an immunoglobulin CDR2. Western blot analysis showed that the reactivities of CD4 molecule in CD4-positive cells or sCD4 molecule with types I-V mAbs were stronger than that with anti-Leu-3a mAb. Flow cytometry showed that no. 35 mAb was faintly reactive with native CD4 molecule on cell surface at the concn showing the inhibitory effects on HIV-1 infection and syncytium formation. In addition, a smaller peptide CD4(66-92), one of the good epitope peptides for no. 35 mAb, also showed strong inhibitory effect on HIV-1 infection as well as a weaker inhibitory effect on syncytium formation. These results suggest that, in addition to the CD4 CDR2-related region, the pre-CDR3-related region is also involved in the early events of the interactions between the host cell and HIV-1.

Amino Acid Sequence↗
PubMed1992

Major core proteins, p24s, of human, simian, and feline immunodeficiency viruses are partly expressed on the surface of the virus-infected cells.

We have previously shown the expression of human immunodeficiency virus type 1 (HIV-1) major gag protein, p24, on the surface of persistently HIV-1-infected cells by using murine monoclonal antibodies (mAb). We now report that the cell surface gag p24 antigen expression is a universal phenomenon among HIV-1, simian immunodeficiency virus (SIV), and feline immunodeficiency virus (FIV). The mAbs prepared by immunization with purified HIV-1 particles were used as antibodies cross-reactive to HIV-1 and SIVagmp24 antigens. The mAbs to FIV p24 were raised against the gag precursor 50 kDa protein of FIV, which was expressed by Baculovirus vector. The p24 antigen expression on the cell surface was detectable in certain combinations of virus-host cell systems in all of these viruses. Since these p24 regions of the animal viruses seem to play as important a role in cell-mediated immunity as that of HIV-1, the p24 applicability as a candidate epitope for vaccine development could be evaluated in those animals.

Animals↗
PubMed1992

Cells surviving infection by human immunodeficiency virus type 1: vif or vpu mutants produce non-infectious or markedly less cytopathic viruses.

Under conditions in which a clonal cell line (M10) isolated from a human T cell lymphotrophic virus type I-transformed MT-4 cell line was completely killed by infection with wild-type human immunodeficiency virus type 1 (HIV-1), equivalent M10 cells survived infection with HIV-1 vif, vpr or vpu mutant virus after transient cytopathic effects. Several cell clones, which were isolated from the proliferating M10 cells after infection with vif and vpu mutant viruses (M10/vif- and M10/vpu-), had heterogeneous HIV-1 phenotypes in terms of HIV-1 antigen expression, their syncytium forming capacity, reverse transcriptase activity and the infectivity of HIV-1 particles produced. When the replication kinetics of the HIV-1 particles produced were assayed in M10 cells, the clones could be classified into three types, i.e. type I producing non-infectious HIV-1, type II producing infectious HIV-1 with low replicative ability and type III producing infectious HIV-1 with a replicative ability similar to that of wild-type HIV-1. HIV-1 major viral cell proteins and virus particle fractions were almost typical in types II and III but not in type I. Electron microscopic examination of particles released by I, II and III clones revealed rare defective, predominantly defective and essentially normal virions, respectively. Northern and Southern blot analyses revealed no apparent deletion in the proviral DNA and mRNA prepared from these clones, except in the case of type I and II clones isolated from M10/vpu- which contained large deletions in the mRNAs for gag and gag-pol proteins. Thus, M10 cells surviving infection with HIV-1 vif or vpu mutants are heterogeneous, persistently expressing HIV-1 antigens and producing non-infectious or less cytopathic virus.

Blotting, Northern↗
PubMed1992

Human immunodeficiency virus type 1 (HIV-1) superinfection of a cell clone converting it from production of defective to infectious HIV-1 is mediated predominantly by CD4 regions other than the major binding site for HIV-1 glycoproteins.

A cell clone, L-2, which produces non-infectious doughnut-shaped human immunodeficiency virus type 1 (HIV-1) particles, was permissive for HIV-1 superinfection, which resulted in the production of infectious particles. The superinfection showed slow kinetics compared with primary HIV-1 infection of M10 cells, the parent of the L-2 cell clone. Inhibition studies on the superinfection of L-2 cells using several CD4-related reagents showed that the CD4 molecule was an essential component of the receptor for superinfection. Strong inhibitory effects were obtained using CD4 peptides such as CD4(68-130), which includes a portion homologous to the immunoglobulin third complementarity-determining region (CDR3), as well as recombinant soluble CD4. In contrast, a CD4(45-60) peptide, which includes most of the CDR2-related region, was not effective, although the Leu-3a monoclonal antibody (MAb), which recognizes a site near the CDR2-related region, did slightly, but significantly, delay the superinfection kinetics. Comparative flow cytometry of L-2 and M10 cells revealed that the cell surface of L-2 cells despite expressing HIV-1 env protein, reacted slightly with OKT4 or anti-CD4(68-130) MAb, but not with Leu-3a or OKT4A MAb. In contrast, no reaction was detected with any of these anti-CD4 MAbs on the surface of another HIV-1 superinfection-resistant cell clone, MOLT-#8IIIB-14, which expresses HIV-1 env proteins but does not produce infectious HIV-1 particles. These results strongly suggest that expression of the CD4 major receptor site for primary HIV-1 infection is preferentially decreased on the surface of L-2 cells, but that the OKT4 epitope and the nearby region corresponding to immunoglobulin CDR3 remain exposed on the cell surface. Consequently, the CD4 CDR3-related region could play a major role as the receptor for the superinfection reported here.

Binding Sites↗
PubMed1992

Highly conserved epitope domain in major core protein p24 is structurally similar among human, simian and feline immunodeficiency viruses.

Linear B cell epitopes were mapped on the major core protein p24 of human immunodeficiency virus type 1 (HIV-1), simian immunodeficiency virus (SIVAGM) and feline immunodeficiency virus (FIV) using a fusion protein-based method and murine monoclonal antibodies reactive against the p24 antigens expressed on the surface of HIV-1- and FIV-infected cells. The results suggest that the sites identified here are encoded at similar positions in the three virus genomes and consist of highly conserved epitopes, which could exhibit immunodominance.

Amino Acid Sequence↗
PubMed1992

A unique monoclonal antibody that recognizes mature p17 of HIV-1 but not its precursor.

The entire and partial gag regions of human immunodeficiency virus type 1 (HIV-1) were overproduced in Escherichia coli and used for epitope mapping of antibodies against p17. We found that a mouse monoclonal antibody to p17, V17 recognizes the mature p17 but not the unprocessed Gag proteins containing the entire p17 moiety. Further analysis revealed that V17 recognizes the C-terminal 12-amino-acid region of p17 having free C-terminus. This monoclonal antibody may be useful for monitoring the maturation of virus particles.

Antibodies, Monoclonal↗
PubMed1992

Altered cell tropism and cytopathicity of feline immunodeficiency viruses in two different feline CD4-positive, CD8-negative cell lines.

Two interleukin-2-dependent feline CD4-positive and CD8-negative cell lines, MYA-1 and the newly established FeL-039, were used as host cells for infection with feline immunodeficiency virus (FIV). All FIV strains used, the Petaluma strain and several new isolates, were highly cytopathic to MYA-1. In contrast, the kinetics of FIV replication in FeL-039 differed greatly depending on the strain tested, i.e., noninfectious strain, highly cytopathic strain, and less cytopathic strain producing a persistent state for a long period. It appears, therefore, that cell tropism for FIV differed with each FIV strain tested even in T-cell lines showing similar cell surface phenotypes. Cytopathicity of FIV is evidently due to both the FIV strain and the host T cell.

Animals↗
PubMed1992

Lymphocyte development from stem cells.

Highly enriched pluripotent and multipotent hematopoietic stem cells (HSCs) are isolated from bone marrow and fetal liver as Thy-1loLin-Sca-1+ cells. Pluripotent HSCs express c-kit receptor on their surface, but the generation and proliferation of early fetal HSCs take place in the absence of steel factor. T precursor cells migrate into the fetal thymus by chemotactic mechanism. CD4lo precursors represent a newly defined phase of T-cell development in the thymus between the bone marrow-derived stem cells and the CD4-8- intrathymic precursors. Only fetal, but not adult, HSCs have the capacity to differentiate into V gamma 3+ and V gamma 4+ T cells under the fetal thymic microenvironment, and HSC themselves may lose some of their developmental potential during ontogeny. It is postulated that HSCs are the locus of a complicated but precise developmental clock that may determine both the time-dependent closure of some gene loci (e.g. V gamma 3 and V gamma 4 T cell receptor, and embryonic and fetal globin) and the activation of others (e.g. the N nucleotide insertion machinery).

Animals↗
PubMed1992

[Polycythemia vera terminating in chronic neutrophilic leukemia: studies on in vitro growth of hematopoietic progenitor cells].

A 57-year-old man, diagnosed as Polycythemia vera (PV), had been treated with administrations of Busulfan since 1984. Three years later, the number of neutrophils in peripheral blood increased to 50,000/microliters with progression of splenomegaly, and the case was diagnosed as Chronic neutrophilic leukemia (CNL) based on the criteria by Miura et al, in November, 1989. In spite of 6MP and Busulfan therapy, marked neutrophilia and splenomegaly progressed, and the patient died due to liver dysfunction in June of 1991. To clarify the pathophysiology of PV and CNL, we studied the in vitro growth kinetics of hematopoietic progenitor cells in bone marrow of this unique case and made a comparison with those of 4 cases of PV and 4 normal volunteers employing methylcellulose culture. As in other cases of PV, erythroid colonies were formed in culture of bone marrow from this patient without addition of erythropoietin. Furthermore, spontaneous colonies derived from CFU-GM and CFU-Mix increased remarkably in this case only. The results suggest that the hematopoietic abnormalities in this case involve the multipotent stem cells as well as erythroid and granuloid-macrophage progenitors.

Colony-Forming Units Assay↗
PubMed1992

[Recurrent pulmonary edema in a patient with acute lymphoblastic leukemia after syngeneic bone marrow transplantation].

Severe respiratory distress appeared in a 14-year-old girl with acute lymphoblastic leukemia 2 months after receiving syngeneic bone marrow transplantation (BMT) with a conditioning regimen of a high-dose of busulfan, etoposide and nimustine. Rapid body-weight gain and edema, especially in eyelids and lower-limbs, were also observed. Without any findings of heart failure nor GVHD, pulmonary edema was recognized on the chest roentgenogram. As soon as the diagnosis of pulmonary edema due to 'capillary leak syndrome' was suspected, the patient was treated with intravenous administration of diuretics, albumin and bolus methylprednisolone in combination of mechanical ventilation. Although the clinical manifestations were improved by the treatment, the disease recurred 5 weeks later. The patient was successfully treated by the same medications, and there has been no recurrence as of the sixth month after discontinuance of the therapy. At present, the mechanism of capillary leak syndrome is still undefined. In this case, however, we speculate that the conditioning regimen for BMT intensified the capillary disturbance initially caused by intensive chemotherapy since remission induction. Furthermore hypoalbuminemia due to severe anorexia might have enhanced the occurrence of the disease.

Adolescent↗
PubMed1991

The junctional modifications of a T cell receptor gamma chain are determined at the level of thymic precursors.

T precursors from fetal liver and adult bone marrow were compared for their ability to give rise to V gamma 4+ T cell development. Fetal thymic lobes were repopulated with fetal liver or adult bone marrow cells, and the organ-cultured thymocytes were analyzed for their T cell receptor expression by the polymerase chain reaction (PCR). Both day 14 fetal liver and adult bone marrow cells gave rise to thymocytes with V gamma 4-J gamma 1 transcripts. However, the average size of the PCR products derived from adult precursors was slightly larger than that from fetal precursors. DNA sequence analysis of the V gamma 4-J gamma 1 transcripts showed that early fetal liver precursors predominantly gave rise to thymocytes with the V gamma 4-J gamma 1 transcripts without N nucleotide insertion, while late fetal liver and adult marrow precursors predominantly gave rise to thymocytes with modified V gamma 4-J gamma 1 junctions. These results suggest the possibility that the level of the N nucleotide insertion is programmed at the level of thymic precursors. This study also supported the model presented previously that the developmental potential of hematopoietic stem cells may change during ontogeny.

Animals↗
PubMed1991

Mouse hematopoietic stem cells and the interaction of c-kit receptor and steel factor.

Hematopoietic stem cells (HSCs) are distinguished from other hematopoietic progenitors in bone marrow by their unique ability to undergo multilineage differentiation and self-renewal. Two mouse mutations, dominant spotting (W) and steel (Sl), have pleiotropic effects on hematopoiesis, gametogenesis, and melanoblast development. These two mutations have been shown to be intrinsic (W) and microenvironmental (Sl) defects. Recently, molecular studies revealed that the W and Sl loci encode the c-kit receptor and steel factor (SLF), respectively. The c-kit receptor is expressed on HSCs and hematopoietic progenitors, while SLF is produced by stromal cells. SLF acts on hematopoietic progenitors synergistically with other growth factors. Here we review the effect of these mutations on mouse hematopoiesis, and show that SLF acts on HSCs and other myeloerythroid progenitors, but that it, in our hands, does not play a critical role in HSC generation or self-renewal. Rather, SLF is the most potent co-mitogen (with IL-1, IL-3, IL-6, G-CSF, GM-CSF, or M-CSF) found that acts on these cells, but the effect of such treatments is the rather specific and massive expansion of myeloerythropoiesis, not lymphopoiesis, and perhaps at the expense of HSC self-renewal.

Animals↗