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K Ohtsuka

Publications and source records attributed to K Ohtsuka.

At least 19 recordsLinked to original sources
PubMed1995

Relationships between intermediate TCR cells and NK1.1+ T cells in various immune organs. NK1.1+ T cells are present within a population of intermediate TCR cells.

Experiments to date have revealed a population of T cells that carry intermediate (int) levels of TCR (or CD3) and express IL-2R beta-chain (IL-2R beta) in mouse liver. Such int TCR cells also reside in other immune organs, although in low numbers. On the other hand, NK1.1+ T cells with int TCR do reside in the thymus and other peripheral organs. To determine the relationship of two types of cells, we characterized int CD3 cells and NK1.1+ T cells throughout the organs in terms of the phenotype, V beta repertoire, and morphology. Although both IL-2R beta+ T cells and NK1.1+ T cells are classified as int CD3 cells, NK1.1+ T cells are present within int CD3 cells. The majority of int CD3 cells in the liver and thymus were NK1.1+, whereas the minority of such cells in the spleen, lymph nodes, and bone marrow were NK1.1+. Among int CD3 cells, double-negative (DN) CD4-8- cells and/or CD4+ were abundant in NK1.1+ subset, whereas CD8+ cells were generally abundant in NK1.1- subset. Self-reactive V beta+ clones estimated by the M1s system were distributed to both NK1.1+ and NK1.1- subsets. High CD3 cells in the thymus and other organs contained neither DN cells nor forbidden clones. Int CD3 cells had the morphology of granular or agranular lymphocytes carrying perforin. Among int CD3 cells, NK1.1+ subset had a higher level of perforin-positive cells than NK1.1- subset. These results clearly demonstrate the relationship between int TCR cells and NK1.1+ T cells in various organs.

Animals↗
PubMed1995

Evidence for extrathymic generation of intermediate T cell receptor cells in the liver revealed in thymectomized, irradiated mice subjected to bone marrow transplantation.

In addition to the major intrathymic pathway of T cell differentiation, extrathymic pathways of such differentiation have been shown to exist in the liver and intestine. In particular, hepatic T cells of T cell receptors or CD3 of intermediate levels (i.e., intermediate T cell receptor cells) always contain self-reactive clones and sometimes appear at other sites, including the target tissues in autoimmune diseases and the tumor sites in malignancies. To prove their extrathymic origin and self reactivity, in this study we used thymectomized, irradiated (B6 x C3H/He) F1 mice subjected to transplantation of bone marrow cells of B6 mice. It was clearly demonstrated that all T cells generated under athymic conditions in the peripheral immune organs are intermediate CD3 cells. In the case of nonthymectomized irradiated mice, not only intermediate CD3 cells but also high CD3 cells were generated. Phenotypic characterization showed that newly generated intermediate CD3 cells were unique (e.g., interleukin 2 receptor alpha-/beta+ and CD44+ L-selectin-) and were, therefore, distinguishable from thymus-derived T cells. The precursor cells of intermediate CD3 cells in the bone marrow were Thy-1+ CD3-. The extrathymic generation of intermediate CD3 cells was confirmed in other combinations of bone marrow transplantation, C3H --> C3H and B10.Thy1.1 --> B6.Thy1.2. The generated intermediate CD3 cells in the liver contained high levels of self-reactive clones estimated by anti-V beta monoclonal antibodies in conjunction with the endogenous superantigen minor lymphocyte-stimulating system, especially the combination of B6 --> (B6 x C3H/He) (graft-versus-host-situation).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Transcriptional inhibition of insulin by FK506 and possible involvement of FK506 binding protein-12 in pancreatic beta-cell.

FK506 (tacrolimus) is a strong immunosuppressant: it has been approved as a drug for liver transplantation in Japan, the United States, and the United Kingdom. One of its main adverse effects is hyperglycemia. Thus, in this study, we investigated the mechanism and the reversibility of the hyperglycemia caused by FK506. FK506 did not affect the glucose uptake by insulin into rat strio-muscle cell line, but suppressed insulin production in rat insulinoma cells. Two-week oral administration of FK506 at 10 mg/kg/day suppressed insulin production time-dependently at the transcriptional step in pancreatic beta-cells, while glucagon content in pancreatic alpha-cells was not affected. When FK506 administration was stopped in these rats, insulin mRNA transcription and insulin production returned to normal. This recovery indicates that the adverse effect of FK506 on the pancreas is reversible. A high content of FK506 binding protein-12 (FKBP-12) in the pancreatic beta-cells was confirmed by immunostaining with anti-human FKBP-12 mAb, but the content was less in the pancreatic alpha-cells and almost negligible in the acinar cells. In contrast, a high content of calcineurin in the pancreatic alpha-cells was confirmed by using anti-calcineurin polyclonal antibody, but this content was less in the pancreatic beta-cells and not found in the acinar cells. Thus, as in the case with NF-AT in T cells, these findings point to the reduction of unidentified nuclear factors for insulin mRNA transcription caused by the binding of FK506 to FKBP-12 and a subsequent inhibition of calcineurin in the beta-cells.

Animals↗
PubMed1995

Interaction between hsp70 and hsp40, eukaryotic homologues of DnaK and DnaJ, in human cells expressing mutant-type p53.

We have recently identified a novel 40-kDa heatshock protein hsp40 as a mammalian homologue of bacterial DnaJ protein. Here we demonstrate the physical interaction between hsp70 (DnaK homologue) and hsp40 in human cells as determined by immunoprecipitation methods. Co-immunoprecipitation of hsp70 with hsp40 was dependent on the presence of ATP or unfolded protein (reduced carboxymethylated alpha-lactalbumin). A mutant type of tumor suppressor gene product, mtp53, was co-immunoprecipitated not only with hsp70 but also with hsp40. These results suggest the existence of a hsp70(DnaK)/hsp40(DnaJ) chaperone system in mammalian cells.

Escherichia coli Proteins↗
PubMed1995

Adhesion molecules on intermediate TCR cells. II. Hepatoprotective effects of hyaluronic acid on acute liver injury.

The liver is a major organ wherein extrathymic T cells and NK cells exist in mice. Due to their unique properties, i.e., extrathymic T cells are TCR (or CD3)-intermediate+ IL-2R beta+ (herein termed intermediate TCR cells) and NK cells are TCR(-)IL-2R beta+, they are easily distinguished from the other lymphocyte subsets by using mAbs in conjunction with immunofluorescence tests. They were recently found to express a higher level of CD44 antigen, which is a ligand for hyaluronic acid, than that of another T cell subset (i.e., thymus-derived T cells or bright TCR cells). Since an intravenous administration of hyaluronic acid was also found to reduce the number of intermediate TCR cells and NK cells in the liver, we examined whether hyaluronic acid had a hepatoprotective effect on acute liver injury. Such injury was induced by LPS injection in mice pretreated with Propionibacterium acnes 1 week earlier. When a single dose of hyaluronic acid was given to these mice 12 hr before LPS injection, a prominent hepatoprotective effect was observed in terms of decreases of mortality (up to 50%), lymphocyte infiltration of the liver, serum transaminase levels, and tissue damage. At this time, liver mononuclear cells isolated from the treated mice showed decreased levels of cytokine production such as TNF and IL-1. These results reveal that intermediate TCR cells and NK cells in the liver actually adhere the sinusoid walls by means of an interaction of CD44 molecules and hyaluronic acid even in the case of acute liver injury. It suggests a possible therapeutic effect of the administration of hyaluronic acid in acute liver injury by eliminating the effector cells and cytokine-producing cells from the liver.

Adjuvants, Immunologic↗
PubMed1995

Intermediate TCR cells with self-reactive clones are effector cells which induce syngeneic graft-versus-host disease in mice.

It has been established that, even after syngeneic bone marrow transplantation, animals treated with immunosuppressive drugs may suffer from graft-versus-host disease, showing autoimmune-like symptoms. Although the major effector cells are known to be T-cell subsets, detailed characterization of such T cells remains to be investigated. In the present study, we characterized them, especially as to whether they are thymus-derived T cells or extrathymic T cells, and how self-reactive clones were distributed among the above T-cell subsets. BALB/c mice (Mls-1b2a) were irradiated (9 Gy), subjected to bone marrow transplantation, and then treated with cyclosporin A (CsA) for 6 weeks. From 2 weeks after the cessation of CsA, these mice displayed signs of GVH disease. The major target organs included the liver and colon. Two-color staining for CD3 and IL-2R beta was applied to identify CD3-IL-2R beta+ NK cells, CD3-intermediate +IL-2R beta+ cells (i.e., intermediate CD3 or TCR cells of extrathymic origin) and CD3-high+IL-2R beta- cells (i.e., high CD3 cells of thymic origin). It was demonstrated that the major expanding lymphocytes were intermediate TCR cells and that self-reactive clones (V beta 3+ and V beta 11+ cells in this strain of mice) were confined to this population. Interestingly, these self-reactive clones had ability to respond to immobilized anti-V beta 3 and anti-V beta 11 mAbs. Liver MNC in mice with GVH disease which contained the highest proportion of intermediate TCR cells were able to mediate the adoptive transfer of GVH disease to other irradiated (6.5 Gy) mice. Intermediate TCR cells also showed potent cytotoxic activity against syngeneic leukemia cells. These results suggest that intermediate TCR cells are the major effector cells for the induction of syngeneic GVH disease.

Animals↗
PubMed1995

Transcranial magnetic stimulation over the posterior cerebellum during visually guided saccades in man.

Recent intensive neurophysiological experiments in the monkey have demonstrated the function of the cerebellum in the control of saccadic eye movements. Microstimulation studies on monkeys have determined that the cerebellar cortex, which is specifically involved in the control of saccades, is located in vermal lobules VIc and VII. The Purkinje-cell axons arising from this vermal area terminate almost exclusively in an ellipsoidal region which protrudes dorsocaudally from the fastigial nucleus. Saccade-related cells in the fastigial nucleus are located exclusively in the ellipsoidal region. Microstimulation of the vermis modulated the activity of saccade-related cells in the fastigial nucleus, and then produced dysmetric saccades. In this study, we investigated effects of cerebellar stimulation on saccade metrics in man using a transcranial magnetic stimulation (TMS) device. Focal TMS was applied over the posterior cerebellum at an area approximately 7 mm lateral and caudal to the inion during horizontal visually guided saccades in six normal subjects. The TMS device was triggered after the onset of saccades with a latency of 0, 20, 40 or 60 ms. We investigated the effect of TMS on the amplitude and velocity of saccadic eye movements. For visually guided saccades directed contralateral to the stimulation side, TMS of the posterior cerebellum with the latency of 0 ms produced hypometric saccades followed by corrective saccades. Transcranial magnetic stimulation with latencies of 20, 40 and 60 ms had no effect on saccade metrics. On the other hand, for ipsilateral saccades, TMS with latencies of 0, 20 and 40 ms produced hypermetric saccades followed by postsaccadic drift.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1995

Role of heat shock cognate 70 protein in import of ornithine transcarbamylase precursor into mammalian mitochondria.

The roles of the 70-kDa cytosolic heat shock protein (hsp70) in import of precursor proteins into the mitochondria were postulated to be related to (i) unfolding of precursor proteins in the cytosol, (ii) maintenance of the import-competent state, and (iii) unfolding and transport of precursor proteins through contact sites, in cooperation with matrix hsp70. We examined roles of cytosolic hsp70 family members in import of ornithine transcarbamylase precursor (pOTC) into rat liver mitochondria, using an in vitro import system and antibodies against hsp70. Immunoblot analysis using an hsc70 (70-kDa heat shock cognate protein)-specific monoclonal antibody and a polyclonal antibody that reacts with both hsc70 and hsp70 showed that hsc70 is the only or major form of hsp70 family members in the rabbit reticulocyte lysate. The hsc70 antibody did not inhibit pOTC import when added prior to import assay. However, when pOTC was synthesized in the presence of the antibody and then subjected to import assay, pOTC import was markedly decreased. pOTC import was also decreased when the precursor was synthesized in the lysate depleted for hsc70 by treatment with hsc70 antibody-conjugated Sepharose. This reduction was almost completely restored by readdition of purified mouse hsc70 during pOTC synthesis. The readdition of hsc70 after pOTC synthesis and only during the import assay was not effective. Thus, once import competence of pOTC was lost, hsc70 was ineffective for restoration. Newly synthesized pOTC lost import competence in the absence of hsc70 somewhat more rapidly than in its presence. These results indicate that hsc70 is required during pOTC synthesis and not during import into the mitochondria. hsc70 presumably binds to pOTC polypeptide and maintains it in an import-competent form.

Animals↗
PubMed1995

Cotranslocation and colocalization of hsp40 (DnaJ) with hsp70 (DnaK) in mammalian cells.

A novel 40-kDa heat-shock protein hsp 40 in mammalian cells has been recently identified to be a homolog of bacterial DnaJ protein. We have previously shown the colocalization of hsc70 (p73, constitutive form) with hsp40 in the nucleoli of heat-shocked HeLa cells. In this report we further investigated intracellular translocation and localization of hsp40 and hsp70 (both constitutive p73 and inducible p72) in several mammalian cells. Translocation kinetics of hsp40 during heating at mild temperature were almost the same as those of hsp70 in HeLa cells. Hsp40 colocalized not only with hsc70 (p73) but also with hsp70 (p72) in heat-shocked HeLa (human), HA-1 (Chinese hamster) and NRK (rat) cells. Direct interaction of hsp40 with hsp70 (p73 and/or p72) was observed in all cells tested by immunoprecipitation methods. Also, treatments of cells with cytoskeleton-acting drugs such as cytochalasin E, colchicine and taxol had no effect on the heat-induced translocation of hsc70 (p73) and hsp40 in NRK cells. These results strongly suggest that hsp40 and hsp70 (p73/p72) form a complex in the cytoplasm at normal temperature, translocate together and colocalize in the nuclei and nucleoli upon heat-shock, and that they may function cooperatively to repair (refold) denatured proteins under stress conditions.

Animals↗
PubMed1995

Adhesion molecules on intermediate TCR cells. I. Unique expression of adhesion molecules, CD44+ L-selectin-, on intermediate TCR cells in the liver and the modulation of their adhesion by hyaluronic acid.

In addition to thymus-derived T cells, it was demonstrated recently that extrathymically differentiated T cells exist in the liver and other immune organs of mice. Since such extrathymic T cells have T-cell receptors (TCR) of intermediate intensity (i.e. intermediate TCR cells) and constitutively express IL-2 receptor beta-chain (IL-2R beta) similar to natural killer (NK) cells, they are easily distinguished from thymus-derived T cells with a TCR-bright+ IL-2R beta- phenotype (i.e. bright TCR cells). In the present study, the expression of adhesion molecules CD44 and L-selectin was compared between these T-cell subsets. Intermediate TCR cells in the liver and other organs were found to be CD44+ L-selectin- and, inversely, bright TCR cells were CD44- L-selectin+. CD3- IL-2R beta+ NK cells were also estimated to be CD44+ L-selectin-. Hyaluronic acid, which is known to adhere to a CD44 ligand, bound to intermediate TCR cells, but not to bright TCR cells. Among many extracellular matrices, hyaluronic acid induced a prominent decrease in the numbers and proportions of intermediate TCR cells and NK cells in the liver from 6 to 24 hr after in vivo administration. The half-life span of injected hyaluronic acid was approximately 7 hr in the plasma. These results suggest that the CD44 molecule, which is uniquely expressed on intermediate TCR cells and NK cells, is eventually associated with their adhesion to the sinusoidal walls in the liver.

Animals↗
PubMed1995

Transcranial magnetic stimulation of the posterior parietal cortex degrades accuracy of memory-guided saccades in humans.

PURPOSE: Previous neurophysiological studies in the monkey have shown that the posterior parietal cortex (PPC) is involved in the control of memory-guided saccades. In this study, the authors investigated the effect of focal transcranial magnetic stimulation (TMS) over the posterior parietal cortex on accuracy of memory-guided saccades in humans to clarify the cortical region related to controlling memory-guided saccades. METHODS: Single TMS pulses were applied systematically at various locations over the parietal cortex during memory-guided saccades in human subjects. Each subject was instructed to fixate on a central target until the target was extinguished. A lateral target was flashed for 100 msec with unpredictable direction and amplitude. The subject executed a saccade to the remembered position of the previously flashed target in total darkness when the central target was extinguished. Transcranial magnetic stimulation was applied after the extinction of the central target with various latencies. Stimulation sites were identified by three-dimensional magnetic resonance imaging (MRI). RESULTS: Transcranial magnetic stimulation of the right PPC degraded accuracy of rightward and leftward memory-guided saccades in all subjects, when TMS was applied 100 msec after the offset of the central target. On the other hand, TMS of the left PPC showed no effects on accuracy of memory-guided saccades. Systematic error of saccade amplitude appeared as a hypometria for rightward saccades. Three-dimensional MRI revealed that the effective stimulation site was located over the posterior portion of the intraparietal sulcus and the superior part of the angular gyrus of the right hemisphere. CONCLUSIONS: These findings strongly suggest that the right PPC is involved in maintaining spatial accuracy of remembered target locations of memory-guided saccades in humans.

Adult↗
PubMed1994

A similar expression pattern of adhesion molecules between intermediate TCR cells in the liver and intraepithelial lymphocytes in the intestine.

Two major populations of extrathymically differentiated T cells exist in the liver and intestine. Such T cells in the liver have TCR of intermediate intensity (i.e., intermediate TCR cells) and constitutively express IL-2 receptor beta-chain (IL-2R beta), whereas those in the intestine, especially intraepithelial lymphocytes, have TCR of bright intensity, consisting of a mixture of IL-2R beta+ and IL-2R beta-. All mature thymocytes and thymus-derived T cells seen in the peripheral immune organs are TCR-bright+ IL-2R beta- under resting conditions. When the expression pattern of adhesion molecules, including CD44, L-selectin, LFA-1 and ICAM-1, was compared among these T-cell populations, they displayed quite unique patterns of expression. All extrathymic T cells in the liver, intestine, and even other organs were CD44+ L-selectin-LFA-1++ICAM-1+, whereas thymocytes and thymus-derived T cells were CD44-L-selectin+LFA-1+ICAM-1-. This inverted expression of adhesion molecules between extrathymic T cells and thymus-derived T cells might be associated with their unique tissue-localization.

Animals↗
PubMed1994

The induction of interleukin-6 (IL-6) and colony-stimulating factors (CSFs) by FK565 and its thrombopoietic activity following in vivo administration.

The induction of macrophage colony-stimulating factor (M-CSF) in monkey plasma following administration of FK565 was observed within 2 h of injection peaked at 4 h, and remained high after 24 h. Interleukin-6 (IL-6) and M-CSF levels increased in monkeys treated with FK565, even at doses as low as 0.01 mg/kg. Granulocyte CSF (G-CSF) levels increased slightly following a dose of 1 mg/kg, but granulocyte macrophage CSF (GM-CSF) was not detected at any doses of FK565 studied. To examine the thrombopoietic activity of FK565 in vivo, single doses of drug (0.01, 0.1 or 1.0 mg/kg) were administered i.v. to cynomolgus monkeys or normal mice on day 0. The promotes platelet (PLT) count after FK565 injection decreased transiently on days 1 and 2, and then increased in a dose-dependent manner on day 5 and was still high on day 14. The experiment using anti-PLT antibody showed that the increased PLT count was not simply due to a rebound phenomenon after the transient decrease in PLT. The effect of i.v. FK565 was studied in mice myelosuppressed with a single dose of mitomycin C (MMC) (5.6 mg/kg). The fall in PLT count was suppressed on day 7 by 0.1 and 1.0 mg/kg FK565. Although intact cells or tissues are necessary for an increase in PLT following FK565 treatment, FK565 suppressed the impaired hematopoietic function seen after chemotherapy. FK565 is proposed as a drug to restore reduced neutrophil and platelet counts found in AIDS or cancer therapy.

Animals↗
PubMed1993

Afferent and efferent connections of the cortical accommodation area in the cat.

Accommodative responses were evoked by microstimulation of a circumscribed area in the lateral suprasylvian (LS) cortex of the cat. We studied anatomical connections of this area with WGA-HRP. We identified an accommodation-related area by systematic microstimulation of the LS cortex in each of nine cats. Accommodative and pupillary responses were monitored by an infrared optometer and a pupillometer, respectively. WGA-HRP was injected into the accommodation-related area where accommodative responses were elicited with low-intensity microstimulation, but pupillary responses were not evoked. Retrogradely labeled cells were found mainly in ipsilateral areas 17, 18 and 19, the pulvinar, the lateral posterior nucleus of the thalamus (LP) and the contralateral LS area. Fewer labeled cells were found in ipsilateral areas 20 and 21, the ventral lateral suprasylvian area (VLS), the splenial visual area (SVA) and the lateral geniculate nucleus (LGN). Anterogradely labeled terminals were located mainly in the ipsilateral LP, the rostral portion of the pontine nuclei and the superficial layers of the ipsilateral superior colliculus which corresponds to the representation of the central visual field. Less dense labeled terminals were also found in the ipsilateral nucleus of the optic tract (NOT) in two cats.

Accommodation, Ocular↗
PubMed1992

Details of an isolation method for hepatic lymphocytes in mice.

The liver comprises a unique lymphocyte population, i.e., extrathymic alpha beta T cells with TcR of intermediate intensity. In the present study, we attempted to determine what pretreatments were appropriate to isolate hepatic mononuclear cells (MNC) containing such intermediate alpha beta TcR cells in mice. Hepatic MNC were isolated from untreated mice and mice subjected to either bleeding or liver perfusion, and the intermediate alpha beta TcR cells in each preparation were identified. For reasons of simplicity, cell purity and cell yields, hepatic lymphocytes should be obtained from mice subjected to total bleeding. Additional information on extrathymic alpha beta T cells obtained by using the recommended method is also presented.

Animals↗
PubMed1992

The relationship between hsp 70 localization and heat resistance.

Using indirect immunofluorescence we have investigated the kinetics of nuclear accumulation and removal of hsp 70 in HA-1 Chinese hamster fibroblasts exposed to elevated temperatures. The kinetics of accumulation of hsp 70 in the nuclei were found to be time/temperature dependent at all temperatures tested (42-45 degrees C). At a given temperature, the fraction of cells manifesting nuclear localization of hsp 70 increased with exposure time. For a given duration of heating, the fraction of cells manifesting nuclear localization of hsp 70 increased with the temperature. The kinetics of the nuclear accumulation of hsp 70 were similar for normal HA-1 cells, their heat-resistant variants, and transiently thermotolerant cells (triggered by prior exposure to a brief heat shock or to sodium arsenite). Upon return to 37 degrees C after heat shock, the kinetics of removal of the hsp 70 associated with the nucleus was dependent on the severity of the initial heat challenge. However, for a given heat dose, the decay of nuclear localization of hsp 70 was more rapid in thermotolerant and heat-resistant cells than in their normal counterparts. These results suggest that the increased levels of hsp 70 associated with the transient or permanently heat-resistant state may play a direct role in restoring and/or repairing heat-induced nuclear and nucleolar alterations associated with heat-induced cell killing. Furthermore, they also suggest that the heat-resistant state may involve ameliorated repair of heat-induced cellular alterations.

Animals↗