bluuebunny/crossref_metadata_2025 huggingface dataset:
The role of the T cell receptor, CD8, and LFA-1 in different stages of the cytolytic reaction mediated by alloreactive T lymphocyte clones.
Authors: Blanchard D , van Els C , Borst J , Carrel S , Boylston A , de Vries J E , Spits H
Published: December 2022

The cytotoxic reaction mediated by cytotoxic T lymphocytes (CTL) consists of three phases: first, the CTL binds to the target cell; next, the CTL is triggered to lyse the target cell; and in the third phase, the CTL detaches from the target cell which is lysed in the absence of the CTL. Recently, we obtained evidence that human alloreactive CTL clones initially adhere to target cells without the involvement of the interaction between the T cell receptor (Tcr) and its specific target antigen. In the present study, we investigated the effect of monoclonal antibodies specific for the Tcr on the cytotoxic reaction of three CD8+ HLA-A2-specific CTL clones, using a single cell assay in which the binding event can be distinguished from the post-binding (lytic) phase of the cytolytic reaction. It was found that monoclonal antibodies directed at a variable part of the Tcr do not affect the binding phase but strongly block the lytic phase of the cytotoxic reaction. An anti-constant region Tcr antibody and an anti-CD3 reagent had a similar effect on the two phases of the reaction as the anti-variable part Tcr antibodies. In contrast, antibodies specific for LFA-1 strongly blocked the adhesion phase but did not affect the lytic phase. Antibodies specific for CD-8 had intermediate effects. They could block both the adhesion as well as the lytic phase. The effect of anti-CD8 appeared to be dependent on the CTL clone tested. One clone was found to be inhibited in the adhesion phase, but not in the lytic phase, whereas anti-CD8 hardly blocked the adhesion phase of two other CTL clones, but affected the lytic step of those clones. Our data indicate that LFA-1 is a major adhesion molecule in the CTL reaction, whereas the Tcr/CD3 complex is implicated in a phase after the initial formation of conjugates. CD8 is associated with both steps in the cytolytic reaction. In addition to its minor role in the adhesion phase, our data suggest strongly that CD-8 is involved in the triggering phase of the cytolytic reaction.
In vitro and in vivo activation of human mononuclear phagocytes by interferon-gamma. Studies with normal and AIDS monocytes.
Authors: Murray H W , Scavuzzo D , Jacobs J L , Kaplan M H , Libby D M , Schindler J , Roberts R B
Published: December 2022

To determine the potential immunotherapeutic role of interferon-gamma (IFN-gamma) as a mononuclear phagocyte-activating agent, we examined the effector cell function of peripheral blood monocytes from healthy donors and acquired immunodeficiency syndrome (AIDS) patients after either in vitro and/or in vivo treatment with recombinant (r) IFN-gamma. When assayed immediately after a 24-hr in vitro pulse with 300 U/ml, normal and AIDS monocytes behaved similarly with little augmentation of their intrinsically high levels of H2O2 release and activity against Toxoplasma gondii; in contrast, activity toward the more resistant intracellular pathogen, Leishmania donovani, was appreciably enhanced by rIFN-gamma. In addition, upon testing 4 to 6 days after in vitro pulsing, both normal and AIDS monocytes showed clear evidence of persistent activation in all three assays. The capacity of IFN-gamma to similarly activate monocytes in vivo was confirmed in all ten treated AIDS patients by examining cells before and after 24-hr infusions of 0.03 and 0.5 mg of rIFN-gamma/square meter (M2) of body surface area. For postinfusion monocytes tested after 1 day in culture, H2O2 release and antitoxoplasma activity were essentially unchanged, but antileishmanial effects were augmented. After 5 to 7 days in culture, monocytes from treated patients showed 3.2- to 5.9-fold increases in H2O2-releasing capacity and increases of 49 to 68% and 35 to 61% in intracellular activity against T. gondii and L. donovani, respectively. These results indicate that the human monocyte can be induced by rIFN-gamma to express signs of both immediate and persistent activation and suggest that, as a direct activator of mononuclear phagocytes, rIFN-gamma may also have potential as an immunotherapeutic agent for patients with intracellular infections.
Anti-T cell monoclonal antibodies in vivo. II. Modulation of acute and chronic experimental allergic encephalomyelitis (EAE) in guinea pigs.
Authors: Sobel R A , Blanchette B W , Russo M A , Williams A M , Colvin R B
Published: December 2022

To determine the effects of anti-T cell monoclonal antibody-induced systemic T cell depletion in neuro-autoimmune disease, we studied the in vivo effects of 8BE6, a mouse anti-guinea pig (GP) pan-T cell monoclonal antibody, on the course and immunopathology of the disease model experimental allergic encephalomyelitis (EAE) in adult Strain 13 GP. Central nervous system (CNS) tissues were studied by routine histology and by an immunoperoxidase staining technique using monoclonal antibodies to T cells, IgM, and macrophages. From 3 days before to 10 days after sensitization with GP spinal cord and complete Freund's adjuvant, the GP were given one or two i.p. doses of 3.4 mg 8BE6 or MOPC 21, the parent mouse myeloma ascites, or normal saline. Eighteen of 18 control-treated GP developed typical acute, paralytic EAE 11 to 21 days after sensitization, whereas acute EAE was prevented in 33 of 49 8BE6-treated GP (67%), and the onset was delayed and disease progression was slowed in the others. Five GP treated with 8BE6 from days 11 to 14 after sensitization, at the onset of neurologic signs, rapidly deteriorated within hours after treatment and had loss of T cell staining, and lymphocytolysis in the CNS. 8BE6-treated GP which did not develop acute EAE were observed daily for up to 700 days (mean = 213 days). Twenty-nine of 39 (74%) had from one to six relapses or fixed neurologic deficits. GP in relapse were additionally treated with 8BE6 (22), MOPC-21 (5), or saline (6) in a cross-over protocol. Clinical scores were improved from days 2 to 12 after treatment (p less than 0.05), and complete recovery within 30 days occurred more frequently (p = 0.046) and more rapidly (p less than 0.01), after 8BE6 as compared with control treatments. Recoveries occurred more often if 8BE6 was given early in the relapse. Multiple treatments led to dose-dependent levels of serum antibodies to mouse immunoglobulin detected by an ELISA. There were no differences between acute and chronic EAE in numbers of inflammatory foci or numbers of macrophages and T cells in CNS infiltrates, but GP with chronic EAE had more extensive demyelination and vascular fibrosis and more numerous IgM+ B cells in parenchymal and meningeal infiltrates than in acute EAE (p less than 0.001).(ABSTRACT TRUNCATED AT 400 WORDS)
Interleukin 1 modulates messenger RNA levels of lymphokines and of other molecules associated with T cell activation in the T cell lymphoma LBRM33-1A5.
Authors: Hagiwara H , Huang H J , Arai N , Herzenberg L A , Arai K , Zlotnik A
Published: December 2022

We have investigated the mechanism of action of IL 1 on T cell activation. For this purpose, we analyzed the content of specific messenger RNA for lymphokines and other genes that are associated with T cell activation in the murine IL 1-dependent T cell lymphoma LBRM33-1A5. Using cloned genes for IL 2, IL 3, TGF-beta, TY5, IL 2 receptor, Ly-1, c-myc, and p53 as probes in the S1 nuclease protection assay, we compared the amount of specific transcripts among total RNA prepared from unstimulated cells, IL 1 alpha or PHA-stimulated cells, and PHA plus IL 1 alpha-stimulated cells. IL 1 alpha augmented the PHA-induced accumulation of IL 2 mRNA with a magnitude comparable to the amount of IL 2 produced, suggesting that IL 1 alpha modulates IL 2 gene expression at the RNA level. Similar results were obtained with IL 3. We also observed that Ly-1 mRNA appears after PHA treatment and its accumulation was augmented by IL 1 alpha addition. On the basis of the effects of IL 1 alpha and/or PHA treatments on gene expression, we classified these genes into four groups. In all cases, IL 1 alpha seemed to affect mRNA levels quantitatively. These observations support previously described characteristics of this cytokine as a co-stimulator of T cell activation.
A B cell-specific differentiation antigen, CD23, is a receptor for IgE (Fc epsilon R) on lymphocytes.
Authors: Yukawa K , Kikutani H , Owaki H , Yamasaki K , Yokota A , Nakamura H , Barsumian E L , Hardy R R , Suemura M , Kishimoto T
Published: December 2022

Two independent L cell transformants expressing human lymphocyte Fc epsilon R were established by using cellular DNA from RPMI 8866 cells. The surface expression of the receptor was confirmed on the basis of the binding of a panel of anti-Fc epsilon R antibodies and its ability to bind IgE. Anti-CD23 antibodies strongly stained the transformants, indicating possible identity or antigenic relationship between Fc epsilon R and CD23. This interesting observation warrants additional clarification as to the role of CD23 and Fc epsilon R in B cell differentiation.
Deficiency of the C3b/C4b receptor (CR1) of erythrocytes in systemic lupus erythematosus: analysis of the stability of the defect and of a restriction fragment length polymorphism of the CR1 gene.
Authors: Wilson J G , Wong W W , Murphy E E , Schur P H , Fearon D T
Published: December 2022

The role of genetic factors in controlling CR1 quantitative expression on erythrocytes (E) of patients with systemic lupus erythematosus (SLE) was reexamined by determining the temporal stability of CR1 numbers and the frequency of a CR1 genomic restriction fragment length polymorphism (RFLP). The mean number of binding sites/(E) for Yz-1 monoclonal anti-CR1 correlated with the number of sites for polyclonal anti-CR1 that had been determined 2 to 4 yr previously in 18 normal persons (p less than 0.001), 18 patients (p less than 0.001), and 28 relatives (p less than 0.001), indicating that CR1 sites/E was a stable characteristic in all three groups. The mean number of Yz-1 sites/E was 281 +/- 34 (+/- SEM) in 28 probands with SLE and 457 +/- 21 in 93 relatives, both determinations being less than that for 100 normal persons, 553 +/- 21 (p less than 0.002). Thirty-six patients and 51 normal individuals were also assessed for the presence of the 7.4 kb and 6.9 kb HindIII CR1 allelic restriction fragments that correlate with high and low expression, respectively, of CR1 on E. The distribution of patients differed from normal (p less than 0.05), with a smaller proportion being homozygous for the 7.4 kb allele. In addition, the mean numbers of Yz-1 sites/E for patients and relatives who were homozygous (p less than 0.02) and heterozygous (p less than 0.05) for the 7.4 kb allele were significantly lower than those for normal persons matched for the HindIII RFLP, suggesting the existence of additional heritable factors that decrease CR1 expression. The stability over time of the CR1 deficiency among patients, the finding of decreased CR1 number among an expanded group of relatives, the altered frequency among patients of CR1 alleles defined by the HindIII RFLP, and the decreased expression of CR1 on E among patients and relatives compared with normal individuals having the same HindIII RFLP indicate a role for genetic factors in CR1 deficiency in SLE.
Antigen presentation by hapten-specific B lymphocytes. IV. Comparative ability of B cells to present specific antigen and anti-immunoglobulin antibody.
Authors: Lichtman A H , Tony H P , Parker D C , Abbas A K
Published: December 2022

The ability of trinitrophenyl (TNP)-binding murine B lymphocytes to present native rabbit IgG (RGG), TNP-modified RGG, and rabbit anti-mouse Ig (RAMG) to an Ia-restricted, RGG-specific helper/inducer T cell clone was compared. By three independent assays (lymphokine secretion, T cell proliferation, and B cell differentiation), TNP-RGG was presented at 10(2)- to 10(3)-fold lower concentrations than RGG, and RAMG at 10(2)- to 10(3)-fold lower concentrations than TNP-RGG. The available data suggest that the efficiency of antigen presentation is dependent primarily on the avidity of binding of a ligand to B cell surface Ig and/or the extent of subsequent endocytosis (modulation). Despite the observed quantitative differences between anti-Ig (RAMG) and specific antigen (TNP-RGG), these results demonstrate that qualitatively both are essentially similar in their ability to mediate specific T-B interactions. Thus, anti-Ig antibodies are valid models for analyzing cognate interactions between antigen-specific B and helper T lymphocytes.
T3-T cell receptor (Ti) complex-independent activation of T cells by wheat germ agglutinin.
Authors: Yachie A , Hernandez D , Blaese R M
Published: December 2022

The T3-Ti complex appears to play a central role in the activation of T cells by antigens and mitogens. Wheat germ agglutinin (WGA) is a unique lectin which inhibits T cell proliferation induced by mitogens, but it also induces marked IL 2 production by peripheral blood T cells. The pattern of responses induced by WGA suggests that this lectin may use a different mechanism of T cell activation other than the mechanism employed by the common T cell stimulants. We first investigated the production of IL 2 by Jurkat cells (E6-1) stimulated with WGA, before and after modulation of the surface T3-Ti complex. IL 2 production was markedly reduced after modulation of the T3 antigen from the cell surface when these cells were stimulated with PHA. In contrast, little change was observed in WGA-induced IL 2 production after modulation. Furthermore, we examined the effect of WGA on a T3-mutant of E6-1 cells (T3.1) which does not produce IL 2 in response to PHA or PHA plus PMA. WGA-stimulated T3.1 cells produced a significant amount of IL 2 with or without added PMA. In addition, a small but consistent rise in intracytoplasmic free calcium was observed when these cells were stimulated with WGA. These results demonstrate the presence of an alternative mechanism of T cell activation independent of the T3-Ti complex.
Human Fc gamma receptors: stable inter-donor variation in quantitative expression on platelets correlates with functional responses.
Authors: Rosenfeld S I , Ryan D H , Looney R J , Anderson C L , Abraham G N , Leddy J P
Published: December 2022

Evidence has recently been presented that a 40,000 dalton membrane sialoglycoprotein (p40) shared by monocytes and granulocytes serves as the human platelet receptor for aggregated IgG. We now report that the platelets of normal donors exhibit stable quantitative differences in the expression of this receptor molecule, as determined by flow cytometry using fluorescent staining with murine monoclonal antibody to p40 (mab IV.3). These inter-donor differences were reproducible on repeated testing over at least 4 mo. In concurrent assays, the binding of mab IV.3 to each donor's platelets was highly correlated with the binding of heat-aggregated human IgG, also assayed by flow cytometry. The biological relevance of this quantitative variation in IV.3 binding is suggested by its reproducible correlation with platelet responsiveness to aggregated IgG measured by aggregometry. Such stable quantitative variation in platelet Fc receptor expression among individual humans could contribute to differences in severity of certain pathologic processes initiated by IgG-containing immune complexes.
Class-specific regulation of anti-DNA antibody synthesis and the age-associated changes in (NZB x NZW)F1 hybrid mice.
Authors: Sekigawa I , Okada T , Noguchi K , Ueda G , Hirose S , Sato H , Shirai T
Published: December 2022

Investigations of regulatory helper and suppressor T cells in the in vitro anti-DNA antibody synthesis in NZB x NZW (B/W) F1 hybrid mice were initiated by the development of an in vitro system in which G10-passed B cells from B/W F1 mice were cocultured with mitomycin C-treated T cells in the presence of Con A and either in the presence or in the absence of LPS. It was revealed that each IgG and IgM anti-DNA antibody synthesis was under the regulation of separate L3T4+ helper and Ly-2+ suppressor T cells. The function of these class-specific regulatory T cells was age-dependent. Although the helper effect of L3T4+ T cells on IgG antibody synthesis increased, the effect of L3T4+ T cells on IgM antibody production decreased in B/W F1 mice with aging. The IgG anti-DNA antibody production in the cocultures of L3T4+ T cells and B cells was suppressed by addition of Ly-2+ T cells from young but not aged B/W F1 mice, whereas the production of IgM anti-DNA antibodies was suppressed by Ly-2+ T cells from aged but not young B/W F1 mice. We also found that although IgM anti-DNA antibody-producing B cells were already present in 2-mo-old mice, B cells producing IgG antibodies under the influence of L3T4+ T cells appeared in mice at 7 mo of age. These data clearly indicate that separate class-specific regulatory T cells are involved in the production of IgM and IgG anti-DNA antibodies and that the total serum level of the antibodies is reflected by both their age-associated changes and the generation of antibody-forming B cells in B/W F1 mice.
The interleukin 1 receptor. Dynamics of interleukin 1 binding and internalization in T cells and fibroblasts.
Authors: Mizel S B , Kilian P L , Lewis J C , Paganelli K A , Chizzonite R A
Published: December 2022

In this study, we have demonstrated that a murine T cell lymphoma, EL 4, and a murine fibroblast cell line, Swiss 3T3, possess a single class of high affinity interleukin 1 (IL 1) receptors that exist in a dynamic state of equilibrium that is influenced by IL 1. In the absence of IL 1, the IL 1 receptor appears to turnover with a t1/2 of approximately 11 hr. However, when cells are incubated in the presence of IL 1, the IL 1 receptor undergoes extensive ligand-induced down-regulation. IL 1 itself is internalized at 37 degrees C; 50% of the surface-bound IL 1 is internalized in 60 to 120 min. IL 1 does not undergo degradation for at least 6 hr after internalization. By using electron microscopy and autoradiography, we observed several important features of the internalization process. When cells having bound 125I-IL 1 at 4 degrees C were shifted to 37 degrees C, IL 1 moved from the cell membrane to the cytoplasm where it was found in proximity to nuclei or within lysosomes. IL 1 appeared to progressively accumulate in nuclei. Six hours after shifting cells to 37 degrees C, 30 to 35% of the internalized 125I-IL 1 is associated with the cell nucleus. The accumulation of relatively high levels of IL 1 in the nucleus raises the interesting possibility that IL 1 may not only interact in a highly specific manner with cell surface receptors, but also with potentially important nuclear receptors.
Purification and biochemical characterization of a human autocrine growth factor produced by Epstein-Barr virus-transformed B cells.
Authors: Buck J , Hämmerling U , Hoffmann M K , Levi E , Welte K
Published: December 2022

Autocrine growth factor for Epstein-Barr virus-transformed human B cells (aBGF), a protein that is constitutively produced by the human EBV-transformed B cell line 5/2, has been purified from serum-free conditioned medium. The purification involved sequential ammonium sulfate precipitation, ion exchange chromatography, gel filtration, and reversed-phase high performance liquid chromatography. The purified protein has a m.w. of 16,000 in NaDodSO4/polyacrylamide gel electrophoresis and an isoelectric point between 7.0 and 8.0. The relative molecular mass 16,000 form exists in equilibrium with dimeric and tetrameric forms. aBGF supports the growth of EBV-transformed B cells, which have been deprived of their own conditioned medium. The purified aBGF is fully effective at 0.5 ng/ml and has no interleukin 1 activity in the lymphocyte activation factor assay. Because several randomly selected lines of EBV-transformed cells and one EBV-negative lymphoma cell line both produce aBGF activity and show growth dependency on aBGF and because stimulation of normal B cells with anti-immunoglobulin M is increased by aBGF, we propose that aBGF has general significance for growth control of human B cells.
cDNA cloning and sequencing reveals that the electrophoretically constant DR beta 2 molecules, as well as the variable DR beta 1 molecules, from HLA-DR2 subtypes have different amino acid sequences including a hypervariable region for a functionally important epitope.
Authors: Wu S K , Yabe T , Madden M , Saunders T L , Bach F H
Published: December 2022

Two-dimensional gel electrophoresis (2D-PAGE) of DR molecules from three different Dw subtypes (Dw2, Dw12, and FJO) of the HLA-DR2 haplotype reveals that at least two DR beta genes are expressed. Protein mixing experiments demonstrate that one of the two expressed DR beta molecules is electrophoretically variable (referred to as DR beta 1), and the other (DR beta 2) migrates constantly among DR2 subtypes. We have constructed cDNA libraries from Dw12 and FJO homozygous typing cells (HTC DHO for Dw12 and HTC FJO for FJO) and isolated DR beta cDNA clones. Four of these clones (FJO-13, DHO-8, FJO-6, and DHO-7) were sequenced, and the deduced amino acid sequences were compared with each other and with two published amino acid sequences for the DR beta molecules derived from a DR2-Dw2HTC. Prediction of the migration patterns on 2D-PAGE from the amino acid sequences of these and other DR beta molecules allows the tentative designation of the two full-length cDNA (DHO-8 and FJO-13) as coding for DR beta 2 molecules and the other two cDNA (DHO-7 and FJO-6) for DR beta 1 molecules. Amino acid sequence comparisons also show that the constantly migrating DR beta 2 molecules, as well as the electrophoretically variable DR beta 1 molecules, from Dw2, Dw12, and FJO have different primary amino acid sequences, including a clustered difference in the third hypervariable region of the polymorphic first domain.
Production and characterization of a monoclonal antibody specific for the human lymphocyte low affinity receptor for IgE: CD 23 is a low affinity receptor for IgE.
Authors: Bonnefoy J Y , Aubry J P , Peronne C , Wijdenes J , Banchereau J
Published: December 2022

In the present study a gamma 1 kappa monoclonal antibody, Mab 25, specific for the receptor for the Fc fragment of IgE on lymphocytes (Fc epsilon RL) was established. This antibody was generated after fusion of spleen cells from mice immunized with the EBV-transformed lymphoblastoid cell line RPMI 8866, which is known to express Fc epsilon RL at high density. Mab 25 inhibits strongly the binding of IgE to RPMI 8866 cells and to other Fc epsilon RL-positive EBV-transformed lymphoblastoid cell lines. A 50% inhibition of IgE binding was observed at a Mab 25 concentration of 10 ng/ml. The binding of IgE was also inhibited by Fab fragments of Mab 25, suggesting that the inhibition is not simply due to steric hindrance or to an eventual binding through its Fc portion. Mab 25 only binds to cell lines expressing Fc epsilon RL. Mab 25 immunoprecipitated a single polypeptide with an apparent m.w. of 42 Kd, pI 4.9. The membrane molecule bound to and eluted from a Mab 25 immunoabsorbent had the same apparent m.w. and pI as the Fc epsilon RL purified from an IgE immunoabsorbent. Additionally, when RPMI 8866 cell lysates were cleared with Mab 25, no Fc epsilon RL could be bound to or eluted from an IgE immunoabsorbent. Mab 25 was found to weakly bind to a minor proportion of blood (1 to 4%), tonsil (2 to 9%) and spleen (4 to 5%) mononuclear cells with a low intensity. By double fluorescence analysis, most of the Fc epsilon RL-positive cells were found to be CD 20-positive B lymphocytes. The staining pattern of Mab 25 and the biochemical characteristics of the antigen detected by Mab 25 were comparable to those of the CD 23 Mab. The four CD 23 Mab MHM 6, PL 13, HD 50, and Tü 1 were found to inhibit the binding of IgE. PL 13 was found to totally inhibit the binding of Mab 25 to RPMI 8866 cells, whereas Tü 1 and MHM 6 only partially inhibited Mab 25 binding. HD 50 was unable to block the binding of Mab 25. The finding that different CD 23/Fc epsilon RL-specific monoclonal antibodies recognizing distinct epitopes have in common the capacity of inhibiting the binding of IgE suggests that upon binding they induce a conformational alteration of the Fc epsilon RL resulting in a loss of the IgE binding capacity. In conclusion, our data demonstrate that the CD 23 antigen is a low affinity receptor for IgE on lymphocytes.
Human large granular lymphocytes express high affinity receptors for murine monoclonal antibodies of the IgG3 subclass.
Authors: Anasetti C , Martin P J , Morishita Y , Badger C C , Bernstein I D , Hansen J A
Published: December 2022

We have evaluated the ability of murine monoclonal antibodies to mediate antibody-dependent cell-mediated cytotoxicity (ADCC) by human lymphoid cells. Purified large granular lymphocytes (LGL) and interleukin 2-dependent cloned LGL lines having a CD2+/CD16+/CD3- phenotype were tested as effector cells in an ADCC assay by using a family of IgG isotype switch variant anti-Thy-1.1 antibodies against 51Cr-labeled Thy-1.1+ murine SL2 thymoma target cells, a system in which human cells have no spontaneous cytotoxicity. Cytotoxicity was greatest when using the IgG3, followed in rank order by the IgG2a and IgG2b. No cytotoxicity was observed with the IgG1 antibody. Because the antigen-binding regions of the antibodies are identical, the differences in cytotoxicity directly reflect the relative affinity of LGL Fc receptors for each antibody isotype.
In vivo activation of quiescent B cells by anti-immunoglobulin. I. Induction of latent VH allotype production in adult rabbits by treatment with heterologous antibodies or antibody fragments.
Authors: Metzger D W , Fried V A , Van Cleave V H
Published: December 2022

We demonstrated previously that injection of adult rabbits with homologous anti-VHa1 allotype antibody induces the appearance in serum of genetically unexpected (latent) a1 immunoglobulin (Ig) and a1-like internal images. We have now investigated the mechanism for this induction effect and have found that treatment of animals with polyclonal goat or monoclonal mouse anti-a1 antibody similarly results in production of unexpected a1 determinants. The N-terminal amino acid sequences of deblocked heavy chains showed that latent a1 Ig induced by monoclonal antibody treatment was identical to nominal a1 Ig at 18 of 19 residues, with the one amino acid difference at position 13 probably due to a single nucleotide change. Like nominal a1, these molecules expressed multiple VHa1 framework epitopes, as demonstrated by direct immunoelectron microscopic visualization of immune complexes. Whereas F(ab)'2 fragments of rabbit anti-a1 antibody retained inducing activity, F(ab) fragments were effective only when administered in an aggregated form; this result suggests that cross-linking of surface Ig receptors plays a critical role in the induction process. We conclude that all rabbits contain dormant B cells expressing germ-line-encoded latent allotypes, and that in vivo administration of anti-Ig antibody causes activation of these cells directly rather than through perturbation of an allotype regulatory network.
A high m.w. form of decay-accelerating factor (DAF-2) exhibits size abnormalities in paroxysmal nocturnal hemoglobinuria erythrocytes.
Authors: Kinoshita T , Rosenfeld S I , Nussenzweig V
Published: December 2022

Decay-accelerating factor (DAF) is a 70,000 Mr membrane protein that inhibits the amplification of the complement cascade on cell surfaces. Monoclonal antibodies against different epitopes of the 70,000 Mr DAF (DAF-1) recognize a second band at the position of 140,000 Mr on a Western blot of total red cell ghost proteins or partially pure DAF subjected to electrophoresis under denaturing conditions. Like DAF-1, this polypeptide (DAF-2) has the ability to accelerate decay of the C3 convertase, C4b2a, and to reincorporate into red cell membranes. A population of erythrocytes from patients with paroxysmal nocturnal hemoglobinuria (PNH) lack DAF-1 and also DAF-2. In addition, in some patients' red cells bearing DAF-1 of normal Mr, DAF-2 is 5,000 to 10,000 Mr smaller than normal. The structural basis for these differences in size of DAF and its PNH variants is unknown.
Monoclonal antibodies specific to the alpha-subunit of the mast cell's Fc epsilon R block IgE binding and trigger histamine release.
Authors: Baniyash M , Alkalay I , Eshhar Z
Published: December 2022

In an attempt to block the interactions between IgE and its receptor on mast cells (Fc epsilon R), we have established anti-Fc epsilon R monoclonal antibodies (mAb) by fusion of myeloma cells with mouse splenocytes immunized with irradiated rat basophilic leukemia (RBL) cells. Two anti-Fc epsilon R mAb were obtained (denoted 4.7 and 5.14) that could specifically bind to RBL and mast cells. This binding could be inhibited by IgE. The mAb and their F(ab')2 fragments inhibited 125I-IgE binding to RBL cell and triggered cell degranulation. The Fab' fragments, on the other hand, could only inhibit IgE binding but did not stimulate cell degranulation. Furthermore, these monovalent fragments inhibited RBL and mast cell degranulation induced by IgE-antigen complexes both in vitro and in vivo in the passive cutaneous anaphylaxis reaction. The number of mAb 4.7 and 5.14 molecules bound per RBL cells was similar to that of IgE; nevertheless, mAb 4.7 and 5.14 recognized different epitopes on the IgE receptor. Immunoprecipitation and immunoblotting analysis demonstrated that the mAb reacted with the alpha-subunit of the Fc epsilon R. Our findings establish the anti-Fc epsilon R mAb as a useful reagent for the isolation and characterization of the Fc epsilon R's alpha-subunit and the monomeric (Fab') for blocking the IgE-Fc epsilon R interactions.
Interaction between cytokines and 8-mercaptoguanosine in humoral immunity: synergy with interferon.
Authors: Goodman M G
Published: December 2022

In previous studies it has been demonstrated that a T cell-like differentiation signal is transmitted by C8-substituted guanine ribonucleosides such as 8-mercaptoguanosine (8MGuo) to antigen-stimulated B cells. A large subset of potentially reactive B cells remains unresponsive to antigen even in the presence of signals provided by these nucleosides except when this signal is preceded by a soluble activity present in mixed lymphocyte culture supernatants. Studies with purified preparations of interleukin (IL)-1, IL-2, IL-3, granulocyte-macrophage colony stimulating factor, B cell stimulatory factor 1 (IL-4), and B cell growth factor II (IL-5) indicated that none of these activities is capable of synergizing with 8MGuo to augment B cell responsiveness to antigen. Therefore, supernatants from a number of cloned cell lines were examined for activity that could synergize with 8MGuo, in order to determine the cellular source of this activity. Soluble products secreted by cloned 24/G1 T cells act synergistically with 8MGuo to evoke enhanced antibody responses to specific antigen in populations of purified B cells. Because concanavalin (Con) A-activated 24/G1 cells produce large quantities of interferon-gamma (IFN-gamma), the possibility that interferons might mediate synergy with 8MGuo was investigated. Purified murine IFN-gamma is unable to interact synergistically with 8MGuo; moreover, treatment of active 24/G1 supernatants with monoclonal anti-IFN-gamma antibodies or at pH 2 fails to abrogate their ability to synergize. In contrast to IFN-gamma, when B cells were supplemented with either IFN-alpha or IFN-beta, antigen-dependent synergy with 8MGuo was observed. However, abrogation of IFN-alpha and IFN-beta activity with specific antibodies fails to interfere with synergy between 8MGuo and mixed lymphocyte culture or Con A supernatants. Therefore, it appears that although IFN-alpha and IFN-beta are not responsible for the synergizing activity present in activated T cell supernatants, they nonetheless represent a previously unrecognized source of synergizing activity.
Equilibrium binding parameters of an autoimmune monoclonal antibody specific for double-stranded DNA.
Authors: Braun R P , Lee J S
Published: December 2022

Two techniques have been developed to estimate binding parameters for Jel 241 under equilibrium conditions. Jel 241 is an autoimmune monoclonal antibody derived from an NZB/NZW mouse which binds to double-stranded DNA. Thermal denaturation profiles of poly[d(AT)] were measured in the presence of increasing concentrations of IgG Jel 241. From these data it was estimated that the IgG occludes 12 base-pairs on duplex DNA, and the binding to double-stranded DNA was at least four orders of magnitude greater than to single-stranded DNA. In addition, intrinsic association constants (K(O)) were measured by a gel filtration technique for the interaction of both Fab and IgG Jel 241 to native calf thymus DNA. K(O) for the IgG was only 60-fold greater than for the Fab fragment for which K(O) was 4.4 X 10(4) M-1 at an NaCl concentration of 150 mM. Also, K(O) for the Fab increased dramatically with decreasing ionic strength, suggesting that there are four phosphates involved in the interaction. These techniques should be applicable to most autoimmune antibodies which bind to nucleic acid polymers.
Generation from tumor-bearing mice of lymphocytes with in vivo therapeutic efficacy.
Authors: Shu S Y , Chou T , Rosenberg S A
Published: December 2022

Systemic transfer of sensitized lymphocytes can effectively mediate the regression of established tumors. However, virtually all prior experimental applications of this approach have utilized lymphocytes from animals that have been immunized to reject tumor challenge. A similar source of cells is not available in the human. With the use of a weakly immunogenic murine tumor, MCA 105, we demonstrate here that following in vitro sensitization (IVS) with viable tumor cells and interleukin 2, the nontherapeutic lymphoid cells from mice bearing a progressively growing tumor acquired antitumor reactivity capable of mediating the regression of established pulmonary metastases. Although the IVS system induced nonspecific lymphokine-activated killer-like cytotoxic activity from lymphoid cells of normal as well as tumor-bearing mice, therapeutically active cells could only be generated from cultures initiated with lymphoid cells from tumor-bearing animals, indicating that the IVS was a secondary in vitro immune response. Without other treatment, the IVS cells could mediate antitumor effects. However, low doses of exogenous interleukin 2 administration could enhance their therapeutic efficacy. By in vivo T cell subset depletion with monoclonal antibodies, the primary effector cells were identified as belonging to cytotoxic/suppressor T cell lineage expressing the Lyt-2 phenotype. In addition, these therapeutic effector cells could be further expanded in numbers in vitro with continuous stimulation by tumor cells in the presence of interleukin 2. Compared to the number of cells initiating the culture, as many as 126 times the number of cells were obtained after 9 days of IVS followed by in vitro expansion for an additional 5 days. Studies on the kinetics of the occurrence of the pre-effector lymphocytes during tumor growth revealed that they were readily obtained from draining lymph nodes of mice with a broad range of tumor burdens as well as durations of tumor growth. The ability to generate and expand, in vitro, therapeutically active lymphocytes from tumor-bearing hosts has important implications for cellular therapy of human cancers.
Chronic experimental autoimmune myasthenia gravis induced by monoclonal antibody to acetylcholine receptor: biochemical and electrophysiologic criteria.
Authors: Gomez C M , Richman D P
Published: December 2022

To determine whether the chronic presence of antibody to acetylcholine receptor (AChR) can account for the neuromuscular abnormalities in myasthenia gravis (MG), rats injected repeatedly with monoclonal antibody (mAb) to AChR were compared with those injected with control mAb. In a previous report, those receiving anti-AChR mAb, studied ultrastructurally, had grossly simplified endplates when compared with normal controls. In this report, animals injected once or chronically for 9 to 12 wk had reduced content of muscle AChR. The chronically injected animals also had diminished miniature endplate potential amplitudes, but to a lesser extent than the reduction in AChR content. These studies establish the pathogenetic role of antibody to AChR in the induction of the ultrastructural, biochemical, and electrophysiologic hallmarks of MG.
A fish oil diet inhibits amyloid P component (AP) acute phase responses in arthritis susceptible mice.
Authors: Cathcart E S , Mortensen R F , Leslie C A , Conte J M , Gonnerman W A
Published: December 2022

Amyloid P component (AP) bears close homology with C-reactive protein and behaves as an acute phase reactant in the plasma of mice but not in man. Our aim was to determine whether AP is influenced by diet, gender, and arthritis severity in a murine model of arthritis. B10.RIII mice were segregated according to gender and diet at 8 wk of age: the source of fat was either corn oil, fish oil, or beef tallow (5% by weight). Four weeks later, each mouse was immunized with 100 micrograms fetal bovine type II collagen, and the incidence and severity of arthritis was noted at weekly intervals. AP was measured by competitive ELISA in plasma taken 5 wk and 15 wk after immunization. AP levels were less in fish oil fed males and females. Under all conditions tested AP levels of females were greater than in males. There was a negative correlation between AP levels and the severity of arthritis. We conclude from these data that although AP levels cannot be used as indices of arthritis severity, there are significant dietary and gender effects on AP concentrations as long as 15 wk after immunization with type II collagen.
Gene induction by gamma-irradiation leads to DNA fragmentation in lymphocytes.
Authors: Sellins K S , Cohen J J
Published: December 2022

An early event in death of interphase lymphocytes exposed in vivo or in vitro to low doses of gamma-irradiation is the degradation of DNA into nucleosome-sized fragments. Induction of fragmentation required RNA and protein synthesis because actinomycin D and cycloheximide, respectively, are able to inhibit DNA fragmentation in irradiated lymphocytes. Studies adding cycloheximide and actinomycin D at various times postirradiation suggest that once the metabolic process is initiated within an individual cell it proceeds to completion. The reversible RNA synthesis inhibitor, 5,6-dichloro-1-beta-d-ribofuranosylbenzimidazole inhibits DNA fragmentation in irradiated thymocytes. When this drug is removed after 6 hr, irradiated thymocytes proceed to fragment their DNA; this suggests that an inducing "signal" that is not simply mRNA persists within the irradiated cell for at least 6 hr after irradiation. In contrast to mitogen-activated T and B lymphoblasts, resting T and B cells show significant DNA fragmentation after exposure to 100 to 500 rad. At 2000 rad, all of the splenic subpopulations die rapidly via a different mechanism. By studying the mechanism of DNA fragmentation induced during the interphase death of lymphocytes, we hope to understand better the extreme sensitivity of resting lymphocytes to radiation and what may be the common final pathway of programmed cell death.
Ia-restricted B-B cell interaction. I. The MHC haplotype of bone marrow cells present during B cell ontogeny dictates the self-recognition specificity of B cells in the polyclonal B cell activation by a B cell differentiation factor, B151-TRF2.
Authors: Ono S , Takahama Y , Hamaoka T
Published: December 2022

We have demonstrated that B cell recognition of Ia molecules is involved in polyclonal B cell differentiation by B151-TRF2. The present study was undertaken to examine the Ia recognition specificity of B151-TRF2-responsive B cells in fully major histocompatibility complex (MHC)-allogeneic P1----P2, semiallogeneic P1----(P1 x P2)F1, and double donor (P1 + P2)----(P1 x P2)F1 and (P1 + P2)----P1 radiation bone marrow chimeras. The B cells from both P1----P2 and P1----(P1 x P2)F1 chimeras could give rise to in vitro immunoglobulin M-producing cells upon stimulation with B151-TRF2 comparable in magnitude to that of normal P1 B cells, and their responses were inhibited by anti-I-AP1 but not by anti-I-AP2 monoclonal antibody even in the presence of mitomycin C-treated T cell-depleted P2 spleen cells as auxiliary cells. In contrast, the B151-TRF2 responses of P1 B cells isolated from both (P1 + P2)----(P1 x P2)F1 and (P1 + P2)----P1 double bone marrow chimeras became sensitive to the inhibition of not only anti-I-AP1 but also anti-I-AP2 monoclonal antibody only when the culture was conducted in the presence of P2 auxiliary cells, demonstrating that they adaptively differentiate to recognize as self-structures allogeneic as well as syngeneic Ia molecules. Moreover, the experiments utilizing B cells from H-2-congenic mice and B cell hybridoma clones as auxiliary cells revealed that B151-TRF2-responsive B cells recognize Ia molecules expressed on B cells. Taken together, these results demonstrate that B151-TRF2-responsive B cells recognize Ia molecules expressed by B cells as self-structures and that their self-recognition specificity is dictated by the MHC haplotype of bone marrow cells present during the B cell ontogeny but not by the MHC haplotype of a radiation-resistant host environment.
The inhibitory effects of K+ channel-blocking agents on T lymphocyte proliferation and lymphokine production are "nonspecific".
Authors: Schell S R , Nelson D J , Fozzard H A , Fitch F W
Published: December 2022

The effect of K+ channel-blocking agents, tetraethylammonium (TEA) and 4-aminopyridine (4AP), on the responses of cloned murine helper and cytolytic T lymphocytes stimulated with mitogen, anti-T cell receptor monoclonal antibody, or interleukin 2 was examined. The addition of TEA and 4AP reduced [3H]thymidine incorporation and lymphokine production to levels observed in unstimulated cells. However, thymidine incorporation by the tumor cell lines P-815 and SP2/0, which replicate autonomously, also was inhibited by these drugs. Treatment of cloned murine helper T lymphocyte, L2, with TEA appeared to inhibit uptake of [3H]thymidine and [3H]phenylalanine after stimulation with interleukin 2. These results suggest that the inhibitory effects of the K+ channel-blocking agents TEA and 4AP may not be specific for the sequence of events that are initiated by activation of T lymphocytes through the antigen receptor. Instead, the observed inhibitory effects by these agents may result from inhibition of transport of thymidine, amino acids, and other essential metabolites across the cell membrane.
Functional competency of T cell antigen receptors in human thymus.
Authors: Weiss A , Dazin P F , Shields R , Fu S M , Lanier L L
Published: December 2022

The T cell antigen receptor is likely to play a role in both positive and negative selection in the thymus. Three populations of thymocytes can be distinguished by the level of expression of the CD3-alpha/beta-chain heterodimer of the T cell antigen receptor (CD3/Ti alpha/beta) complex. Cells which fail to express these receptors or express low levels of receptors are contained in a population of thymocytes which express low levels of the CD5 antigen and are predominantly CD4+/CD8+. Thus, these cells appear to be relatively immature phenotypically. In contrast, the cells which express high levels of CD3/Ti alpha/beta co-express high levels of CD5 and are predominantly contained in the more mature single positive cells which express either CD4 or CD8. With the calcium-sensitive dye, Indo-1, and immunofluorescence, we demonstrated that, despite the relative phenotypic immaturity of cells which express low levels of CD3/Ti alpha/beta, these antigen receptors are able to mediate transmembrane signaling when stimulated with CD3 monoclonal antibodies. Although increases in calcium were observed in these CD3/Ti alpha/beta-low expressing cells in response to anti-CD3, no proliferative response was observed, even in the presence of phorbol myristate acetate. Proliferative responses were observed in the more mature cells which express high levels of CD3/Ti alpha/beta. These results suggest that, rather than a defect in the functional capability of the antigen receptor complex to mediate transmembrane signaling events, cellular responses to signals generated by the antigen receptor may differ at various stages of thymocyte development.
Beta-adrenergic-receptor-mediated suppression of interleukin 2 receptors in human lymphocytes.
Authors: Feldman R D , Hunninghake G W , McArdle W L
Published: December 2022

Adrenergic receptor agonists are known to attenuate the proliferative response of human lymphocytes after activation; however, their mechanism of action is unknown. Since expression of interleukin 2 (IL-2) receptors is a prerequisite for proliferation, the effect of beta-adrenergic receptor agonists on lymphocyte IL-2 receptors was studied on both mitogen-stimulated lymphocytes and IL-2-dependent T lymphocyte cell lines. In both cell types the beta-adrenergic receptor agonist isoproterenol blocked the expression of IL-2 receptors, as determined with the IL-2 receptor anti-TAC antibody. To determine the effect of beta-adrenergic agonists on expression of the high affinity IL-2 receptors, [125I]IL-2 binding studies were performed at concentrations selective for high affinity sites. No significant effect of beta-adrenergic agonists on high affinity IL-2 receptor sites could be detected. The data demonstrate that beta-adrenergic receptor agonists down-regulate IL-2 receptors primarily affecting low affinity sites.
The expression of membrane and secreted immunoglobulin during the in vitro differentiation of the murine B cell lymphoma CH12.
Authors: Stockdale A M , Dul J L , Wiest D L , Digel M , Argon Y
Published: December 2022

The expression of membrane and secreted IgM was analyzed during mitogen-induced differentiation of the murine B cell lymphoma CH12. To characterize the Ig genes used by CH12, the nucleotide sequences of the variable gene segments (V mu and V kappa) were determined. The expressed V mu gene segment belongs to the VHII NPb-related family. The D (FL16.1a) and J (JH2) segments are the same as those used by the NP-specific hybridoma B1-8. The V kappa used by CH12 is almost identical to those used by the oxazolone-specific hybridomas NQ5.89.4 and NQ7.7.1. Treatment with lipopolysaccharide (LPS) induces up to 80% of CH12 cells to secrete IgM within 48 hr of culture. The steady state levels of secreted mu (mu s) and kappa mRNA increase four to fivefold over this period in cells stimulated with LPS compared with unstimulated cells. The kinetics are similar for both mRNA and parallel the increase in IgM secretion. EL-4 supernatants induce comparable changes in m mu s and kappa transcript levels. The simultaneous increase in m mu s and kappa transcripts suggests that coordinate control of RNA levels is used to increase the synthesis of secretory IgM during differentiation. The level of mRNA encoding the membrane form of mu (mu m) remains constant in stimulated cells and increases slightly in unstimulated cells. While the net rates of synthesis of membrane-bound mu-chains remain similar during LPS stimulation, the level of surface IgM on secreting cells is reduced three to fivefold. These observations suggest that the level of surface IgM expression during differentiation of CH12 is controlled largely by post-translational mechanisms. Our results demonstrate that the CH12 cell line regulates the expression of membrane and secreted IgM differently during its differentiation. The changes in IgM expression in CH12 parallel those occurring in normal B cells after mitogen or antigen challenge. Thus, the in vitro differentiation of CH12 is a good model for the analysis of late stages of B cell development.