Biomedical subjects

U Yamashita

Publications and source records attributed to U Yamashita.

At least 19 recordsLinked to original sources

PubMed1999

Interleukin-1alpha regulates G1 cell cycle progression and arrest in thyroid carcinoma cell lines NIM1 and NPA.

This study provides the first report that the same cytokine (interleukin-1 (IL-1)) can induce opposite effects on cyclin-dependent kinases (Cdks) and Cdk inhibitors (Cdkis) in the G1 phase even in the same type of cancer cells (papillary thyroid carcinoma cells). Cell cycle analysis revealed an increase in NIM1 cells and a decrease in NPA cells in the S and G2+M phases after treatment with IL-1alpha. The addition of IL-1alpha to NIM1 cells reduced the expression of p16 and p21 protein and induced the expression of Cdk2 and Cdk4 protein, which leads to the phosphorylation of retinoblastoma protein. The addition of IL-1alpha to NPA cells induced the expression of p27 protein and reduced the expression of Cdk2 protein, which leads to induction of p107 protein expression. It is of interest that p21 protein expression was not observed in NPA cells. These results suggest that several Cdks and Cdkis play a regulatory role in the G1 cell cycle progression and arrest induced by IL-1alpha in thyroid carcinoma cell lines.

Blotting, Western↗

PubMed1998

[Antigen-recognition mechanism of T cells].

The antigen-recognition mechanism of T cells was reviewed from the standpoint of MHC-restriction. T cells recognize antigens presented by antigen-presenting cells in a MHC-restricted manner. CD4+ helper T cells recognize antigens with the context of MHC class II molecules, and CD8+ cytotoxic T cells recognize antigens with MHC class I molecules. The MHC class I molecule is composed of alpha-chain and beta 2-microglobulin, and MHC class II molecule is composed of alpha-chain and beta-chain. The T cell antigen receptor is composed of alpha-chain and beta-chain, or gamma-chain and delta-chain, and CD3 complex. Antigens are processed and presented by antigen-presenting cells to T cells in the context of MHC molecules. Furthermore, costimulatory factors such as adhesion molecules and cytokines are required for the full activation of T cells. The biological significance of the T cell antigen recognition mechanism is discussed from the standpoint of clinical application.

Antigen-Presenting Cells↗

PubMed1998

Stimulatory effect of lymphocyte-derived factor on catecholamine efflux from cultured bovine adrenal medullary cells.

The effects of lymphocytes and their conditioned medium on catecholamine efflux and uptake were examined in cultured bovine adrenal medullary cells. Co-culture of adrenal medullary cells with lymphocytes for 3 days caused an increase in appearance of catecholamines in the culture medium. Treatment of adrenal medullary cells with a conditioned medium prepared from lymphocytes also enhanced the appearance of catecholamines in culture medium in time- (8-48 h) and concentration-dependent manners. Heat treatment of the conditioned medium at 60 and 100 degrees C for 10 min reduced its stimulatory effect to 59 and 20% of control, respectively. After gel filtration on a Sephadex G-25 column or dialysis (<8 kDa molecular mass cutoff), the stimulatory activity of the conditioned medium was found in a high molecular fraction. The conditioned medium had little effect on the activity of lactate dehydrogenase in the medium of cultured adrenal medullary cells and on desipramine-sensitive [3H]norepinephrine uptake by the cells. These findings suggest that lymphocytes release a heat-sensitive factor(s) (molecular mass of more than 8 kDa) which increases efflux of catecholamines from cultured adrenal medullary cells.

Adrenal Medulla↗

PubMed1998

Induction of expression of MHC-class-II antigen on human thyroid carcinoma by wild-type p53.

Mutation of the tumor-suppressor gene p53 is involved in carcinogenetics. We investigated the role of p53 in the induction of anti-tumor immune responses by establishing a thyroid carcinoma cell line (1F3) prepared by transfection of wild-type human p53 gene into a p53-deficient cell line (FRO). Our results showed for the first time the involvement of p53 in the induction of anti-tumor immune responses, as demonstrated by: (i) expression of the major-histocompatibility-complex(MHC)-class-II antigen on 1F3, but not FRO; (ii) mRNA of class-II gene was expressed both in 1F3 and in FRO, but was stable at post-transcriptional level in FRO, which restrained protein synthesis; (iii) 1F3 induced MHC-class-II-specific CD4+ cytotoxic-T-cell activity through allo-antigen presentation and co-stimulation. Although our novel results are limited to the wild-type-p53-expressing clone from a p53-deficient cell line, we suggest that the absence of p53 in carcinoma cells may reduce the induction of CD4+ cytotoxic-T-cell activity against carcinoma cells by diminishing the expression of class-II antigen.

Blotting, Western↗

PubMed1998

Effects of retinoic acid and TGF-beta 1 on the proliferation and melanin synthesis in chick retinal pigment epithelial cells in vitro.

PURPOSE: The purpose of this study was to investigate the effects of all-trans retinoic acid (RA) and transforming growth factor-beta 1 (TGF-beta 1) on proliferation and melanin synthesis of chick retinal pigment epithelial (RPE) cells in vitro. METHODS: Chick RPE cells were cultured in the presence or absence of RA (10(-5) to 10(-7) M) or TGF-beta 1 (1 to 100 U/ml) for a 10-day period. The effects of RA and TGF-beta 1 on RPE cell proliferation were assessed by counting the number of cells, and their effects on melanin synthesis were evaluated by measuring the melanin content in the cells. RESULTS: RA significantly inhibited RPE cell proliferation in a time-dependent manner. The most marked inhibition was observed on day 10 of culture, and the most effective concentration of RA was 10(-6) M. Melanin synthesis was increased on day 10, and the most effective concentration of RA was also 10(-6) M. TGF-beta 1 also inhibited RPE cell proliferation and increased melanin synthesis. CONCLUSIONS: These findings indicate that RA and TGF-beta 1 have regulatory effects on the proliferation and differentiation of chick RPE cells in vitro.

Animals↗

PubMed1998

Five monoclonal antibodies against glycophorin A of human erythrocyte recognize glycoprotein of bovine erythrocyte.

To study heterophile blood antigens on erythrocytes between human and experimental or domestic animals, we have produced 295 monoclonal antibodies (MAbs) to human erythrocyte membrane protein. According to the affinity, reactivity, and titre of the MAbs, we selected 40 clones to study the heterophile blood antigens between human and bovine, chicken, guinea pig, horse, rabbit, sheep, and swine. Five MAbs commonly reacted with human type A, type B, and type O erythrocytes and reacted with bovine erythrocytes as well but did not react with erythrocytes from other species. Other MAbs did not react with erythrocytes from all the tested animals. These five MAbs reacted with the same erythrocyte membrane protein, 90 KD glycophorin A (GPA) of human or 200 KD major glycoprotein and other two components of bovine by immunoblotting and GPA competitive inhibition assay. Furthermore, by enzyme treatment and monosaccharide competitive inhibition assay, it was confirmed that these five MAbs recognized antigen epitope of glycosylation free amino acid portion but not glycosylation portion of GPA of erythrocyte membrane.

ABO Blood-Group System↗

PubMed1997

Staphylococcal enterotoxin B induces arthritis in female DBA/1 mice but fails to induce activation of type II collagen-reactive lymphocytes.

It has been proposed that superantigens are involved in the pathogenesis of autoimmune diseases. To test the possibility of superantigens inducing arthritis in naive mice, V beta 8-reactive superantigen staphylococcal enterotoxin B (SEB) was injected into naive mice. We used female DBA/1 mice, because they were susceptible to collagen-induced arthritis (CIA), in which the pathogenic T cells were supposed to preferentially use limited V betas of T cell receptors including V beta 8. Mild monoarthritis developed in uninjected hindlimbs of mice administered with SEB in higher frequency (an average incidence of 24%) than the control phosphate-buffered saline-injected mice (4.2%). Autoimmune responses in mice administered with SEB were compared with those in mice developing CIA. However, activation of type II collagen (IIC)-reactive T cells was not detected in SEB-injected mice. Production of autoantibodies, anti-IIC antibody and rheumatoid factor was also undetected. Although exact mechanisms of pathogenesis of this arthritis remain to be known, V beta 8+ T cells were activated for a long period and the unresponsiveness of V beta 8+ T cells was not detected in this strain. From these results, we discuss the pathogenesis of arthritis induced by SEB and the possibility that superantigen may play a role in the induction of autoimmune diseases.

Animals↗

PubMed1997

Cytokine- and neuropeptide-mediated differentiation in retinal pigment epithelial cells in vitro.

To determine the mechanism of growth and differentiation of retinal pigment epithelial (RPE) cells it is important to understand the pathogenesis of several retinal diseases. Recently it has been reported that several cytokines and neuropeptides regulate the growth of RPE cells. In this study, the role of cytokines and neuropeptides in melanin synthesis, which is one indication of the RPE cell differentiation, was examined using chick RPE cells in vitro IL-1beta, TNF-alpha, substance P, beta-endorphin and methionine-enkephalin stimulated the melanin synthesis of RPE cells in a dose-dependent manner. The most effective concentrations of these agents on RPE cell melanin synthesis were not the same as that for RPE cell proliferation. These results indicate that cytokines and neuropeptides play an important role not only for the growth but also for the differentiation of RPE cells.

Animals↗

PubMed1997

B cell stimulating activity of seaweed extracts.

The activity of seaweed extracts on murine and human lymphocytes was studied in vitro. The extracts of some kind of seaweed, such as Hizikia fusiformis and Meristotheca papulosa, stimulated normal mouse spleen cells to proliferate. The responder cells are B cells, because the response was depleted by the treatment of spleen cells with anti-immunoglobulin (Ig) antibody and complement and being passed through a nylon wool column. This response is not due to lipopolysaccharide (LPS) contamination, because seaweed extracts can stimulate spleen cells of C3H/HeJ mice which are LPS low responders. Seaweed extracts also enhanced Ig production by B cells and tumor necrosis factor (TNF) production by macrophages. Furthermore, seaweed extracts stimulated human lymphocytes to proliferate. All these B cell stimulating activities of seaweed extracts associated with glycoproteins whose molecular weights resided in 100 kD. These results suggest that seaweed extracts have stimulating activity on B cells and macrophages and this ability could be clinically for the modulation of immune responses.

Animals↗

PubMed1997

Immunomodulating activity of Chinese medicinal herbs and Oldenlandia diffusa in particular.

The effect of eight different Chinese medicinal herbs (CMHs) on lymphocytes was studied in vitro using murine spleen cells. Among the studied eight CMHs, Astragalus membranaceus and Oldenlandia diffusa markedly stimulated murine spleen cells to proliferate. The responder cells for CMHs were B cells, because the response was depleted by the treatment of spleen cells with anti-immunoglobulin (i.g.) antibody and complement and after purification by nylon wool column. This response was not due to contamination by lipopolysaccharide (LPS), because CMHs could stimulate C3H/HeJ spleen cells which are low responders to LPS. CMHs enhanced the production of Ig. CMHs also enhanced the induction of allo-antigen specific cytotoxic T lymphocytes. However, CMHs had no effect on natural killer cells. Furthermore, CMHs stimulated macrophages to produce interleukin-6 and tumor necrosis factor. The electroelution of the proteins from SDS-PAGE gel showed that the active components of Oldenlandia diffusa had an apparent molecular weight of 90-200 kD and were sensitive to pronase E and NaIO4 treatment, suggesting glycoproteins in nature. These results suggest that CMHs have immunomodulating activity in vitro and this activity could be used clinically for the modulation of immune responses.

Adjuvants, Immunologic↗

PubMed1997

Human T cell responses to recombinant mite antigens of Dermatophagoides farinae.

We studied T cell responses to four glutathione S transferase (GST)-fused mite antigens prepared in our laboratory using peripheral blood lymphocytes from mite-sensitive patients with bronchial asthma. Of the four recombinant antigens, purified GST-Mag3 had the strongest ability to cause patients' lymphocytes to proliferate, and its potency was almost comparable to that of crude mite bodies (Dfb) and faeces (Dff) antigens and a purified major antigen, Der f 2. The responder lymphocytes were mainly T cells, because the proliferative response was depleted by the treatment of lymphocytes with anti-CD3 antibody and complement, but not with anti-CD20 antibody and complement. The responsiveness of lymphocytes to GST-Mag3 correlated with that to Der f 2, but GST-Mag3 displayed slightly higher activity to stimulate lymphocytes than Der f 2. Simultaneously, the levels of Dff- and GST-Mag3-specific IgE antibodies correlated with the responsiveness of lymphocytes to GST-Mag3. These results suggest that Mag3 is a new valuable antigen for the response of T cell proliferation in mite-sensitive patients.

Allergens↗

PubMed1997

Enhancing effect of polysaccharides from an edible brown alga, Hijikia fusiforme (Hijiki), on release of tumor necrosis factor-alpha from macrophages of endotoxin-nonresponder C3H/HeJ mice.

An enhancing activity for the release of tumor necrosis factor-alpha from macrophages of C3H/HeJ mice was detected in the hot water-soluble extract of an edible brown alga, Hijikia fusiforme (Hijiki in Japanese). This activity was divided into the polysaccharide and nonpolysaccharide fractions, with the former showing much higher activity than the latter. The active components in the polysaccharide fraction were further purified by ion-exchange column chromatography and gel permeation system of high-performance liquid chromatography; they were identified as polysaccharides with apparent molecular mass of about 2,000 and 70 kDa and were designated Hijiki-derived polysaccharides I and II (HPS-I and HPS-II), respectively. They also enhanced macrophage-dependent suppression against the growth of EL-4 tumor cells in an in vitro culture experiment, with HPS-I exhibiting much higher immunologic activity than HPS-II. Furthermore, other comparative experiments confirmed that the immunoenhancing activities of polysaccharides from H. fusiforme are associated with the functions of polysaccharides themselves, but not with the artificial activity induced by contaminated endotoxins. Some biochemical properties of immunoenhancing polysaccharides were partially characterized, and the significance of this finding is discussed from the viewpoint of the protective role of edible seaweeds against carcinogenesis.

Adjuvants, Immunologic↗

PubMed1997

Identification of T-cell epitope sequences on an important mite antigen.

BACKGROUND: T-cell epitopes on Der 1 and Der 2 groups, the major mite allergens, have been intensively analysed, while those on the other important allergens remain to be elucidated. We have cloned four cDNAs coding for important mite allergens on the basis of frequency and capacity of IgE binding. Stimulatory action of glutathione S-transferase-fused Mag1 on lymphocytes from mite-allergic patients was relatively high among them. OBJECTIVE: To identify T-cell epitopes on Mag1, we studied the stimulating activity of truncated Mag1 proteins and synthetic peptides on proliferative response of lymphocytes from mite antigen-immunized mice and mite-sensitive patients. METHODS: Truncated Mag1 proteins were expressed as a fusion protein with beta-galactosidase in Escherichia coli pop2136 carrying a variety of deleted Mag1 inserts. Murine T-cell epitope regions were estimated by the truncated antigen-induced lymphocyte proliferation assay. Overlapping peptides covering the whole sequence of the presumed T-cell epitope regions were synthesized to identify the epitope core sequences using murine and human Mag1-specific T-cell lines. RESULTS: Amino acid range 56-70 on Mag1 molecule showed remarkable stimulatory action on murine T cells, while amino acid ranges 51-65 and 86-100 had potent stimulatory activity on human T cells. CONCLUSIONS: These results suggest that Mag1 is a valuable antigen suitable for studies on T-cell responses and T-cell epitopes in mice and humans.

Animals↗

PubMed1997

Interleukin-1 as an autocrine stimulator in the growth of human ovarian cancer cells.

The role of interleukin-1 (IL-1), a multifunctional cytokine which mediates important immune responses, was investigated in the growth of ovarian cancer cell lines in vitro. The messenger RNA for IL-1 alpha and IL-1 beta was expressed in six and four ovarian cancer cell lines, respectively out of eight. Measurement of IL-1 in the eight cell lines by enzyme-linked immunosorbent assay revealed that two lines, MCAS and TYK-nu, secreted a high amount of IL-1 alpha, but that none secreted IL-1 beta after 72 hours of incubation. The growth of these cells was significantly stimulated by the addition of recombinant IL-1 alpha (rIL-1 alpha) in a concentration-dependent manner in a 96 hour culture. The maximum response was obtained with 10 ng/ml of IL-1 alpha by counting the viable cell number using trypan blue. [3H]-thymidine incorporation by these cells was also stimulated by a 72 hour incubation with rIL-1 alpha. The spontaneous growth of these cells was inhibited by the addition of anti-IL-1 alpha antibody, anti-IL-1 receptor antibody or IL-1 receptor antagonist. These cells expressed two classes of IL-1 binding receptors on their surface as detected by [125I]-labeled rIL-1 alpha. These results indicate that IL-1 alpha is an autocrine growth stimulator for some ovarian cancer cells and suggest that IL-1 alpha plays an important role in the progression of this disease.

Antibodies, Monoclonal↗

PubMed1996

B cell mitogenic activity of house dust mite, Dermatophagoides farinae, antigens.

The effect of mite antigens on murine and human lymphocytes was studied in vitro. Antigens prepared from Dermatophagoides farinae feces and bodies stimulated normal murine spleen cells to proliferate in a dose-dependent manner. The responder cells are B cells, because the response was reduced by the treatment of spleen cells with anti-immunoglobulin antibody and complement, but not with anti-Thy 1 antibody and complement. Furthermore, nylon column-purified T cells did not respond. The stimulation of B cells with mite antigens was not due to the contamination of lipopolysaccharide, a representative B cell mitogen, because C3H/HeJ spleen cells which are low responders to lipopolysaccharide could respond to mite antigens. These antigens induced not only proliferative response of murine B cells, but also immunoglobulin production. By gel-filtration column chromatography, the active fractions were eluted around the molecular weight of 150-155 kDa. Furthermore, mite antigens also stimulated human B cells to proliferate and to produce immunoglobulin. All these results suggest that mite antigens are a potent B cell mitogen and this activity might concern the induction of allergic reaction.

Animals↗

PubMed1996

Biphasic changes in behavioral, endocrine, and sympathetic systems in adjuvant arthritis in Lewis rats.

Adjuvant arthritis (AA) is an experimental model for rheumatoid arthritis, and is induced most easily in inbred Lewis rats by an intradermal injection of heat-killed Mycobacterium tuberculosis (MT) in incomplete Freund's adjuvant. Susceptivity to the arthritis in Lewis rats is thought to be related to a defect in their responses of the hypothalamo-pituitary-adrenal (HPA) axis to the disease. Because the use of an inbred strain is necessary for our immunological studies, we examined in Lewis rats changes in behavior, the HPA axis, and sympathetic nerve activities during development of the adjuvant arthritis. Following intradermal injections of heat-killed MT in adjuvant, the arthritis began to develop on day 12, reaching its maximum severity on day 21, and remained at the level for over a month. The body temperature rose from day 0 to 5 (the primary phase--before the onset of the arthritis). It then fell to normal temperature, and again rose from day 10 to 21 (the secondary phase--with fully developed arthritis). The behavioral (physical activity, food, and water intake) and hormonal parameters [plasma adrenocorticotropic hormone (ACTH) and corticosterone levels] also changed in two phases, similar to those observed in the temperature responses. No change in plasma vasopressin level was observed. Sympathetic nerve activities, assessed by changes in plasma noradrenalin levels, increased more in the primary than in the secondary phase. The possible causes for the biphasic changes associated with development of arthritis are discussed.

Animals↗

PubMed1996

Induction of autoimmune hepatitis and autoantibodies to liver antigens by neonatal thymectomy in mice.

We examined development of autoimmune hepatitis in neonatally thymectomized C3H/HeN mice and tried to characterize the nature of liver antigens recognized by the autoantibodies at the molecular level. Autoantibodies to crude liver proteins detected by ELISA were found in 12(67%) of 18 mice thymectomized 2 days after birth. However, autoantibodies were not detected in mice thymectomized 7 days after birth. The autoantibodies mainly consisted of IgG and reached the maximum level 8 weeks after birth. Hepatic inflammation, mononuclear cell infiltration in the portal area, was seen in 5 (28%) of 18 mice thymectomized 2 days after birth, but not in mice thymectomized 7 days after birth. Most infiltrating cells were Thy-1+ lymphocytes. The serum autoantibody level to crude liver proteins in mice with hepatitis was much higher than that in mice without hepatitis. We fractionated crude liver proteins by a Sepharose 6B column and examined the reactivity against the autoantibodies. The autoantibodies of three of five mice with hepatitis reacted with th approximately 150kD liver proteins other than liver-specific protein (LSP). By Western immunoblotting of SCS-PAGE using LSP and fractionated liver proteins, we found that the molecular weights of the target antigens were 52kD in LSP and 150kD (strong band), 138, 128, 120 and 110kD (weak band) in fractionated liver proteins other than LSP. This 150-kD target molecule in crude liver proteins was found only in liver. These results indicate that hepatitis and autoantibodies to liver proteins are induced spontaneously by neonatal thymectomy in mice, and the candidates of autoantigen in this hepatitis model are 52-kD protein in LSP and 150-kD liver proteins different from LSP. Still more, we regard the 150-kD molecule as a new autoantigen related to hepatitis.

Age Factors↗

PubMed1996

Concomitant transcriptional activation of nitric oxide synthase and heme oxygenase genes during nitric oxide-mediated macrophage cytostasis.

During in vitro activation of mouse peritoneal macrophages with interferon-gamma (IFN-gamma) and lipopolysaccharide (LPS), their synthesis of peroxynitrite and their cytostatic activity against mouse lymphocytic leukemia (L1210) cells were examined. The activation of the genes for nitric oxide synthase (iNOS) and heme oxygenase (HO-1) was also determined during the activation of the macrophages. Results showed that activation of peroxynitrite synthesis in macrophages was accompanied by the transcriptional activation of iNOS and HO-1 genes. Both genes seem to be activated simultaneously upon activation of the macrophages. Simultaneous activation of iNOS and HO-1 genes may be important because degradation of heme by HO-1 is one of the most important reaction that produces CO in higher organisms, and nitric oxide (NO) and carbon monoxide (CO) can react with heme-containing guanylate cyclase.

Animals↗