Biomedical subjects

S Yonehara

Publications and source records attributed to S Yonehara.

At least 91 records · Page 5Linked to original sources

PubMed1995

A recessive mutant of the U937 cell line acquired resistance to anti-Fas and anti-p55 tumor necrosis factor receptor antibody-induced apoptosis.

Human monocytic leukemia U937 cells readily undergo apoptosis when cells are treated with various stimuli including antitumor agents, tumor necrosis factor (TNF)-alpha and anti-Fas antibody. However, the signal transduction mechanism resulting in apoptosis is unclear. To study the mechanism of apoptosis, we isolated and characterized a mutant, UK110, from U937 cells, which was resistant to TNF-alpha and anti-Fas antibody-induced apoptosis but was less resistant to etoposide-induced apoptosis. TNF-alpha induced signals are mediated by two types of TNF receptors (TNFR), p55- and p75-TNFR, and p55-TNFR is homologous to the Fas antigen. Interestingly, UK110 cells showed resistance to apoptosis by agonistic anti-p55-TNFR antibody, indicating that UK110 cells were resistant to Fas- and p55-TNFR-mediated apoptosis. Because expression of apoptosis-associated molecules, such as c-Myc, Bcl-2, and Bax, was similar between U937 and UK110 cells an undetermined pathway for apoptosis through Fas and p55-TNFR could be mutated in UK110 cells. To clarify the genetic phenotype of UK110 cells, we performed somatic cell hybridization with parental U937 and the UK110 cells. All of the hybrid clones were as sensitive as the parental U937 cells to apoptosis by both anti-Fas and anti-p55-TNFR antibodies, indicating that the apoptosis resistance in UK110 cells resulted from recessive genotype.

Antibodies↗

PubMed1994

IL-5 receptor-mediated tyrosine phosphorylation of SH2/SH3-containing proteins and activation of Bruton's tyrosine and Janus 2 kinases.

Interleukin 5 (IL-5) induces proliferation and differentiation of B cells and eosinophils by interacting with its receptor (IL-5R) which consists of two distinct polypeptide chains, alpha and beta (beta c). Although both IL-5R alpha and beta c lack a kinase catalytic domain, IL-5 is capable of inducing tyrosine phosphorylation of cellular proteins. We investigated the role of IL-5R alpha in tyrosine phosphorylation of molecules involved in IL-5 signal transduction, using an IL-5-dependent early B cell line, Y16 and transfectants expressing intact or mutant IL-5R alpha together with intact beta c. The results revealed that the transfectants expressing truncated IL-5R alpha, which entirely lacks a cytoplasmic domain, together with beta c, showed neither protein-tyrosine phosphorylation nor proliferation in response to IL-5. This confirms that IL-5R alpha plays a critical role in protein-tyrosine phosphorylation which triggers cell growth. IL-5 stimulation results in rapid tyrosine phosphorylation of beta c and proteins containing Src homology 2 (SH2) and/or SH3 domains such as phosphatidyl-inositol-3 kinase, Shc, Vav, and HS1, suggesting their involvement in IL-5-mediated signal transduction. IL-5 stimulation significantly enhanced activities of Janus 2 and B cell-specific Bruton's tyrosine kinases (JAK2 and Btk) and increased the tyrosine phosphorylation of JAK2 kinase. These results and recent data on signaling of growth factors taken together, multiple biochemical pathways driven by tyrosine kinases such as JAK2 and Btk are involved in IL-5 signal transduction.

Agammaglobulinaemia Tyrosine Kinase↗

PubMed1994

Serum alleviates the requirement of the granulocyte-macrophage colony-stimulating factor (GM-CSF)-induced Ras activation for proliferation of BaF3 cells.

Deletion analysis of the beta subunit of the human granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor previously defined two cytoplasmic regions required for distinct signaling. The membrane-proximal region is responsible for induction of c-myc and pim-1, and is indispensable for GM-CSF-dependent proliferation of mouse BaF3 transfectants. The distal region is required for activation of Ras, Raf-1, MAP kinase and p70 S6 kinase as well as induction of c-fos and c-jun, but is dispensable for GM-CSF-dependent proliferation of transfectants under normal culture conditions containing serum. Here we show that signals induced by the distal region of the beta subunit are also required for proliferation. GM-CSF supported proliferation of BaF3 transfectants expressing the normal beta subunit, even in serum-free medium. However, in the absence of seru, GM-CSF did not support proliferation of BaF3 transfectants that have the beta deletion mutants lacking the distal region. Serum-induced activation of Ras, phosphorylation of MAP kinase and expression of c-fos in parental BaF3 cells and antisense oligonucleotide against c-raf blocked DNA synthesis of BaF3 cells. These results indicate that proliferation of BaF3 cells requires signals induced by the proximal as well as the distal region of the beta subunit of the GM-CSF receptor, and that serum alleviates the requirement of signals induced by the distal region.

Animals↗

PubMed1994

Cytotoxicity of fresh NK1.1+ T cell receptor alpha/beta+ thymocytes against a CD4+8+ thymocyte population associated with intact Fas antigen expression on the target.

Recent studies have revealed that 10-20% of CD4+8- or CD4-8- thymocyte populations contain NK1.1+ T cell receptor (TCR)-alpha/beta+ cells. This subpopulation shows characteristics that are different from NK1.1- CD4+ or NK1.1- CD8+ T cells and seems to have developed in a manner different from NK1.1- T cells. Although extensive studies have been performed on the NK1.1+ TCR-alpha/beta+ thymocytes, the physiological role of the NK1.1+ TCR-alpha/beta+ thymocytes has been totally unclear. In the present study, we found that freshly isolated NK1.1+ TCR-alpha/beta+ thymocytes, but neither whole thymocytes nor lymph node T cells, directly killed CD4+8+ thymocytes from normal syngeneic or allogeneic mice by using a long-term cytotoxic assay in which flow cytometry was used to detect the cytotoxicity. However, only weak cytotoxicity was detected against thymocytes from lpr mice on which the Fas antigen that transduces signals for apoptosis into the cells is not expressed. Furthermore, the NK1.1+ TCR-alpha/beta+ thymocytes exhibited high cytotoxicity against T lymphoma targets transfected with fas genes as compared with the parental T lymphoma targets or target cells transfected with mutated fas genes, which lack the function of transducing signals. On the other hand, NK1.1+ effector thymocytes from gld mice that carry a point mutation in Fas ligand did not kill thymocyte targets from normal mice. The present findings, thus, consistently suggest that the NK1.1+ TCR-alpha/beta+ thymocytes kill a subpopulation among CD4+8+ thymocytes via Fas antigen and in this way regulate generation of T lineage cells in the thymus.

Animals↗

PubMed1994

Fas and its ligand in a general mechanism of T-cell-mediated cytotoxicity.

To investigate the mechanisms of T-cell-mediated cytotoxicity, we estimated the involvement of apoptosis-inducing Fas molecule on the target cells and its ligand on the effector cells. When redirected by ConA or anti-CD3 monoclonal antibody, a CD4+ T-cell clone, BK1, could lyse the target cells expressing wild-type Fas molecule but not those expressing death signaling-deficient mutants. This indicates the involvement of Fas-mediated signal transduction in the target cell lysis by BK1. Anti-CD3-activated but not resting BK1 expressed Fas ligand as detected by binding of a soluble Fas-Ig fusion protein, and the BK1-mediated cytotoxicity was blocked by the addition of Fas-Ig, implicating the inducible Fas ligand in the BK1 cytotoxicity. Ability to exert the Fas-mediated cytotoxicity was not confined to BK1, but splenic CD4+ T cells and, to a lesser extent, CD8+ T cells could also exert the Fas-dependent target cell lysis. This indicates that the Fas-mediated target cell lytic pathway can be generally involved in the T-cell-mediated cytotoxicity. Interestingly, CD4+ T cells prepared from gld/gld mice did not mediate the Fas-mediated cytotoxicity, indicating defective expression of functional Fas ligand in gld mice.

Animals↗

PubMed1994

Anti-Fas on nonhematopoietic tumors: levels of Fas/APO-1 and bcl-2 are not predictive of biological responsiveness.

Fas/APO-1 is a cell surface protein known to trigger apoptosis in a variety of cell types upon specific antibody binding. Although extensively studied on normal and malignant hematopoietic cells, little is known about Fas/APO-1 on nonhematopoietic cells. In the study presented here, we have examined Fas/APO-1 expression and function on 11 human tumors of nonhematopoietic origin. By flow cytometric analysis, Fas/APO-1 was expressed on 10 of the 11 tumors at levels comparable to those previously reported for lymphoid cells sensitive to the cytolytic effects of anti-Fas. Despite abundant cell surface expression, only 4 of the 10 Fas-positive tumors were sensitive to the cell-killing effects of anti-Fas. Moreover, anti-Fas enhanced the growth of 2 of 10 Fas-positive tumors. Additional studies using cycloheximide demonstrated that de novo protein synthesis was required for anti-Fas-triggered growth stimulation and, at least in one case, was responsible for the resistance to antibody-induced apoptosis. The biological effects initiated by anti-Fas engagement, however, were not correlated with endogenous bcl-2 expression. This report documents that: (a) Fas/APO-1 is widely expressed on cultured nonhematopoietic tumors; (b) the inherent susceptibility to anti-Fas-induced apoptosis is not correlated with expression of the Fas/APO-1 protein; (c) Fas/APO-1 engagement can result in growth enhancement; and (d) protective/growth-promoting proteins other than bcl-2 may contribute to the diverse spectrum of biological effects induced by anti-Fas engagement of the Fas/APO-1 protein.

Antigens, Neoplasm↗

PubMed1994

Anti-Fas antibody induces different types of cell death in the human histiocytic cell line, U937, and the human B cell line, B104: the role of single-strand DNA breaks and poly (ADP-ribosyl)ation in cell death.

We investigated the anti-Fas antibody-induced cell death in two different types of human cell lines, U937 and B104. IFN-gamma increased the surface expression of Fas antigen and susceptibility to anti-Fas Ab-induced cell death of B104 and U937 cells. Anti-Fas Ab-induced death of U937 and B104 cells required neither a Ca2+ influx nor macromolecular synthesis. U937 cells treated with anti-Fas Ab represented apoptosis with DNA fragmentation, whereas anti-Fas Ab-treated B104 cells did not. Single-strand DNA breaks, however, appeared in the B104 cells. Zinc ions prevented DNA fragmentation and the morphological features of apoptosis in anti-Fas Ab-treated U937 cells, but did not inhibit cell death. However, zinc ions, when used in combination with the poly(ADP-ribosyl)ation inhibitors, inhibited anti-Fas Ab-induced U937 cell death. The inhibitors by themselves did not inhibit anti-Fas Ab-induced U937 cell death, but did inhibit anti-Fas Ab-induced B104 cell death. A substantial decrease in NAD pools was observed in anti-Fas Ab-treated B104 and U937 cells in parallel with the increase of DNA strand breaks before cell death became apparent. These results suggest the involvement of single-strand DNA breaks and poly(ADP-ribosyl)ation in the mechanisms of anti-Fas Ab-induced U937 and B104 cell death.

Antigens, Surface↗

PubMed1994

Alteration in length of telomeric repeats in lung cancer.

We investigated the relationship between telomere length and various characteristics of tumor cells in 46 lung cancer specimens (40 primary lesions and six metastatic lesions). Three variant patterns of telomere length were observed in 16 cases (34.8%): reduction in 13 cases, elongation in two cases, and convergence in one case. These variant patterns were frequently observed in small cell carcinomas, in metastatic lesions, and in cases which possessed the S-type allele of the L-myc gene. All three cases with telomere elongation or convergence were associated with a poor prognosis. This is compatible with the previous report suggesting that telomerase activity may be an indicator of immortality in vitro. In adenocarcinoma, telomere reduction or elongation was also observed in the early stages with a low percentage of cells in the S-phase, while in cases with other histologic types, these changes were observed only in late stage, in metastatic lesions, or in cancerous tissues with a high percentage of cells in the S-phase. Although the reduction of telomere length in these tissues may be a result of many cell divisions, it may represent another stage of carcinogenesis in early-stage adenocarcinoma.

Aged↗

PubMed1994

Tumorigenicity study of ferric citrate administered orally to mice.

Ferric citrate (FC) was orally administered at concentrations of 0.12 (maximum tolerated dose) or 0.06% in the drinking water to male and female B6C3F1 mice. Treatment was continued for 96 wk and the experiment was terminated at wk 100. There was no significant difference between treated and control groups in the tumour incidence or in the distribution of different types of tumour. Thus the long-term oral administration of FC to mice did not yield any evidence of chronic toxicity or tumorigenicity.

Administration, Oral↗

PubMed1994

p53 mutations in lung cancers from Japanese mustard gas workers.

Mustard gas (MG) is a mutagenic and carcinogenic alkylating agent, and is a known risk factor for occupational lung cancer. Our hypothesis is that lung cancers from MG workers contain mutations (G:C to A:T transitions) as the result of MG-produced DNA promutagenic adducts in the p53 tumor suppressor gene. We analyzed 12 primary lung cancers from Japanese MG factory workers and 12 lung cancers from non-exposed individuals. Genomic DNA was isolated from archival paraffin-embedded tissues. Exons 5-8 were amplified by polymerase chain reaction using p53-specific primers, and sequenced by dideoxy termination methods. Six out of 12 lung cancers from MG workers contained a total of eight somatic point mutations: two cases had double G:C to A:T transitions; one had a G:C to T:A transversion; one case had an A:T to G:C transition; and two cases had single base deletions. Four of the six mutated purines occurred on the non-transcribed, DNA-coding strand. Out of 12 unexposed cases, there were six single base mutations in six cancers, and no double mutations. The p53 mutational frequency in the MG-exposed cases is similar to the non-exposed controls and the usual smoking-related lung cancers reported previously. However, the distinctive double mutations (G:C to A:T transition) observed in two cases are unusual and may be related to MG exposure.

Aged↗

PubMed1994

Involvement of apoptosis antigen Fas in clonal deletion of human thymocytes.

Apoptosis appears to play a major role in the differentiation and selection of T and B lymphocytes, but the mechanisms of clonal deletion of T cells in thymus are not well understood. We have prepared an anti-human Fas IgM mAb with associated apoptosis-inducing activity in Fas antigen-positive target cells including human T cells. We analyzed the expression of apoptosis antigen Fas on human thymocytes by cytofluorometry showing low, but significant amounts of Fas antigen on double-negative and double-positive undifferentiated thymocytes. On the contrary, most of the differentiated thymocytes (single-positive or CD3-brightest) expressed undetectable levels of Fas antigen. About 1-2% of thymocytes expressed high amounts of Fas antigen, and these cells, which were CD3-bright, were CD4-bright and CD8-low at the stage of late double-positive lineage. Immunohistological analysis shows these Fas-bright cells on the edge of the medulla. Stimulation through the TCR complex was shown to induce the expression of Fas antigen on thymocytes at the late double-positive stage and prolonged stimulation through the TCR complex rendered the Fas-bright thymocytes sensitive to apoptosis-inducing activity of anti-Fas. To show the involvement of the Fas system in the negative selection/clonal deletion of thymocytes, we organ-cultured human thymus in the presence of the superantigen, staphylococcus enterotoxin B (SEB), and new antagonistic anti-Fas mAb, which can inhibit the apoptosis-inducing activity of the original anti-Fas mAb. The SEB-reactive TCR complex on thymocytes was at first down-regulated by SEB, then the SEB-reactive clone was deleted by apoptosis, which was inhibited by an antagonistic anti-Fas mAb. Thus, Fas antigen is shown to be involved in the negative selection/clonal deletion of superantigen-reactive thymocytes.

Antibodies, Monoclonal↗

PubMed1994

Expression of Fas antigen on keratinocytes in vivo and induction of apoptosis in cultured keratinocytes.

Fas antigen, which belongs to a nerve growth factor/tumor necrosis factor receptor superfamily, is a membrane protein that induces apoptosis. In humans, distribution of Fas antigen has been reported on cell lines and lymphocytes. Immunohistochemical studies revealed Fas antigen on the keratinocytes of lesional epidermis in lichenoid drug eruption, erythema multiforme, contact dermatitis, bullous pemphigoid, pemphigus vulgaris, and herpes zoster; it is co-expressed with intercellular adhesion molecule-1. Cultured keratinocytes expressing Fas antigen increased from 8.4% to 34.6% after stimulation with interferon gamma for 24 h. Treatment of interferon-gamma-stimulated keratinocytes with anti-Fas for 48 h resulted in DNA fragmentation and death of 32% of cells, suggesting that Fas antigen may mediate apoptosis. The expression of Fas antigen on keratinocytes in lesional skin suggests that death via Fas antigen may play an important role in the pathogenesis of keratinocyte cytotoxicity.

Antibodies↗

PubMed1994

Further experimental evidence of selective nerve regeneration in aortic Y-chambers.

To explain the discrepancy between Weiss and Taylor's results and our recent findings about axonal growth patterns in a fresh aortic chamber, we examined whether regenerating axons show preferential growth toward a nerve piece in an aortic Y-chamber. In our previous studies, we could not reproduce their results, which showed that regenerating nerve fibres from the proximal stump of a severed peripheral nerve grew randomly in the two channels with and without a nerve segment; our results showed preferential regeneration toward a nerve segment channel. In the present study, we retraced the steps of Weiss and Taylor using white rats, the longitudinal section of the specimen, and silver staining. Despite using the same methods we were still unable to show that the growth pattern was random, but we did find preferential axonal growth into the nerve segment channel of a fresh aortic Y-chamber. The evidence of selective nerve regeneration was unequivocal and we cannot support the findings of Weiss and Taylor reported in 1944.

Animals↗

PubMed1993

Unidirectional cross-phosphorylation between the granulocyte colony-stimulating factor and interleukin 3 receptors.

The mouse interleukin-3 (IL-3)-dependent hemopoietic precursor cell line 32DC13 responds to granulocyte colony-stimulating factor (G-CSF) for proliferation and differentiation. We established a subline (32D-FH) of the 32DC13 cells which has lost the ability to respond to G-CSF. When murine G-CSF receptor cDNA was introduced into the 32D-FH cell line, the transformants responded to G-CSF as well as to IL-3 for proliferation. Adding G-CSF to the transformants rapidly induced tyrosine phosphorylation not only of the G-CSF receptor but also of both subtypes of the IL-3 receptor beta-chain (AIC2A and AIC2B molecules). On the other hand, stimulation of the transformants with IL-3 induced tyrosine phosphorylation of the AIC2A and AIC2B but not of the G-CSF receptor. These results indicate that the tyrosine kinase activated through the G-CSF receptor interacts with the IL-3 receptor beta-chains but not vice versa. This unidirectional cross-phosphorylation between the G-CSF and IL-3 receptors may play a role in granulopoiesis induced by G-CSF.

Amino Acids↗

PubMed1993

Overcoming tumor necrosis factor and drug resistance of human tumor cell lines by combination treatment with anti-Fas antibody and drugs or toxins.

Monoclonal mouse anti-Fas antibody is directed against Fas antigen, a M(r) 36,000 encoded polypeptide that belongs to the family of cell surface proteins which includes nerve growth factor receptor, tumor necrosis factor (TNF) receptors, B-cell antigen CD40, and T-cell antigens OX40. Anti-Fas antibody mimics TNF-alpha in its cytolytic activity but not in other TNF-alpha-mediated activities. Thus, we examined if anti-Fas antibody synergizes in cytotoxicity with toxins and drugs. The present studies demonstrate that anti-Fas antibody in combination with diphtheria toxin (DTX), Adriamycin, or cis-platinum results in enhanced cytotoxicity and synergy and also overrides resistance to TNF, drugs, or toxins when tested against a battery of human tumor cell lines. Synergy with anti-Fas and DTX requires that DTX is enzymatically active, since inhibitors of DTX-mediated protein synthesis inhibition resulted in loss of synergy. When the plant toxin ricin was used, there was no synergy with anti-Fas antibody but rather an additive effect. The synergy was not obtained in a TNF receptor-negative line but was achieved with other anti-Fas-resistant lines. Cell lines resistant to either Adriamycin or cis-platinum were rendered sensitive by the combination of drug and anti-Fas antibody. Further, combination treatment of anti-Fas and Adriamycin overcame resistance of the gp 170-expressing, multidrug-resistant MDR ovarian line. In all cases, cytotoxicity was augmented by pretreatment of target cells with gamma-interferon which upregulates Fas antigen expression. These results show that anti-Fas antibody can synergize in cytotoxicity with toxins and chemotherapeutic drugs, and combination treatment can reverse resistance to TNF, toxins, and/or drugs.

Antibodies, Monoclonal↗

PubMed1993

Thymic metastasis from prostatic carcinoma: report of a case.

The thymus is an important organ involved in cell-mediated immunological function, and to our knowledge, there has never been a case of thymic metastasis reported. We recently examined a 65-year-old man who presented at our department with a cough and shortness of breath on exertion. He had a history of prostatic carcinoma for which he had undergone an orchiectomy 11 years previously. Investigations disclosed a mediastinal tumor, 14 x 9 cm in size, and histological examination of the resected tumor confirmed a diagnosis of thymic metastasis from prostatic carcinoma.

Adenocarcinoma↗

PubMed1993

Rapid acceleration of neutrophil apoptosis by tumor necrosis factor-alpha.

We demonstrate here that human necrosis factor-alpha, a potent neutrophil activator, induces rapid (within 3 h) apoptosis of these cells, i.e. neutrophils treated with this cytokine exhibit (i) light and electron microscopic changes characteristic to apoptotic cells, (ii) reduced propidium iodide binding to DNA, and (iii) the ladder form of DNA, as shown by agarose gel electrophoresis. These results suggest that apoptosis acceleration may be involved in processes by which neutrophils are prevented from damaging tissues.

Apoptosis↗

PubMed1993

IL-5 receptor positive B cells, but not eosinophils, are functionally and numerically influenced in mice carrying the X-linked immune defect.

Mouse IL-5 (mIL-5) acts on B cells and eosinophils to induce growth and differentiation through the mIL-5 specific receptor (mIL-5R). The functional high-affinity mIL-5R is a heterodimer composed of alpha and beta chains. We investigated the expression of mIL-5R and the responsiveness of B cells and eosinophils to mIL-5 in X-linked immunodeficient (xid) mice. mIL-5R expression analyzed by using mAbs specific for alpha and beta chains revealed that xid B cells had fewer mIL-5R alpha +mIL-5R beta + than BALB/c B cells. In particular, a decrease in the number of peritoneal mIL-5R+ B cells among Ly-1 B cells (known as B-1 cells) was remarkable. Furthermore, the frequency of precursors of mIL-5 responsive B cells in xid mice was approximately 100-fold lower than that of BALB/c mice. Interestingly, sorted mIL-5R+ peritoneal B cells from xid mice displayed a low response to mIL-5. Intraperitoneal injection of mIL-5 into BALB/c mice induced polyclonal IgM production and an increase in the number of eosinophils. The same regimen failed to induce an increase in the same parameters in xid mice. However, xid mice showed mIL-5-induced eosinophilia in peripheral blood to a similar extent as BALB/c mice. Eosinophils from mIL-5-injected xid mice expressed both alpha and beta chains of mIL-5, and responded to mIL-5 with prolonged in vitro survival.

Animals↗