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Biomedical subjects

K Katamura

Publications and source records attributed to K Katamura.

At least 37 records · Page 2Linked to original sources

Physical interaction with monocytes rescues human mature CD4+ T-cell lines from anti-CD3-induced apoptosis.

Crosslinking of the TcR-CD3 complex with immobilized anti-CD3 antibodies without sufficient co-stimulation induced cell death in human mature CD4+ T-cell lines. In these T cells, DNA fragmentation and morphological characteristics of apoptosis were seen. The anti-CD3-induced apoptosis was inhibited by co-culture with monocytes. The rescue signal provided by monocytes does not need to be present simultaneously with signals mediated by anti-CD3. When T cells were precultured with monocytes for 24 h before anti-CD3 stimulation and then the monocytes were removed from the culture, anti-CD3-induced T-cell apoptosis was also inhibited. To determine whether the monocyte-derived rescue signals were transduced by soluble factors or by direct cell-to-cell interaction with monocytes, we precultured T cells with monocytes separated by a micropore membrane which prevented T cell-monocyte physical interaction but not the diffusion of secreted molecules. In this system, rescue signals could not reach the T cells. To further assess the importance of physical interaction, we precultured T cells with fixed monocytes. T cells could not be rescued from apoptosis under these experimental conditions, either. The results considered collectively suggest that sufficient physical interaction with viable monocytes is important for the rescue of anti-CD3-induced apoptosis of CD4+ T cells.

Antibodies, Monoclonal↗

Role of LFA-1/ICAM-1-dependent cell adhesion in CD40-mediated inhibition of anti-IgM antibody-induced B-cell death.

Cross-linking of surface IgM by anti-IgM antibody caused activation-induced cell death of a surface IgM+, IgD+ human B lymphoma cell line, B104. The dying B104 cells did not show the morphology of apoptosis but did show that of necrosis. However, anti-IgM antibody caused apoptosis of another surface IgM+, IgD+ human B lymphoma cell line, DND-39. The influx of extracellular Ca2+ was necessary for the cell deaths of B104 and DND-39 caused by anti-IgM antibody. Their cell deaths were inhibited by cyclosporine. The anti-IgM antibody-induced cell death of DND-39, but not that of B104, was prevented by costimulation with anti-CD40 antibody. In human peripheral blood B-cells, anti-IgM antibody inhibited cell cycle transition induced by Staphylococcus aureus Cowan I at the G2/M interphase without inhibition of DNA synthesis. In this system, too, anti-CD40 antibody canceled the inhibitory signal transduced through surface IgM and increased the number of M phase cells. Blocking antibodies against the leukocyte function-associated antigen-I/intercellular adhesion molecule-1 system decreased the rescue effect of anti-CD40 antibody in both DND-39 cells and peripheral B-cells, which shows that leukocyte function-associated antigen-1/intercellular adhesion molecule-1-dependent cell adhesion plays an important role in the CD40-mediated inhibition of surface IgM-mediated negative signals.

Animals↗

Selective induction of interleukin-4- and interferon-y-producing T cells from cord blood naive T cells. Effects of costimulatory signaling through CD28.

We investigated the effect of costimulation through CD28 and CD11a on the differentiation of human naive CD4+ T cells with restricted cytokine production profiles. Interleukin (IL)-4 and interferon-gamma (IFN-gamma) were measured by ELISA and IL-2 was detected by a bioassay. Naive CD4+ T cells proliferated and produced IL-2 upon cross-linking of CD3, and costimulation through CD28 enhanced IL-2 production. After repeated stimulation, CD4+ T cells which were stimulated in the absence of costimulation through CD28 lost their ability to secrete IL-2 and started secreting IL-4 and IFN-gamma. Instead in the presence of costimulation through CD28, they secreted IL-2, IL-4 and IFN-gamma. Blocking of endogenous IL-4 activity with anti-IL-4 Ab suppressed the IL-4 secretion and proliferation of T cells.

CD28 Antigens↗

IgM-mediated B cell apoptosis.

Cross-linking of surface immunoglobulin M (sIgM) on normal mature B cells induces different signaling consequences, including DNA synthesis (positive signaling) and cell cycle arrest and/or death by apoptosis (negative signaling). Presumably, the difference depends on the intensity of sIgM cross-linking: relatively weak cross-linking induces DNA synthesis, moderate cross-linking induces DNA synthesis with cell cycle arrest at the G2/M interphase, and intense cross-linking induces apoptosis. In vivo experiments with transgenic mice have shown that relatively weak cross-linking of sIgM by soluble antigens induces anergy in autoreactive B cells, whereas intense sIgM cross-linking by membrane-bound forms of antigens induces deletion of them. However, it is still unknown whether the different intensities of sIgM cross-linking generate qualitatively different signals responsible for DNA synthesis or cell death or whether they generate qualitatively the same but quantitatively different signals, and the quantitative difference is responsible for the induction of positive or negative signaling. The sIgM-mediated negative signaling presumably plays an important role in the induction and maintenance of B cell tolerance, and sIgD and sIgG also possess the machinery necessary for negative signaling. Negative signaling through sIgM is dependent on tyrosine kinase(s) and Ca2+ influx and is sensitive to cyclosporin A in certain types of B cells but not in all B cells. It has been suggested that there are different intracellular signaling pathways that transduce negative signaling via sIgM, and that activation-induced B cell death by sIgM cross-linking does not necessarily show DNA fragmentation and the morphology of apoptosis. On the other hand, sIgM-mediated B cell death may be inhibited in the presence of appropriate co-stimulators such as IL-4, alpha-, and beta-interferons and CD40-mediated signaling. The CD40-mediated signaling effectively inhibits sIgM-mediated B cell apoptosis in many but not all experimental systems. Although homotypic cell adhesion through the LFA-1/ICAM-1 dependent pathway was shown to be involved in certain types of CD40-mediated inhibition of sIgM-mediated negative signaling, it is still not known how the cytokines and CD40-mediated signaling inhibit sIgM-mediated B cell death. The molecular mechanisms responsible for sIgM-mediated negative signaling and for the inhibitory signaling against sIgM-mediated negative signaling need further elucidation.

Animals↗

Involvement of LFA-1/intracellular adhesion molecule-1-dependent cell adhesion in CD40-mediated inhibition of human B lymphoma cell death induced by surface IgM crosslinking.

B cells have been shown to receive negative signals for their growth through crosslinking of surface IgM (sIgM), and it has been demonstrated that anti-IgM Abs induce B cell death. Proliferation of B cells in response to Ag stimulation in vivo may thus require additional signals that inhibit the sIgM-transduced negative signals. Signaling through CD40 has been proposed as a candidate for such costimulatory signals. To investigate the role of CD40-transduced signals in sIgM-mediated B cell death, we used a human B cell line (DND-39) that expresses sIgM, sIgD, and CD40. Crosslinking of sIgM, but not sIgD, by Abs induced DND-39 cell death. The dying cells showed the morphology of apoptosis and DNA fragmentation. Anti-CD40 Abs induced homotypic adhesion of DND-39 cells and rescued them from anti-IgM Ab-induced cell death. Anti-CD40 Abs inhibited anti-IgM Ab-induced cell death when added within 3 h after stimulation with anti-IgM Ab. Treatment with Abs against CD11a, CD18, or CD54 inhibited not only the homotypic adhesion but also the inhibition of anti-IgM Ab-induced apoptosis by anti-CD40 Ab. CD11a antisense decreased the surface CD11a expression, the anti-CD40 Ab-induced homotypic adhesion, and the inhibitory effect of anti-CD40 Ab on anti-IgM Ab-induced apoptosis. The data show that LFA-1/ICAM-1-dependent cell adhesion induced by signaling through CD40 plays an important role in the inhibition of anti-IgM Ab-induced apoptosis of DND-39 cells.

Antigens, CD↗

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↗

Positive and negative signals transduced through surface immunoglobulins in human B cells.

Cross-linking of surface IgM and surface IgD by anti-IgM antibodies and anti-IgD antibodies, respectively, showed different effects on the growth of normal human peripheral blood B cells and the human B lymphoma cell line, B104. Only cross-linking of surface IgM transduced signals that inhibited cell division of peripheral blood B cells and B104 cells at the G2/M interphase. In B104 cells, the inhibition of cell division was followed by rapid B104 cell death. The negative signals were inhibited by cyclosporin A and FK-506 at lower concentrations than those that inhibited proliferation of the B cells. Anti-IgM antibody-induced B104 cell death was dependent on Ca2+ influx and macromolecular synthesis. B104 cells treated with anti-IgM antibodies showed neither DNA fragmentation or morphology of apoptosis but showed DNA single-strand breaks and morphology of necrosis. Nicotinamide inhibited anti-IgM antibody-induced B104 cell death and the involvement of poly(adenosine diphosphate-ribosyl)ation was suggested in the process of the B104 cell death. With regard to the intracellular mechanisms responsible for the different signals, however, no qualitative difference was detected in putative signal transducers, including tyrosine phosphorylated protein, phosphatidyl inositol turnover, Ca2+ influx, activation of protein kinase C, and messenger ribonucleic acid expression of c-fos and Egr-1 when surface IgM and surface IgD were crosslinked. Further investigations of the mechanisms responsible for the different signals transduced through surface IgM and surface IgD will provide better understanding of immunodeficiencies and autoimmune diseases.

B-Lymphocytes↗

Induction of phosphatidylinositol turnover and EGR-1 mRNA expression by crosslinking of surface IgM and IgD in the human B cell line B104.

We have previously shown that a human B lymphoma cell line, B104, expressed surface IgM (sIgM) and surface IgD (sIgD), and that crosslinking of sIgM and sIgD by anti-IgM antibody (Ab) and anti-IgD Ab, respectively, induced Ca2+ influx to almost the same degree, whereas only sIgM-crosslinking caused B104 cell death. Here, we investigated the accumulation of cyclic AMP (cAMP), the hydrolysis of inositol phosphates, protein kinase C (PKC) activity and the induction of Egr-1 and c-fos mRNA expression by sIgM- and sIgD-crosslinking to examine differences in the signals mediated through sIgM and sIgD in B104 cells. Both sIgM- and sIgD-crosslinking with antibodies induced elevation of cAMP levels, phosphatidylinositol turnover, PKC activation and expression of Egr-1 and c-fos mRNA, although sIgM-crosslinking was more effective than sIgD-crosslinking, presumably due to the higher expression of sIgM than of sIgD. Egr-1 mRNA expression induced by sIgM- and sIgD-crosslinking was inhibited by H7, erbstatin and genistein, but not by HA1004. Erbstatin and genistein inhibited the sIg-crosslinking-induced Egr-1 mRNA expression in a dose-dependent manner parallel to that observed in the inhibition of sIg-crosslinking-induced protein tyrosine phosphorylation. Phorbol myristate acetate induced Egr-1 mRNA expression but forskolin and dibutyryl cyclic AMP did not. These findings suggest that the Egr-1 mRNA activating signals through sIgM and sIgD are protein tyrosine kinase- and PKC-dependent, but protein kinase A-independent. Cyclosporin A (CsA) and FK506 rescued B104 cells from death induced by anti-IgM Ab, but did not affect the expression of Egr-1 and c-fos mRNA, showing that CsA and FK506 affect signal transducers differently from or downstream to these molecules. The difference in signals transduced through sIgM and sIgD in B104 cells is discussed.

B-Lymphocytes↗

Effects of cyclic AMP on expression of LFA-1, Mac-1, and VLA-4 and eosinophilic differentiation of a human leukemia cell line, EoL-1.

We examined the effect of dibutyryl cAMP (dbcAMP) on the expression of LFA-1 (CD11a/CD18), Mac-1 (CD11b/CD18), and VLA-4 (CD49/CD29) and on eosinophilic differentiation of a human leukemia cell line, EoL-1. Dibutyryl cAMP induced eosinophilic differentiation of EoL-1 cells from 6-9 days after the start of culture with down-regulation of CD11a, CD18, and CD49 expression and up-regulation of CD11b expression. Changes in integrin expression, except for CD18, were seen predominantly in the fraction containing eosinophilic granule-positive cells, suggesting that the changes were dependent on eosinophilic differentiation. On the other hand, dbcAMP-induced changes of integrin expression were reversible and were not seen on day 9 when dbcAMP was removed on day 3, whereas eosinophilic differentiation was still present. A combination of G-CSF and TNF-alpha, which also induced eosinophilic differentiation of EoL-1 cells, increased CD11b expression slightly but had no significant effect on the expression of the other integrins. Butyrate and PMA up-regulated CD11b expression without eosinophilic differentiation. The results collectively suggest that the regulation of integrin expression on EoL-1 cells is partly dependent and partly not dependent on eosinophilic differentiation. The possible involvement of protein kinase A and protein kinase C in these changes is suggested.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

Induction of phosphatidylinositol-linked Fc gamma receptor III expression on an eosinophilic cell line, EoL-1, by dibutyryl cyclic AMP and interferon-gamma.

The expression of Fc gamma receptor III (Fc gamma RIII) on a human eosinophilic leukemia cell line, EoL-1, was examined and compared with its expression on normal blood eosinophils. Surface Fc gamma RIII expression on EoL-1 cells could be induced in vitro with a combination of dibutyryl cAMP (dbcAMP) and gamma-interferon (IFN-gamma), but not with IFN-gamma or dbcAMP alone. Pretreatment of EoL-1 cells with dbcAMP induced EoL-1 cells to express Fc gamma RIII when stimulated with IFN-gamma, but EoL-1 cells pretreated with IFN-gamma and then stimulated with dbcAMP failed to express Fc gamma RIII. Cyclic AMP was shown to play a role in the effect of dbcAMP. Both the treatment with phosphatidyl-inositol-specific phospholipase C (PI-PLC) and the restriction enzyme digestion of Fc gamma RIII cDNA showed that the Fc gamma RIII on EoL-1 cells was a phosphatidylinositol-linked form. On the other hand, freshly isolated blood eosinophils constitutively expressed few, if any, Fc gamma RIII, and IFN-gamma induced Fc gamma RIII expression on them in vitro. Dibutyryl cAMP did not induce Fc gamma RIII expression and even suppressed the IFN-gamma-induced Fc gamma RIII expression on normal eosinophils. The EoL-1 cell line appears to be a useful in vitro model for the expression and function of the phosphatidylinositol-linked form of Fc gamma RIII on eosinophils.

Base Sequence↗

Cyclosporin A and FK506 block the negative signaling mediated by surface IgM cross-linking in normal human mature B cells.

Cross-linking of surface IgM (sIgM) or sIgD by anti-IgM Ab or anti-IgD Ab, respectively, induced DNA synthesis in peripheral blood B cells (PBL-B). Cell division, determined by the increase in the number of M phase cells, was also induced when PBL-B were stimulated with anti-IgD Ab plus IL-4 or Staphylococcus aureus Cowan I (SAC), but far less by stimulation with anti-IgM Ab plus IL-4. Anti-IgM Ab did not suppress the DNA synthesis induced by SAC or anti-IgD Ab plus IL-4, but it did suppress the cell division induced by them. Thus, sIgM cross-linking generates both positive and negative signaling to B-cell proliferation. Cyclosporin A (CSA) and FK506 suppressed DNA synthesis and cell division at relatively high concentrations. On the other hand, CSA and FK506 at lower concentrations blocked the anti-IgM Ab-generated inhibition of cell division without affecting DNA synthesis. Low concentrations of CSA did not affect the cell division induced by anti-IgD Ab plus IL-4 but did increase the cell division induced by SAC or anti-IgM Ab plus IL-4, suggesting that stimulation with SAC, as well as with anti-IgM Ab plus IL-4, generates both positive and negative signals to cell division, whereas sIgD lacks the ability to transduce negative signaling.

Antibodies, Anti-Idiotypic↗

Characterization of an eosinophilic leukemia cell differentiation factor (ELDF) produced by a human T cell leukemia cell line, HIL-3.

An adult T cell leukemia cell line, HIL-3, constitutively secretes a factor which induces the phenotypical and functional eosinophilic differentiation of a human eosinophilic leukemia cell line, EoL-1. Biochemical characteristics of the factor, termed eosinophilic leukemia cell differentiation factor (ELDF), were examined. ELDF was precipitated by 35 to 65% saturated ammonium sulfate from the culture supernatants of HIL-3 cells (HIL-3 sup). ELDF was eluted in a peak corresponding to a molecular weight of 30 to 40 kd by gel filtration. Isoelectric focusing in the Rotofor showed that ELDF had isoelectric points of 5 to 6. ELDF was trypsin-sensitive and stable to heat treatment at 65 degrees C for 30 minutes but labile at 80 degrees C or pH lower than 3. Half of the activity adhered to lentil-lectin but not to Con-A, indicating that a part of ELDF is glycoprotein with an N-linked carbohydrate moiety, which did not seem to be essential for ELDF activity. The biochemical characteristics of ELDF and blocking experiments using cytokine-specific neutralizing antibodies suggest that ELDF is different from gamma-interferon (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-5 (IL-5) and interleukin-2 (IL-2), which may exist in HIL-3 sup, and that ELDF may be a previously unrecognized leukemia differentiation factor.

Cell Differentiation↗

Anti-IgM antibody-induced cell death in a human B lymphoma cell line, B104, represents a novel programmed cell death.

We investigated the mechanisms of anti-IgM antibody-induced cell death in a recently established human surface IgM+ IgD+ B lymphoma cell line, B104, the growth of which is irreversibly inhibited by anti-IgM antibody but not by anti-IgD antibody, and compared it with the cell death of T cells via TCR/CD3 complex and with the cell death of a murine anti-IgM antibody-sensitive B lymphoma cell line, WEHI-231. The rapid time course of B104 cell death and its requirements for de novo macromolecular synthesis and Ca2+ influx suggest that anti-IgM antibody-induced B104 cell death is an active Ca(2+)-dependent programmed cell death. Moreover, cyclosporin A rescued B104 cells from this lethal signal, via surface IgM, suggesting that the intracellular mechanisms involved are quite similar to those of T cell death. DNA fragmentation, which has been reported in TCR/CD3 complex-mediated T cell death, apoptosis, was not involved in the B104 cell death process, but the possible involvement of DNA single-strand breaks was suggested. Observations under light microscopy and transmission electron microscopy indicated that the morphologic features of dying B104 cells resembled necrosis rather than apoptosis. B104 cell death was shown to be quite distinct from that of WEHI-231 in cell death kinetics, the mode of cell death, and the response to cyclosporin A. These data collectively indicate that the death of B104 cells resulting from surface IgM cross-linking represents a hitherto undefined mode of programmed cell death.

Antibodies, Anti-Idiotypic↗

Tyrosine phosphorylation of IgM- and IgD-associated molecules of a human B lymphoma cell line B104.

We investigated tyrosine phosphorylation and structural properties of the IgM-associated molecules in comparison with IgD-associated molecules in a recently established human surface IgM+, IgD+ B lymphoma cell line, B104, the growth of which was irreversibly inhibited by anti-IgM mAbs but not by anti-IgD mAbs. Tyrosine kinase activity and tyrosine phosphorylated proteins were detected in anti-IgM and anti-IgD immunoprecipitates from digitonin lysates of B104 cells with the use of an in vitro kinase assay followed by a re-immunoprecipitation experiment with anti-phosphotyrosine mAbs. Tyrosine phosphorylated proteins of 74, 58-44, 41, and 39 kDa were detected in anti-IgM immunoprecipitates, whereas tyrosine phosphorylated proteins of 74, 58-44, and 39 kDa, but not 41 kDa, were detected in anti-IgD immunoprecipitates. Crosslinking of surface IgM and surface IgD stimulated rapid tyrosine phosphorylation of different sets of proteins which included tyrosine-phosphorylated proteins of the same or similar molecular weights as those detected in the anti-IgM and anti-IgD immunoprecipitates respectively. After deglycosylation by N-glycosidase, both the IgM- and IgD-associated phosphoproteins (pp58-pp39) gave rise to the same three bands of 29, 27, and 26 kDa. Proteolytic peptide mapping of these three deglycosylated proteins showed that the primary structures of the IgM- and IgD-associated molecules are identical, suggesting that the IgM- and IgD-associated phosphoproteins (pp58-pp39) are the products of the same or closely related genes. One of the products, pp41, may be associated with IgM, but not with IgD, although the same gene product may be associated with IgD in a different glycosylation pattern.

Antibodies, Anti-Idiotypic↗

Disseminated intravascular coagulation syndrome in anorexia nervosa.

A 15 year old boy with anorexia nervosa developed disseminated intravascular coagulation syndrome (DIC). Because of severe cachexia he had been admitted to the Shimane Prefectural Central Hospital. During his hospitalization he developed generalized massive ecchymosis. Laboratory data revealed not only DIC but also multiple organ complications. The patient was treated intravenously with FOY (gabexate mesilate, a protease inhibitor), heparin, a transfusion of fresh frozen plasma, antithrombin III concentrates and platelets. Intravenous hyperalimentation was also administered. The laboratory data, the general condition and the emotional state of the patient improved remarkably. We emphasize the importance of keeping in mind coagulopathy as a complication in anorexia nervosa.

Adolescent↗

IL-4-producing murine T helper cell line provides help for in vitro production of IgE.

Studies of the regulation of the IgE synthesis have been hampered by the difficulties in making lymphocytes produce IgE in vitro. Synthesis of IgE, IgG1 and IgM were investigated in a murine coculture system in which a conalbumin-specific T helper cell line of Th2 type was cultured together with nonadherent splenocytes from normal MHC-matched mice. Help provided by the antigen-stimulated T cell line induced significant IgE production (20 ng/ml), along with IgG1 (5 micrograms/ml) and IgM (250 micrograms/ml). Immunoglobulin synthesis in cultures was detectable at day 3-4 and culminated at day 7-8. IL-4 production was observed within the first 2 days of culture. The kinetics of the synthesis of the three isotypes were parallel. Stimulation of the Th2 cell line with Con A produced dose-response curves resembling those after antigen stimulation, whereas supernatant from these cells was unable to induce synthesis of IgE. The presence of interferon-gamma in the cultures, inhibited synthesis of IgE, IgM and IgG1, but this inhibition was not due to interference with the IL-4 production, which increased in the presence of high doses of interferon-gamma.

Animals↗

Association of glycosylation-inhibiting factor with plasma membranes of T suppressor cell hybridomas.

The glycosylation inhibiting factor (GIF) was detected in EGTA extracts of the OVA-specific Ts cell hybridoma, 231F1 cells and 71B4 cells, which constitutively secrete GIF. The lymphokine in both culture supernatants and EGTA extracts failed to bind to OVA-Sepharose. Association of GIF with the plasma membrane was confirmed by surface labeling of the 231F1 cells with 125I. The major species of GIF in the extract was 14.4-kDa peptide as determined by SDS-PAGE, and was identical to that detected in culture supernatants. Pretreatment of the cells with monoclonal anti-GIF switched the cells from the formation of unglycosylated IgE-BF to the formation of glycosylated IgE-BF, indicating that the membrane-associated GIF is involved in the determination of the nature of IgE-binding factor during their biosynthesis. When the hybridoma was stimulated with OVA-pulsed APC, EGTA extracts of the cells contained GIF having affinity for OVA. The binding of the OVA-binding GIF in the EGTA extracts to OVA-Sepharose was inhibited by a synthetic peptide, which corresponds to amino acid residues 307-317 in the OVA molecule and represents the epitope recognized by TCR on the cells. The OVA-binding GIF in the extracts bound to the monoclonal anti-TCR-alpha chain, H-28-710 and the mAb 14-12, which is specific for the Ag-binding chain of effector type suppressor factor, and suppressed the in vivo antibody response of BDF1 mice to DNP-OVA in a carrier-specific manner. Evidence was obtained that indicated that the Ag-binding chain was associated with nonspecific GIF chain on the cell surface of the Ag-stimulated cells.

Animals↗

Biochemical identification of glycosylation inhibiting factor.

Rat monoclonal antibody against mouse glycosylation inhibiting factors was obtained, and radiolabeled glycosylation inhibiting factors from the mouse T-cell hybridoma, 231F1 cells, were purified by using the monoclonal antibody and antilipomodulin antibody. Analysis of the affinity-purified lymphokine by PAGE demonstrated two proteins of 14.4 kDa and 41 kDa. Both proteins migrated similarly under the reduced and unreduced conditions, indicating that each of the two species consist of a single polypeptide chain. Biologic activity of the lymphokine was recovered by extraction of the proteins from gel slices followed by renaturation. Evidence was obtained that suggested the 14.4-kDa peptide was a degradation product of the 41-kDa molecules. The 14.4-kDa peptide was also recovered by extraction of the surface-labeled 231F1 cells with Ca2+ chelator, indicating that the glycosylation inhibiting factor is associated with the cell surface.

Animals↗