Release of lymphokines following Epstein-Barr virus infection in vitro of blood lymphocytes from patients with autoimmune diseases.
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Adult↗
Biomedical subjects
D A Carson
Publications and source records attributed to D A Carson.
Lymphoblastoid B cell lines produce an interleukin-1-like activity that can be serologically distinct from macrophage interleukin-1.
Antigen presentation by macrophages is accompanied by the production of interleukin-1 (IL-1) for successful T cell triggering. We have described EBV infected B cell lines that served as antigen presenting cells (APC) and now report the ability of these cells and their supernates, as well as EBV negative B cell lines, to support IL-2 production by a T cell line JM in the presence of the monoclonal antibody that recognizes the CD3 complex. Both EBV-transformed B cell lines and EBV-negative lines, and their supernates, exhibited IL-1-like activity in two different IL-1 dependent assays. These IL-1-like molecules were released constitutively from five of six EBV positive lines and three of five non-infected B cell lines. The activity from both, one virally infected and one non-infected B cell line, eluted from Sephadex G-75 in two peaks at 15-18K and 30-35K and could not be neutralized by antibody specific for macrophage IL-1. Supernates having IL-1-like molecules also contained a higher molecular weight inhibitor of proliferation. These data indicate that: both EBV-positive and EBV-negative B cell lines are capable of elaborating the IL-1-like activity; and the IL-1-like activity from these cells can be serologically distinct from macrophage IL-1. This suggests the presence of a family of molecules with IL-1 activity that derive from different cell types.
Animals↗
Expression of a cross-reactive idiotype on rheumatoid factor in patients with Sjogren's syndrome.
Primary Sjogren's syndrome (SS) is a systemic autoimmune disorder characterized by lymphocytic infiltration of salivary and lacrimal glands. These patients have evidence of marked B cell hyperactivity, including the production of autoantibodies such as rheumatoid factor (RF) and an increased frequency of non-Hodgkin's lymphoma. We now demonstrate that RF from 12/15 SS patients contains a cross-reactive idiotype (CRI) on their kappa light chain defined by a monoclonal antibody (MoAb 17-109) and immunoblotting. This CRI was associated with immunoglobulin (Ig) A-RF, and to a lesser extent with IgM-RF molecules on the basis of direct binding studies. With the use of immunoperoxidase techniques to stain frozen tissue sections, B cells containing cytoplasmic Ig reactive with MoAb 17-109 were detected in the salivary gland biopsies from 11/12 SS patients at high frequencies, and in the blood from the same patients at much lower frequencies. One patient with pre-existant SS developed non-Hodgkin's lymphoma with tumor cells and RF paraprotein reactive with MoAb 17-109. Evaluation of serial biopsies over a 4-yr period showed a progressive increase in the proportion of B cells bearing the CRI. In contrast, synovial membrane biopsies from RA patients lacking sicca symptoms did not contain B cells expressing the CRI. Because previous studies have demonstrated that MoAb 17-109 detects a CRI on RF paraproteins from patients with lymphoma, B cells bearing this CRI may have increased frequency of neoplastic transformation. SS patients provide an opportunity to study the expression of this CRI and to understand the transition of B cell clones from autoimmune proliferation to neoplastic transformation.
Adolescent↗
Inhibition of anti-CD3 (T3) antibody-induced activation of T cells by cyclosporine.
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Antibodies, Monoclonal↗
Characterization of a crossreactive idiotype in Sjögren's syndrome.
Sjögren's syndrome (SS) is characterized by lymphoid infiltration of the salivary glands and autoantibody production. Rheumatoid factor (RF) in patients with primary SS (1 degree SS) contains a crossreactive idiotype (CRI) defined by a monoclonal antibody (MoAb 17-109). This CRI was located on the kappa light chain by immunoblotting methods. A high frequency of CRI+ B cells was found in SS salivary gland biopsies, suggesting this tissue as the site of production for this autoantibody. Further characterization of CRI+ RF from SS patients was performed using antibodies prepared against synthetic peptides corresponding to the hypervariable region of RF paraproteins from patients with Waldenström's macroglobulinemia (WM). These results demonstrate a close structural relationship between RF in SS and WM patients. To analyze the genes that encode these RF in SS patients, B cell hybridomas that secreted CRI+ immunoglobulin were created and their DNA analyzed by Southern blot techniques. These hybridomas will allow us to determine the DNA sequence of kappa genes encoding the CRI and to identify adjacent regulatory genes that may promote high levels of CRI expression.
Cross Reactions↗
The common occurrence of internal image type anti-idiotypic antibodies in rabbits immunized with monoclonal and polyclonal human IgM rheumatoid factors.
We have previously reported that rabbits immunized with a polyclonal human rheumatoid factor (RF) autoantibody preparation could induce anti-idiotypic antibodies bearing the 'internal image' of the Fc fragments of human IgG. The 'internal image' anti-idiotype have been shown to react with both the RF molecules as well as with the RF receptors on B lymphocytes. Under what conditions these anti-idiotypes occur is not known. Presently, we report that these anti-idiotypic antibodies occur more frequently than previously thought and could be isolated in sera of rabbits immunized with either monoclonal paraproteins with RF activity or other purified human polyclonal serum RFs. Immunization of rabbits with a peptide corresponding to the second complementarity-determining regions of a monoclonal RF did not induce this anti-idiotype. Immunization of goats with human RF similarly did not result in induction of such anti-idiotype. Induction of these anti-idiotypes thus depended upon immunization with the intact RF antigen as well as the species of animal immunized. The repeated isolation of 'internal image' anti-idiotypic antibodies from RF immunized rabbits suggests that the antigenic conformations recognized by human RF autoantibodies are restricted, and that 'internal image' anti-idiotypic species to RF may pre-exist within the rabbit immune network. Such broadly cross-reactive anti-idiotypic reagents provide unique reagents for studying the regulation of RF autoantibody synthesis.
Animals↗
Resistance to multiple adenine nucleoside and methionine analogues in mutant murine lymphoma cells with enlarged S-adenosylmethionine pools.
Adenosine and many adenosine analogues exert toxicity to mammalian cells at the nucleoside level. The mechanism of action of these agents is controversial. Previous experiments suggested that adenosine toxicity could be mediated by the accumulation of S-adenosylhomocysteine (AdoHcy), a potent inhibitor of S-adenosylmethionine (AdoMet) dependent methylation reactions. To analyze this question genetically, adenosine resistant, adenosine kinase deficient mutant clones of a murine T-lymphoma cell line (R1.1) have been selected and analyzed. Compared to parental lymphoma cells, the adenosine resistant mutants had severalfold elevated levels of AdoMet and an increased AdoMet:AdoHcy ratio. The activity of methionine adenosyltransferase was also raised in the mutants. The mutant cells were cross-resistant to agents postulated to cause accretion of AdoHcy, formation of AdoHcy analogues, impairment of AdoMet synthesis, or direct interference with AdoMet dependent reactions. These included 3-deazaadenosine, carbocyclic adenosine, carbocyclic 3-deazaadenosine, formycin A, 8-azaadenosine, 5'-deoxy-5'-methylthiotubercidin, 5'-deoxy-5'-methylthioadenosine, 5'-deoxy-5'-S-isobutylthioadenosine, adenine, cycloleucine, L-ethionine, seleno-DL-ethionine, and (+/-)-2-aminobicyclo[2.1.1]hexane-2-carboxylic acid. These results suggest that diverse purine nucleoside and methionine analogues may block the growth of adenosine kinase deficient cells by interference with AdoMet synthesis and degradation. An increase in AdoMet pools can render mammalian cells cross-resistant to multiple drugs affecting this essential metabolic pathway.
Adenosine↗
IgM rheumatoid factor autoantibody and immunoglobulin-producing precursor cells in the bone marrow of humans.
The natures of the IgM rheumatoid factor (RF)-, IgM-, and IgG-secreting cells in the human bone marrow as compared to the peripheral blood, have been investigated by (1) response to the polyclonal B-cell activator, the Epstein-Barr virus (EBV), (2) sensitivity to the S-phase specific antimetabolite hydroxyurea, (3) presence of the BA-1 and Ia antigens on the cell surface, and (4) cell size, as determined by counter flow elutriation. The EBV-inducible bone marrow IgM-RF precursors derived from medium to large B cells that were inhibited by hydroxyurea pretreatment. The marrow total IgM response derived from small to medium size cells, and was only partially inhibited by hydroxyurea. Hydroxyurea had no effect on IgM-RF or IgM synthesis by peripheral blood cells. These results indicate that the marrow EBV-induced IgM-RF response is not representative of the response by peripheral blood cells, moreover; the marrow RF secreting response arises from a dividing cell pool that may represent newly generated autoreactive B cells.
Aging↗
Characterization of human rheumatoid factors with seven antiidiotypes induced by synthetic hypervariable region peptides.
Recently, we have used synthetic peptides corresponding to the complementarity-determining regions (CDR) of Ig molecules to induce antiidiotypic antisera. Peptide PSH3, representing the third CDR of the IgM rheumatoid factor (RF) Sie heavy (H) chain, induced a private antiidiotype that reacted with only one out of five IgM-RF. Peptide PSL2, corresponding to the second CDR of Sie light (L) chain, induced an antibody against a crossreactive idiotype (CRI), expressed by 10 out of 12 human IgM-RF analyzed. Herein, we report that five additional antiidiotypic antibodies were generated by immunization with synthetic peptides identical to the third L chain CDR of IgM-RF Sie (PSL3), the second and third H chain CDR of IgM-RF Wol, and the second and third CDR of IgM-RF Pom. As analyzed by immunoblot assay, both anti-PSL3 and anti-PSL2 reacted with the majority of 16 IgM-RF. In contrast, all five antiidiotypes induced by the H chain peptides reacted only with the parent proteins, except anti-PSH3, which reacted weakly with one additional RF. These results suggest that one (or very few) VL gene(s), but a larger number of VH genes, are used to encode IgM-RF autoantibodies.
Antibodies, Anti-Idiotypic↗
Synthetic peptides corresponding to third hypervariable region of human monoclonal IgM rheumatoid factor heavy chains define an immunodominant idiotype.
Synthetic peptides corresponding to eight individual heavy chain complementarity-determining regions (CDR) of three human monoclonal IgM anti-IgG (rheumatoid factor [RF]) paraproteins elicited rabbit antibodies with markedly different properties. All antisera recognized the immunizing peptide, and several reacted with the isolated IgM heavy chain on immunoblots. However, only the antisera against peptides representing the third CDR bound consistently and specifically to the intact IgM-RF molecule. These data indicate that the third CDR of human mu chains comprises an immunodominant idiotype, and suggest that the D gene segment may be especially important in creating idiotypic diversity. Synthetic peptides corresponding to the third heavy chain CDR of human paraproteins may be clinically useful for the specific induction of antiidiotypic antibodies.
Amino Acid Sequence↗
Differential regulation of polyamine synthesis and transmethylation reactions in methylthioadenosine phosphorylase deficient mammalian cells.
The consumption of S-adenosylmethionine during polyamine synthesis and transmethylation reactions yields stoichiometric amounts of 5'-deoxy-5'-methylthioadenosine and S-adenosylhomocysteine, respectively. Information concerning the regulation of the two routes of S-adenosylmethionine metabolism in viable cells under changing growth conditions is limited. The present experiments have measured the time-dependent accumulation of 5'-deoxy-5'-methylthioadenosine and L-homocysteine in the medium of four malignant human and murine cell lines deficient in 5'-deoxy-5'-methylthioadenosine phosphorylase (5'-methylthioadenosine: orthophosphate methylthioribosyltransferase). Included in this group were anchorage-independent and anchorage-dependent cells. The enzyme-deficient cells did not detectably cleave 5'-deoxy-5'-methylthioadenosine, and did not appreciably metabolize homocysteine. A comparison of 5'-deoxy-5'-methylthioadenosine and homocysteine excretion therefore provided a noninvasive method for estimating the relative rates of polyamine synthesis and transmethylation. Early after the release of human CEM lymphoblasts from density dependent growth arrest, 5'-deoxy-5'-methylthioadenosine production increased, and exceeded homocysteine synthesis. 5'-Deoxy-5'-methylthioadenosine formation reached a maximum of 0.9 nmol/12 h per 10(6) cells prior to the onset of exponential growth. The kinetics of homocysteine synthesis were different. Homocysteine accumulation was proportional to the specific growth rate, and achieved a peak of 3.1 nmol/12 h per 10(6) cells during mid-exponential phase, at which time 5'-deoxy-5'-methylthioadenosine production was falling. Similar patterns of 5'-deoxy-5'-methylthioadenosine and homocysteine excretion were observed in other 5'-deoxy-5'-methylthioadenosine phosphorylase deficient cell lines. These data show that polyamine synthesis and transmethylation are differentially regulated during the growth cycle of mammalian cells.
Adenosine↗
Characterization of an epibody. An antiidiotype that reacts with both the idiotype of rheumatoid factors (RF) and the antigen recognized by RF.
Recently, an antiidiotype to human monoclonal IgM anti-IgG autoantibodies (rheumatoid factors) was found to react also with human IgG. This peculiar antiidiotype was called an 'epibody'. We describe the induction of a similar epibody by immunization with a synthetic peptide (corresponding to one hypervariable region of the IgM-RF Glo). The results confirm the existence of epibodies, and provide the possible molecular basis of the epibody phenomenon.
Animals↗
Regulation of Epstein-Barr virus infection by recombinant interferons. Selected sensitivity to interferon-gamma.
Interferons (IFN) are antiviral proteins that may be important in mediating cellular defenses against Epstein-Barr virus (EBV) infection. However, the means by which IFN-alpha, -beta and -gamma modify EBV infectivity are not clear. We have evaluated the effects of purified recombinant preparations of all three classes of IFN on EBV-induced B lymphocyte proliferation and Ig secretion. When added early after EBV infection, all three recombinant IFN reduced B cell outgrowth and Ig secretion. IFN-gamma exerted a 7-10-fold more potent antiviral effect than IFN-alpha or -beta. All three types of IFN act directly on B cells. Monocytes and natural killer cells are not necessary for the anti-EBV activity. Of the three recombinant IFN, only IFN-gamma reduced EBV-induced proliferation and Ig secretion when added 3-4 days after virus infection; IFN-alpha/beta were only effective up to 24 h. B lymphoblastoid lines already transformed by EBV are insensitive to the anti-proliferative actions of all three types of IFN. On the basis of these findings, we propose three phases of regulation during EBV infection. In the early phase, EBV-infected cells can be regulated by all IFN. Subsequently, there is an intermediate period where only IFN-gamma is capable of directly affecting EBV-induced B cell responses. In the third phase, B lymphocytes become insensitive to direct actions of all IFN and are now subject to regulation only by cytotoxic cells.
B-Lymphocytes↗
Phytohemagglutinin binds to the 20-kDa molecule of the T3 complex.
Current findings have suggested that the T3 molecular complex is an essential antigenic signal transducer during T cell activation. Lectins, such as phytohemagglutinin (PHA), activate T cells nonspecifically. Conceivably, lectins may mediate their stimulatory action by affecting the T3 complex. In the present investigation we have studied the involvement of the T3 molecular complex in the PHA-mediated activation of T cells. We selectively modulated the surface expression of T3 molecules by anti-T3 antibody and subsequently tested the ability of the modulated cells to respond to PHA. Reduction of T3 expression by 70% resulted in 80% inhibition of the PHA response. This effect was specific for T3 since modulation of other T cell surface molecules (T4, T8) did not affect the PHA-induced mitogenesis. To determine if PHA could interact directly with the T3 complex, immunoblotting (Western blot) analyses of anti-T3 immunoprecipitates were performed. A 20-kDa member of the T3 complex reacted not only with the anti-T3 antibody, but also with PHA itself. These results provide the first evidence for direct binding of PHA to one of the molecules of the T3 complex. The combined data suggest that a major pathway of PHA-induced T cell activation involves the T3 complex. Possible activation mechanisms are discussed.
Antibodies, Monoclonal↗
The role of the T3 molecular complex on human T lymphocyte-mediated cytotoxicity.
The above overview of the experimental data clearly indicates that the T3 molecular complex is intimately involved in T cell activation. The precise role of the T3 complex in the activation process, however, is not clearly understood. The surface association of the T3 complex with the antigen receptor, along with the ability of antibodies to these molecules to render T cells receptive to IL2, reveal a possible mechanism by which specific antigen initiates T cell activation and growth. However, it would be difficult to reconcile this specific effect of anti-T3 antibodies with their effect on CTL function. Since the T3 complex is not a specific marker of any particular effector T cell population, but it is found on all T lymphocytes, we favor the hypothesis that the complex is involved in a more fundamental step of T cell activation, and we believe that triggering of the 'lethal hit', expression of IL2 receptors, and secretion of IL2 are mere manifestations of this basic process. The natural ligand of the antigen receptor is obviously the specific antigen. However, the natural ligand of the T3 complex is unknown. Possibly, its natural ligand is the antigen receptor itself after it has interacted with antigen. A simple scenario, then of the early events of T cell activation would include antigen recognition and binding, followed by an interaction between the antigen receptor and the T3 complex which then activates or allows expression of specific pathways depending on the particular effector population involved. Thus, the inhibition of CTL function by anti-T3 antibodies could be explained by interference with the antigen receptor-T3 complex interaction following target cell recognition. This interaction may be the event that signals the initiation of the 'lethal hit' process.
Antibodies, Monoclonal↗
Characterization of Epstein-Barr virus-specific T-cell hybridomas derived from infectious mononucleosis.
Somatic cell hybridization was utilized to produce hybrids with surface receptors that would pertain directly to those expressed in vivo during Epstein-Barr virus (EBV) infection. Lymphocytes were obtained during acute infectious mononucleosis (IM) and fused to a double mutant of the JM human T-lymphoma cell lines. Hybrid cells that reacted with autologous EBV-infected lymphoblasts were detected by the release of Interleukin-2 into the culture medium. Reactive hybridomas also released IL-2 following coculture with allogeneic EBV-infected cells when those cells shared HLA-DR antigens of the primary parental cells. In contrast, stimulator cells with no shared HLA-DR or without evidence of EBV infection never induced IL-2 release. These results suggest the existence of a population of T cells that arise during acute IM and could account for the known proliferative phase of the disease. The requirements of IL-2 stimulation are currently under study using this system.
Herpesvirus 4, Human↗
Possible involvement of human D minigenes in the first complementarity-determining region of kappa light chains.
The nucleotide sequences of the complementary strands of two human diversity region (D) minigenes, D2 and D4, show stretches of homology with two human variable region kappa chain (V kappa) genes, NG9 and HK101, respectively, in the first complementarity-determining region. In one V kappa sequence, the homology includes the 5' flanking region of D minigenes, which may comprise a recombinase recognition signal. It is thus conceivable that gene conversions involving D minigenes may contribute to V kappa diversity.
Antibody Diversity↗
Metabolic basis for immune dysfunction in adenosine deaminase deficiency.
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Adenosine Deaminase↗