bluuebunny/crossref_metadata_2025 huggingface dataset:
Newborn Xid mice carry the genetic information for the production of natural autoantibodies against DNA, cytoskeletal proteins, and TNP.
Authors: Dighiero G , Poncet P , Rouyre S , Mazie J C
Published: December 2022

Spleen cells from 6-day-old unimmunized male and female (CBA/N X BALB/c)F1 mice, fused with the nonsecreting hybrid SP2/0, gave 184 hybrids secreting immunoglobulins (183 IgM class; 115 from males and 69 from females) which were screened for antibody (Ab) activity against actin, tubulin, myosin, TNP-BSA, dsDNA, and denatured DNA. Eleven hybrids from the male series (9.65%) and eight hybrids from the female series (11.55%) secreted immunoglobulins with significant Ab activity against at least one of the six antigens tested. Four of these clones reacted with a single antigen, while 15 others reacted simultaneously with at least two antigens. Three hybrid clones exhibited Ab activity against all of the antigens tested. The monoclonality and specificity of the immunoglobulins from tissue culture supernatants were respectively assessed by two-dimensional gel electrophoresis after biosynthetic labeling and competitive enzyme-linked immunosorbent assay. These results confirm our previous findings with newborn and adult BALB/c mice, demonstrating a high frequency of this auto-Ab repertoire, especially in newborn mice, and widespread polyspecificity found among these natural auto-Ab. Xid mice are characterized by the absence of a Lyb-3+,5+ B cell population; nevertheless, these studies suggest--but do not prove--that the Lyb-3-,5- lymphocyte population seems to carry genetic information for the production of natural auto-Ab, because no marked differences were observed in the secretory patterns of hybrids obtained from male mice bearing the Xid defect and from normal female mice. Hence, we postulate that the Xid gene might be a regulatory gene because Xid mice, in contrast with normal mice, are unable to secrete anti-DNA auto-Ab on mitogenic stimulation, despite carrying the genetic information.
Suppression of antigen-specific interleukin 2 production in the MRL/MpJ-lpr/lpr mouse.
Authors: Naides S J
Published: December 2022

Several murine strains with spontaneously occurring systemic lupus erythematosus-like disease demonstrate defects in immunoregulation. The MRL/MpJ-lpr/lpr (MRL-1) strain develops a severe age-progressive defect in interleukin 2 (IL 2) production in response to mitogen or antigen. In this study, we demonstrate in vitro the presence of suppressor cells in the lymph nodes of naive mice of the MRL background. Suppression by MRL-1 lymph node cells was partially reversed by treatment with anti-Lyt-1.2 monoclonal antibody and complement and was moderately radiosensitive. Suppression by lymph node cells from the congenic MRL/MpJ-+/+ (MRL-+) mouse was somewhat more resistant to treatment with anti-Lyt-1.2 and complement, or radiation. Lymph node cells from the H-2-syngeneic mouse strain, C3H/HeJ, failed to suppress. Thus, lymph nodes from mice of the MRL background contain cells capable of suppressing in vitro IL 2 responses. We next performed cell transfers to determine whether suppressor cells contribute in vivo to the IL 2 defect. Lymph node cells, but not spleen cells, from MRL-1 mice by 5 to 6 mo of age suppressed antigen-specific IL 2, CTL, and DTH responses when transferred into young MRL-+ recipients. Transfer of identical numbers of lymph node cells from age-matched MRL-+ mice failed to suppress IL 2 production. Transfer of suppression was sensitive to treatment with monoclonal anti-Lyt-2.1 and complement, and to 250 rad of radiation. Thus, this study suggests a role for active suppression of IL 2 production in the establishment of the IL 2 defect in the MRL-1 mouse. Further, suppression may involve phenotypically distinct T lymphocyte subpopulations.
Surface-bound capsular polysaccharide of type Ia group B Streptococcus mediates C1 binding and activation of the classic complement pathway.
Authors: Levy N J , Kasper D L
Published: December 2022

The role of surface-bound type Ia group B Streptococcus (GBS) capsular polysaccharide in antibody-independent binding of C1 and activation of the classic complement pathway was investigated. In a radiolabeled bacterial-polymorphonuclear leukocyte (PMN) association assay, a measure of bacterial opsonization, preincubation of 3H-type Ia GBS with purified F(ab')2 to the organism blocked the association of the bacteria with PMN', and the inhibitory effect was dose dependent. The specificity of F(ab')2 blocking was shown after adsorption of F(ab')2 with type Ia polysaccharide-sensitized erythrocytes. Polysaccharide-adsorbed F(ab')2 had a 70% decrease in ability to block the association of bacteria with PMN. Evidence for the requirement of the capsular polysaccharide in classic complement pathway activation came from a C1 transfer assay with the use of neuraminidase-digested type Ia GBS. Neuraminidase digestion removed 80% of the terminal sialic acid residues from the native polysaccharide. These neuraminidase-digested organisms had a 72% decrease in binding and transfer of purified C1 compared with non-enzyme-treated organisms. Type Ia capsular polysaccharide bound to sheep erythrocytes promoted classic complement pathway-mediated hemolysis of the cells. The role of C1 inhibitor (INH) in modulation of C1 activation by the organisms was investigated. The possibility existed that the C1 INH could be bound by the bacteria, allowing C1 activation to occur in the fluid phase. The inhibitor was purified from human serum, and its activity was measured before and after incubation with type Ia GBS. The organisms had no effect on C1 INH activity. Thus surface-bound capsular polysaccharide of type Ia GBS mediates C1 binding and classic pathway activation, and this does not involve the C1 INH.
Tyrosine phosphorylation of a 66,000 Mr soluble protein in lectin-activated human peripheral blood T lymphocytes.
Authors: Wedner H J , Bass G
Published: December 2022

Protein phosphorylation was studied in human T lymphocytes stimulated with the mitogenic lectins phytohemagglutinin (PHA) and concanavalin A (Con A). The T lymphocytes were prepared from the venous blood of normal volunteers, their intracellular ATP pools were labeled with [32P]orthophosphate, and protein phosphorylation was assayed in the soluble fraction by two-dimensional gel electrophoresis and autoradiography. When lymphocytes stimulated with PHA or Con A were compared to unstimulated control cells, there was a general increase in protein phosphorylation and the specific phosphorylation of a soluble protein with Mr = 64.9 to 69 KD and pI = 5.6 to 5.8. Phosphorylation of this protein, designated TPP-66, was observed as early as 2 min after the addition of lectin with a gradual increase in the level of phosphorylation over the next 120 min. In the majority of experiments, there was no phosphorylation seen in the unstimulated lymphocytes; however, in some experiments, there was appreciable phosphorylation, which was seen beginning 60 min after the labeling period. When the TPP-66 spot from stimulated lymphocytes was excised from gels, was eluted, and was subjected to limited base hydrolysis followed by single-dimension high voltage electrophoresis, the major phosphorylated residue migrated with phosphotyrosine. In some experiments, there was phosphorylation of serine residues in both the stimulated and control cells; tyrosine phosphorylation was never seen in the unstimulated cell population. These data suggest that, like other stimuli for cell growth, the induction of lymphocyte growth by lectins is associated with the activation of a tyrosine-specific kinase. Thus, tyrosine phosphorylation may play a key role in the transmission of the signal for lymphocyte growth from the exterior to the interior of the cell.
Mechanisms of tumor cell capture by activated macrophages: evidence for involvement of lymphocyte function-associated (LFA)-1 antigen.
Authors: Strassmann G , Springer T A , Somers S D , Adams D O
Published: December 2022

The lymphocyte function-associated (LFA)-1 molecule is expressed on certain populations of macrophages that have an augmented capacity to capture tumor cells. Accordingly, we analyzed the role of LFA-1 in the establishment of such cell-cell interactions. F(ab')2 fragments of the M17/4, anti-LFA-1 monoclonal antibody (MAb) inhibited the interaction between activated macrophages and tumor cells by up to 80% in a dose-dependent manner. The anti-LFA-1 MAb reduced (between 55 to 79%) the number of P815, LSTRA, or EL-4 tumor cells bound to trypsin-sensitive structures on bacillus Calmette Guerin activated macrophages. The inhibition appeared selective, because a F(ab')2 fragment of anti-Mac-1 did not inhibit such binding. Inhibition of tumor cell capture could be observed as soon as 15 min after the onset of the cell-cell interaction between activated macrophages and tumor cells. Optimal inhibition occurred when both tumor targets and macrophages were precoated with the MAb. Although P815, LSTRA, EL-4, and BW5147 tumor cells all expressed LFA-1, only the first three but not BW5147 cells were bound by activated macrophages. Furthermore, endotoxin-pulsed macrophages elicited by thioglycollate broth expressed the LFA-1 antigen but did not exhibit selective tumor cell capture. Finally, anti-LFA-1 inhibited the development of weak into strong binding. Taken together, the results suggest that LFA-1 molecules can participate in the interaction between activated macrophages and neoplastic cells.
Interaction of antigen-specific T cell factors with unique "receptors" on the surface of mast cells: demonstration in vitro by an indirect rosetting technique.
Authors: Kops S K , Ratzlaff R E , Meade R , Iverson G M , Askenase P W
Published: December 2022

Picryl (trinitrophenyl) chloride (PCL) contact sensitization of mice induces T cells that release an antigen-binding T cell factor (PCLF) that plays an important role in the initiation of contact sensitivity responses, in part via activation of mast cells. The current study employs an in vitro indirect rosette assay to demonstrate that PCLF can interact with the mast cell surface. Sheep red blood cells (SRBC) were hapten conjugated with trinitrophenyl (TNP), dinitrophenyl (DNP), or oxazolone (OX). When TNP-conjugated SRBC were coated with PCLF, monoclonal anti-DNP IgE, or anti-DNP IgG1, they produced 40 to 50% rosettes with purified normal mouse peritoneal mast cells. Analogous antigen-binding factors, from lymphoid cells of OX and dinitrofluorobenzene contact-sensitized mice, gave similar mast cell rosetting levels with OX-SRBC and DNP-SRBC, respectively. PCLF demonstrated a high degree of hapten specificity in that it formed rosettes with TNP-SRBC but not with DNP-SRBC, unlike IgE and IgG1, or DNPF, which formed rosettes with either SRBC type. Similarly, soluble TNP-BSA could inhibit PCLF rosette-forming capacity, but soluble DNP-BSA could not. In addition to mouse mast cells, PCLF formed rosettes with rat basophil leukemia cells, mouse peritoneal exudate macrophages, mouse alveolar macrophages, and J 774 cultured mouse macrophages; it did not form rosettes with rat mast cells, rat alveolar macrophages, or mouse spleen cells. Thus, PCLF-formed rosettes were antigen specific, relatively species specific, and mast cell/macrophage specific. PCLF-mediated rosette-forming activity could be detected in the presence of nanogram quantities of PCLF. More than 10 times greater IgE was needed to produce IgE-mediated rosettes. Reduction and alkylation eliminated the rosetting activity of IgE, but the rosetting activity of PCLF was not affected. PCLF, but not IgE rosette-forming activity, could be removed by and eluted from affinity columns linked with a monoclonal antibody specific for T cell-derived antigen-binding factors, whereas PCLF rosetting activity was not retained by an anti-immunoglobulin affinity column. Preincubation of mast cells with rat myeloma IgE or mouse monoclonal IgE of various specificities blocked IgE rosettes but not PCLF-induced rosettes. Other immunoglobulin isotypes likewise did not block PCLF rosettes. However, PCLF rosettes could be blocked by preincubation of mast cells with OX factor (OXF),and OXF-mediated rosettes could be blocked similarly by PCLF. These results suggest that the antigen-binding T cell factor PCLF interacts with a unique receptor on the surface of mouse mast cells.
Quantitative aspects of lupus anti-DNA autoantibody specificity.
Authors: Edberg J C , Taylor R P
Published: December 2022

In this study we have attempted to define the cross-reactive potential of SLE anti-DNA antibodies (in 19 representative sera and plasmas) in both the solution phase and the solid phase. We used the Farr and RBC-CF solution phase assays to measure quantitatively the ability of a variety of negatively charged structurally unrelated molecules to inhibit antibody binding to both native DNA (nDNA) and denatured DNA (dDNA). The inhibitors used were of two types: 1) phospholipids (cardiolipin, phosphatidyl glycerol, and phosphatidic acid) and 2) repeating negatively charged molecules (poly-glutamic acid, heparin sulfate, and chondroitin sulfate). We found in both assays that the phospholipids could inhibit antibody binding to nDNA and dDNA, but a large excess (about 1500-fold) of these molecules was needed relative to DNA to achieve equivalent levels of inhibition. The repeating negatively charged molecules did not inhibit DNA binding at equivalent molar levels as the phospholipids; generally, at least a 10,000-fold excess was needed relative to the nucleic acids to achieve any appreciable inhibition. Results of a dDNA binding-inhibition solid-phase ELISA for cross-reactivity of the anti-DNA antibodies gave quite similar results. Finally, we found that eight of the SLE samples did have anti-cardiolipin antibodies, as demonstrated in a cardiolipin-based ELISA. These results suggest that previous reports describing an apparent cross-reactivity of anti-DNA antibodies may not represent physiologically relevant interactions between anti-DNA antibodies and non-nucleic acid antigens.
Comparison of the N-linked glycopeptides of DQw1 and DR1 molecules.
Authors: Iturbe S , Narasimhan S , Merrick J M , Falk J A , Letarte M
Published: December 2022

HLA class II molecules have been isolated from a [3H]mannose-labeled GM3104 B lymphoblastoid cell line with the phenotype DQw1, DR1. The DQw1 molecules were purified by affinity to 77-34 IgG specifically reactive with the DQw1 specificity. The DR1 molecules were separated into two subsets, DR1a (70 to 80%) and DR1b (20 to 30%), by sequential affinity to 21r5-IgG and 21w4-IgG Sepharose. The alpha- and beta-chains of [3H]mannose-labeled DQw1, DR1a, and DR1b molecules were separated by SDS-PAGE and were recovered by electrophoretic elution. The isolated chains were digested with pronase and the glycopeptides were fractionated by sequential lectin chromatography on immobilized concanavalin A (Con A), Lens culinaris (Lens), and Ricinus communis agglutinin type I (RCA). The N-linked glycopeptides derived from the alpha-chains of DQw1, DR1a, or DR1b showed similar profiles on Con A Sepharose: 45% unbound (ConA I), 25% weakly bound (ConA II), and 30% tightly bound (ConA III). The glycopeptides derived from the beta-chains of DQw1 or DR1 molecules were found almost exclusively (80%) in the fraction unbound to Con A Sepharose, with only 11% and 9% in ConA II and ConA III fractions, respectively. The observation that most of the binding to Con A is associated with the alpha-chain glycopeptides suggests that binding of membrane-associated class II molecules to that lectin must be mediated by the alpha-chains. Binding to Lens Sepharose was higher for beta-(50%) than for alpha-(15%) chain glycopeptides, suggesting that within the intact glycoproteins, the beta-chains are responsible for the interaction with Lens. The ConA I fractions derived from the alpha-chain glycopeptides of either DQw1 or DR1 molecules were separated on RCA-agarose as follows: 60% unbound, 17% retarded, and 20% bound and eluted with 0.1 M galactose. The ConA I fractions derived from the beta-chain glycopeptides of either subset of class II molecules also had a similar profile on RCA-agarose: 70% unbound, 16% retarded, and 10% bound and eluted specifically. After removal of sialic acid residues, all of the ConA I fractions of alpha- and beta-chains bound to RCA-agarose. A high degree of similarity was observed between the corresponding glycopeptides of the three subsets of class II molecules and between the complex N-linked structures of alpha- and beta-chains. Minor variations were observed between DR1a and DR1b glycopeptides which appear greater than those observed between DR1 and DQw1 glycopeptides.(ABSTRACT TRUNCATED AT 400 WORDS)
The effect of fibronectin on the processing of C1q- and C3b/bi-coated immune complexes by peripheral blood monocytes.
Authors: Sorvillo J M , Gigli I , Pearlstein E
Published: December 2022

In the past several years, it has been demonstrated that plasma fibronectin (Fn) binds to the C1q subunit of the complement system. The effect of Fn on the processing of immune complexes containing C1q and C3b by human peripheral blood monocytes was investigated. Preincubation of monocytes with Fn causes a significant increase in attachment of sheep erythrocytes coated with IgM and C1q (EIg-MC1q), but does not mediate their ingestion. EIg-MC1q attach to the Fn-treated monocytes via the C1q receptor because Fab anti-Fn antibodies do not inhibit their attachment to the monocytes. In addition, Fn-treated monocytes exhibit no change in C1q receptor number or affinity compared with monocytes treated with buffer. Fn mediates the phagocytosis of C3b/bi-coated particles, and C1q can enhance this process in two ways. First, phagocytosis of particles bearing C3b/bi and Fn is enhanced by the presence of C1q on the immune complex. Second, monocytes on Fn-coated surfaces ingest more particles if they are coated with both C3b/bi and C1q, compared with particles coated with either C3b/bi or C1q alone.
The immunosuppressive activity of L-leucyl-L-leucine methyl ester: selective ablation of cytotoxic lymphocytes and monocytes.
Authors: Thiele D L , Lipsky P E
Published: December 2022

L-Leucyl-L-leucine methyl ester (Leu-Leu-OMe), a dipeptide condensation product of L-leucine methyl ester generated by human monocytes (M phi) or polymorphonuclear leukocytes, eliminates all natural killer cell (NK) function from mixed lymphocyte populations. In the present studies, the specificity of the action of Leu-Leu-OMe was examined. It was found that a variety of tissue culture cells and tumor lines of nonlymphoid origin were completely resistant to any demonstrable Leu-Leu-OMe-mediated toxicity. Furthermore, the erythroleukemia line K562, the T cell line Molt-4, the B cell lines HS-Sultan and Daudi, and EBV-transformed B cell lines were unaffected by concentrations of this compound that completely eliminated NK cells. Similarly, the vast majority of OKT4+ lymphocytes manifested no significant toxicity after Leu-Leu-OMe exposure. Furthermore, they retained the capacity to proliferate normally in response to allogeneic cells as well as the ability to provide help for the generation of immunoglobulin-secreting cells (ISC). However, Leu-Leu-OMe caused partial depletion of OKT8+ cells from mixed populations of lymphocytes. After such exposure, the remaining OKT8+ cells were still capable of proliferating in mixed lymphocyte cultures, but the suppressive effect of these cells on ISC generation was abolished. Furthermore, both precursors and activated effectors of cytotoxic T lymphocyte (CTL) and activated NK-like activity generated in mixed lymphocyte cultures were eliminated by exposure to low concentrations of Leu-Leu-OMe. Indeed, both OKT4+ and OKT8+ CTL were eliminated by Leu-Leu-OMe. In addition, both peripheral blood M phi and U937 cells, a human cell line with many M phi-like characteristics, were sensitive to Leu-Leu-OMe-mediated toxicity, although only at two- to fivefold higher concentrations than those completely eliminating NK cells. These findings indicate that Leu-Leu-OMe has selective toxicity for NK cells, CTL, and M phi without adverse effects on a variety of other lymphoid or nonlymphoid cell types.
Augmentation of respiratory mast cell secretion of histamine caused by vagus nerve stimulation during antigen challenge.
Authors: Leff A R , Stimler N P , Munoz N M , Shioya T , Tallet J , Dame C
Published: December 2022

We studied the effect of vagus nerve stimulation on the mast cell secretion of histamine after intraarterial (i.a.) administration of Ascaris suum antigen (AA) into the bronchial circulation of 10 randomly selected, natively allergic dogs in vivo. Respiratory mast cell response was measured as the arteriovenous difference (AVd) in histamine concentration across the bronchus. Plasma histamine concentration was determined simultaneously from right atrium, right ventricle, and femoral artery 60 and 15 sec before and 15, 30, 45, 60, 75, and 90 sec after i.a. injection of sham (Kreb-Henseleit) diluent, 1:100, and 1:30 concentrations of AA. The mean AVd in plasma histamine for five parasympathetically blocked animals (neural blockade with hexamethonium and beta-adrenergic blockade with propranolol) was 1.28 +/- 0.61 ng/ml (sham), 5.16 +/- 19.7 ng/ml (1:100 AA), and 36.6 +/- 11.1 ng/ml (1:30 AA). Substantial augmentation was obtained when AA was administered during parasympathetic stimulation in five other animals (beta-adrenergic blockade, no neural blockade), which was caused by continuous bilateral electrical stimulation of the vagus nerves. A mean AVd in plasma histamine of 110 +/- 27.6 ng/ml was obtained after 1:100 AA (p less than 0.001 vs parasympathetic blockade) and 166 +/- 32.4 ng/ml for 1:30 AA (p less than 0.001 vs parasympathetic blockade). Parasympathetic stimulation alone did not cause secretion of histamine. In contrast to the AVd response, parasympathetic stimulation did not augment nonrespiratory mast cell secretion after AA challenge. We conclude that vagus nerve stimulation augments secretion of histamine from respiratory mast cells during antigen challenge. We demonstrate that parasympathetic stimulation may potentiate the response to antigen challenge in central airways through augmented mast cell secretion of mediator.
Subcellular characterization of the endocytosis of small oligomers of mouse immunoglobulin G in murine macrophages.
Authors: Finbloom D S
Published: December 2022

To characterize the internalization and degradation of model immune complexes in murine macrophages, the endocytosis of well-defined radiolabeled IgG dimers and heavy oligomers (5 to 7 IgG molecules per complex), which were covalently cross-linked at the antigen-combining site, was studied. Of those heavy oligomers which were bound to the cell at 4 degrees C, 50 to 60% (400,000 molecules of IgG) were internalized within 30 min at 37 degrees C and, subsequently, were completely degraded over a period of 3 hr. Low pH had little effect on the dissociation of the oligomer from its receptor. The degradation of oligomers was markedly inhibited when macrophages were treated with monensin, a proton ionophore which raises organelle pH. Because this treatment did not prevent the delivery of oligomer into the lysosome, the transport of a soluble complex of IgG from the cell surface to the lysosome was not a pH-dependent event. On the other hand, 25 to 30% (50,000 molecules) of those dimers capable of binding to the cell entered the macrophage, but only 5000 molecules were degraded. When macrophages were studied by using density gradient centrifugation, within 15 min, heavy oligomers were found in a vesicle which sedimented at a density between that of the plasma membrane and lysosome. The density of this vesicle was similar to that of endosomes studied in other receptor-ligand systems. Heavy oligomers were within lysosomes shortly thereafter. Incubation of cells at 18 degrees C prevented the appearance of heavy oligomer within the lysosomes and resulted in the concentration of oligomers within an intracellular compartment of a density slightly heavier than that of plasma membrane. At 37 degrees C, dimers sedimented in a similar region of the gradient. But unlike heavy oligomers, dimers never entered lysosomes. These data suggest that the degree of Fc receptor clustering induced by oligomers of IgG influenced the intracellular fate of the ligand.
Differences between basophils and mast cells: failure to detect Charcot-Leyden crystal protein (lysophospholipase) and eosinophil granule major basic protein in human mast cells.
Authors: Leiferman K M , Gleich G J , Kephart G M , Haugen H S , Hisamatsu K , Proud D , Lichtenstein L M , Ackerman S J
Published: December 2022

Human eosinophils contain several distinctive proteins including eosinophil granule MBP and the membrane-associated CLC protein (lysophospholipase). Human basophils also contain these proteins, indicating biochemical similarities between eosinophils and basophils. To determine whether MBP or CLC protein is present in connective tissue mast cells, we studied human lung and cutaneous mast cells by immunofluorescence by utilizing specific antibodies to CLC and MBP. Cytocentrifuge slides of enriched lung mast cells and mast cells in sections of formalin-fixed, paraffin-embedded cutaneous tissue from urticaria pigmentosa lesions were stained for CLC and MBP. Neither pulmonary nor cutaneous mast cells stained for CLC protein or MBP. In contrast, lung and cutaneous eosinophils in the same preparations showed bright staining for both proteins. The failure to find CLC protein and MBP in mast cells provides additional evidence of dissimilarity between mast cells and basophils, and an immunochemical means to distinguish between them.
LPS regulation of specific protein synthesis in murine peritoneal macrophages.
Authors: Largen M T , Tannenbaum C S
Published: December 2022

Two-dimensional polyacrylamide gel electrophoresis (2D PAGE) analysis of biosynthetically labeled proteins of murine peritoneal macrophages elicited by inflammatory and activating stimuli indicated that the accumulation of a small number of cell-associated proteins was altered after in vitro treatment with bacterial lipopolysaccharide (LPS). Both increases and decreases in the accumulation of specific proteins were observed after LPS stimulation. Proteins of approximately 87, 43, 37, 30, and 28 Kd were similarly regulated by LPS in proteose peptone-, P. acnes-, and M. bovis BCG-elicited macrophages. Thioglycollate-elicited and resident peritoneal macrophages showed very few changes in the pattern of proteins synthesized after LPS treatment. Many of the proteins whose accumulation was increased by LPS in the elicited macrophages (proteins of approximately 87, 52, 43, 37, and 28 Kd) were already synthesized at high levels in resident macrophages. LPS stimulation also altered the accumulation of many of the same proteins in bone marrow-derived macrophages, indicating the lack of T lymphocyte influence on the LPS-induced changes in macrophages. LPS stimulation of highly purified B cells caused changes in the accumulation of several proteins of 70 and 78 Kd, which were different from those regulated by LPS in peritoneal macrophages.
Non-H-2-linked genetic regulation of cytotoxic responses to hapten-modified syngeneic cells. I. Non-H-2-linked Ir gene defect expressed on T cells is not predetermined at the stage of bone marrow cells.
Authors: Ogata M , Shimizu J , Tsuchida T , Takai Y , Fujiwara H , Hamaoka T
Published: December 2022

Spleen cells from C3H/He or BALB.K mice immunized to the newly synthesized amino-reactive hapten 5-sulfo-1-naphthoxy acetic acid N-hydroxysuccinimide ester (AED-NH2) were stimulated in vitro with AED-NH2-modified syngeneic cells. After 5 days of culture, effector cells were assayed for their cytotoxic activity against AED-NH2-modified target blast cells. C3H/He and BALB.K mice exhibited the respective high and low anti-AED-NH2 cytotoxic T lymphocyte (CTL) responses. This contrasted with the observation that both of these H-2k strains generated potent CTL responses against aminoreactive haptens, e.g., trinitrophenyl (TNP). Because C3H.SW and BALB.B strains, which are the H-2b counterpart of the above two strains, also represented the respective high and low responders to AED-NH2 hapten, this hapten model enabled us to investigate cellular mechanisms underlying the above non-H-2-associated genetic regulation of CTL responses (C3H vs BALB non-H-2 backgrounds). The results demonstrated that there was no detectable difference between C3H/He and BALB.K strains in the lysability of target cells and the ability of stimulating cells to activate primed spleen cells. Anti-AED-NH2 CTL responses were only marginal when antigen-presenting cells (APC) were eliminated from the primed spleen cells of high responder C3H/He or (C3H/He X BALB.K)F1 mice. The addition of APC to cultures free of APC regained an appreciable CTL response in C3H/He or (C3H/He X BALB.K)F1 mice, irrespective of whether APC were derived from high (C3H/He) or low (BALB.K) responders. We have also demonstrated that allogeneic radiation bone marrow chimera (BALB.K----C3H/He) exhibited a CTL response comparable to that induced by C3H/He mice, whereas the reverse direction of allogeneic chimera (C3H/He----BALB.K) induced a marginal CTL response. These results indicate that this non-H-2-associated Ir gene defect is expressed on T cells (CTL precursors and/or helper T cells) rather than APC, and that this T cell defect is not predetermined at the level of bone marrow cells. The results are discussed in the light of the genetic and cellular mechanisms underlying non-H-2-linked Ir gene control.
Evidence that Lyb-5 is a differentiation antigen in normal and xid mice.
Authors: Smith H R , Yaffe L J , Kastner D L , Steinberg A D
Published: December 2022

These studies addressed the question of whether the Lyb-5 determinant on mouse B cells might represent a marker for a unique lineage of B cells as has been thought, or rather, a differentiation antigen. Previous studies have addressed this question by treating splenic B cells with anti-Lyb-5 + complement and then culturing them with various antigens. In this study, we first immunized in vivo or in vitro and then determined susceptibility to anti-Lyb-5. These studies demonstrate that a substantial proportion of antibody-forming cells produced in response to immunization with sheep red blood cells, trinitrophenyl (TNP)-keyhole limpet hemocyanin, TNP-Brucella abortus, TNP-lipopolysaccharide, as well as TNP-Ficoll, are eliminated by anti-Lyb-5 + complement treatment at the end of culture. Some generation of Lyb-5+ antibody-forming cells occurred in the last 24 hr of culture. These studies suggest that the Lyb-5 marker represents a differentiation antigen on relatively mature B cells.
Tolerance defects in New Zealand Black and New Zealand Black X New Zealand White F1 mice.
Authors: McCoy K L , Kendrick L , Chused T M
Published: December 2022

The susceptibility of autoimmune NZB and (NZB X NZW)F1 mice to the induction of tolerance by monomeric BSA was compared with several normal mouse strains. Unresponsiveness in T and B lymphocyte compartments was probed by challenging with DNP8BSA and measuring anti-DNP and anti-BSA antibodies separately. Tolerance induced by monomeric BSA was carrier specific, and there was no evidence of epitope-specific suppression. Normal NZW, NFS, and B10.D2 mice were easily rendered tolerant with monomeric BSA and did not produce anti-DNP or anti-BSA antibodies after challenge with DNP8BSA. By contrast, the lack of anti-DNP antibody response in similarly treated NZB mice was dependent on the dose of monomeric BSA, indicating that the helper T cells were partially resistant to tolerance induction. NZB mice treated with a high dose of monomeric BSA produced anti-BSA, but not anti-DNP, antibodies after immunization. Thus, the anti-carrier B cells in NZB mice may have been primed by monomeric BSA. The presence of the xid gene on the NZB background rendered the mice susceptible to induction of tolerance, suggesting that the tolerance defect in NZB mice involves the B cell compartment. This abnormal antibody response was a dominant trait: (NZB X NFS)F1 and (NZB X B10.D2)F1 mice had the same characteristics as NZB mice. These F1 hybrids do not develop autoimmune disease, indicating that resistance to experimental tolerance induction expressed at a B cell level may not be sufficient for disease development. In contrast to NZB and other NZB F1 hybrids, (NZB X NZW)F1 hybrids treated with monomeric BSA and challenged with DNP8BSA responded to both DNP and BSA. The contribution of a B cell defect to the tolerance abnormality of (NZB X NZW)F1 mice was examined by analyzing the effect of the xid gene on the progeny of (NZB.xid X NZW)F1 mice. Unlike the effect of the xid gene on NZB mice, both phenotypically normal heterozygous female and phenotypically xid hemizygous male mice produced anti-DNP and anti-BSA antibodies after tolerance induction and immunization, demonstrating that a major helper T cell abnormality was present in (NZB X NZW)F1 mice. The (NZW X B10.D2)F1 hybrid was rendered tolerant by this procedure, indicating that the helper T cell defect (NZB X NZW)F1 mice may have resulted from gene complementation with the NZB mice contributing partial resistance of T helper cells to tolerance induction.(ABSTRACT TRUNCATED AT 400 WORDS)
Restoration of in vitro responsiveness of xid B cells to TNP-Ficoll by 8-mercaptoguanosine.
Authors: Ahmad A , Mond J J
Published: December 2022

8-Mercaptoguanosine (8sGuo) has been reported to enhance responses of normal mice to the type 2 antigen trinitrophenol (TNP)-Ficoll. In this report, we demonstrate that this immune adjuvant restores the immune responsiveness of B cells from mice with the x-linked immune defect (xid), which are nonresponsive to the type 2 antigen TNP-Ficoll. The data demonstrate that TNP-Ficoll, which by itself cannot stimulate anti-TNP responses in CBA/N mice, is able to initiate the initial steps of cell activation in xid B cells and render them sensitive to the subsequent differentiative effects of 8sGuo. We propose that the unresponsiveness of xid B cells to type 2 antigens reflects not the inability of these antigens to stimulate xid B cells from G0 to G1, but rather the inability of these antigen-activated cells to respond to a second signal to which these immune defective B cells are poorly responsive and can be substituted for by 8sGuo.
Expression of a T cell receptor-like molecule on normal and malignant murine T cells detected by rat monoclonal antibodies to nonclonotypic determinants.
Authors: Chan P Y , Takei F
Published: December 2022

A T cell receptor-like molecule with a dimer structure of 45 kilodaltons (Kd) under reducing and 90 Kd under nonreducing conditions was detected on the surface of two murine T lymphoma lines, EL-4 and MBL-2, by two rat monoclonal antibodies. The two antibodies seemed to react with different determinants on the same molecule. The antibodies did not react with the surface of normal T cells as tested by flow cytometric analysis of cell surface staining. Two-dimensional gel electrophoresis (IEF vs SDS-PAGE) and tryptic peptide analysis revealed the molecule to consist of two chains with different isoelectric points and different tryptic peptides. A conventional antiserum was raised against the heterodimer purified from EL-4 cells. The immune serum did not bind to the surface of normal T cells. However, the immune serum as well as the monoclonal antibodies immunoprecipitated the dimer molecules from detergent-solubilized normal thymocytes and spleen cells. The dimer molecule was detected on both immature and mature thymocytes. These results suggest that the antibodies detect non-clonotypic determinants on a T cell receptor-like protein. The determinants are masked on the surface of normal T cells, whereas they are exposed on the surface of at least two T lymphoma cell lines. Three polypeptides of 30 Kd, 25 Kd, and 15 Kd were also coprecipitated with the heterodimer from MBL-2 cells. These proteins may associate with the heterodimer and may be masking the antigenic determinants on normal T cells. The relationship between the heterodimer molecule described here and the T cell antigen receptor or the human T cell antigen 9.3 is still unknown.
Regulation of the synthesis of the third component of complement and factor B in cord blood monocytes by lipopolysaccharide.
Authors: Sutton M B , Strunk R C , Cole F S
Published: December 2022

We compared the regulation of C3 and factor B synthesis in cord blood and adult monocytes by using techniques for identification and quantification of newly synthesized proteins, lipopolysaccharide (LPS) from several Gram-negative organisms, and precursors of LPS. Synthesis of C3 and factor B in cord blood monocytes was unaffected by lipid A (the active moiety of LPS extracted by the Westphal procedure). In contrast, adult monocytes increased C3 synthesis by 11.5-fold and factor B synthesis by 3.1-fold in response to LPS. This difference in cord blood monocyte response to LPS was specific in that other LPS-induced monocyte functions (superoxide production and phagocytosis) were stimulated comparably in both cord blood and adult monocytes by LPS. To characterize further this regulatory difference, the roles of LPS precursors, arachidonic acid metabolites, and of factor(s) released by adult monocytes were examined. Precursors of the lipid portion of LPS (lipid X and lipid Y), LPS isolated by trichloroacetic acid extraction, and endotoxin-associated protein (EAP) increased C3 and factor B synthesis in cord blood monocytes. Inhibitors of the lipoxygenase pathway (dexamethasone, ETYA) but not of the cyclooxygenase pathway (indomethacin) abrogated the response of adult monocytes to lipid A and EAP and of cord blood monocytes to EAP. Finally, co-incubation of adult monocytes and cord blood monocytes in LPS-containing medium resulted in enhancement of C3 and factor B synthesis in cord blood monocytes. These data suggest that the difference in LPS response between cord blood and adult monocytes may result from differences in lipid processing or protein recognition of LPS, differences in the production of lipoxygenase pathway products, and/or one or more regulatory factors. The availability of human mononuclear phagocytes which exhibit distinct differences in biosynthetic responsiveness to LPS should permit investigation of the molecular mechanism(s) by which LPS affects C3 and factor B gene expression.
In vivo treatment of mice and hamsters with antibodies to asialo GM1 increases morbidity and mortality to pulmonary influenza infection.
Authors: Stein-Streilein J , Guffee J
Published: December 2022

The role of natural killer (NK) cells in host defenses against influenza virus infections in the lung was investigated by using rabbit antiserum to asialo GM1 (RAGM1), a neutral glycosphingolipid expressed on the plasma membrane of NK cells and some mouse pulmonary macrophages. Intravenous or intratracheal (i.t.) administration of RAGM1 resulted in depletion of the (in vitro) NK activity in lung and spleen or lung alone, respectively. The NK activity was depleted as early as 12 hr post-inoculation of antiserum, but returned to the normal range of activity by 4 days after antibody administration. RAGM1 serum treatment had no effect on the cytotoxic macrophage activity expressed by the plastic-adherent mononuclear cell populations isolated from mouse or hamster lung. Treatment of mice or hamsters with an i.t. or i.v. inoculation of RAGM1 rendered both species of laboratory animals susceptible to increased morbidity and mortality during a pulmonary influenza infection. These data support the hypothesis that a population of NK cells exist in an extravascular compartment within the lung, and that this local population of NK cells in the lung is crucial to the early natural pulmonary defenses during influenza infection.
Intraepithelial leukocytes contain a unique subpopulation of NK-like cytotoxic cells active in the defense of gut epithelium to enteric murine coronavirus.
Authors: Carman P S , Ernst P B , Rosenthal K L , Clark D A , Befus A D , Bienenstock J
Published: December 2022

Initially the intraepithelial leukocytes (IEL) of specific pathogen free (SPF) mice were compared with those of mice held without isolation and were found to differ markedly in total number and distribution of cell surface antigens. The IEL from SPF mice expressed significantly less Thy-1, Lyt-1, and Lyt-2 antigens than their conventional counterparts. The local cell-mediated immune response of mucosal lymphocytes to an enteric murine coronavirus (MHV-Y) was studied in inbred strains of naive SPF mice. A potent in vitro cytotoxic activity was demonstrated by mucosal leukocytes, especially IEL, and spleen cells for MHV-Y-infected syngeneic and allogeneic target cells. The cytotoxicity was not restricted by the major histocompatibility complex. Targets infected with Pichinde virus, an enveloped nonenterotropic virus, were not lysed by these cells. The phenotype of the IEL effector cell was asialo GM1+, Thy-1-, Lyt-1-, Lyt-2-. This cell represents a small subpopulation of the total IEL. After the in vivo administration of anti-asialo GM1 sera, the virus-specific cytotoxic function of the IEL was markedly diminished in in vitro assays, and there was enhanced persistence of virus in gut tissues in vivo. The IEL effector population is defined as a natural killer-like cell that appears to be active in the defense of the gut epithelium to a murine enteric coronavirus.
Lectin-dependent and anti-CD3 induced cytotoxicity are preferentially mediated by peripheral blood cytotoxic T lymphocytes expressing Leu-7 antigen.
Authors: Phillips J H , Lanier L L
Published: December 2022

Monoclonal antibodies against the CD3 antigen and certain lectins can induce interleukin 2 dependent antigen-specific T cell clones to mediate non-antigen specific cytotoxicity. On the basis of this observation, we predicted that it may be possible to identify cytotoxic T lymphocytes (CTL) in peripheral blood without knowing the antigen specificity of these in vivo primed CTL. By using this strategy, peripheral blood lymphocytes were separated into low and high-density fractions on Percoll gradients and were tested for cytotoxic activity in the presence or absence of concanavalin A (Con A) or anti-Leu-4 antibody. Lectin-dependent cellular cytotoxicity (LDCC) and anti-CD3 induced cytotoxicity against both natural killer (NK)-insensitive and NK-sensitive targets were exclusively mediated by low-density CD3+ T lymphocytes. Additional studies indicated that low-density CD3+ T lymphocytes co-expressing Leu-7 antigen preferentially mediated this activity, although in some individuals, significant activity was also observed in the low-density T cells lacking Leu-7. In contrast, high-density CD3+ T lymphocytes and CD16+ (Leu-11+) NK cells (both Leu-7 and Leu-7+) did not mediate nonantigen-specific cytotoxicity under these conditions. The finding that NK cell-mediated cytotoxicity was unaffected by these lectins refutes the hypothesis that lectin-dependent cellular cytotoxicity is simply a result of effector and target agglutination. T cell-mediated cytotoxicity was both lectin and antibody specific. Phytohemagglutinin, Con A, and pokeweed mitogen induced cytolytic activity in the Leu-7+ T cells, whereas wheat germ agglutinin did not. Of the antibodies against T cell-associated differentiation antigens (anti-Leu-2,3,4, and 5), only anti-Leu-4 induced cytotoxicity. This anti-CD3-induced cytotoxicity was essentially completely inhibited by the presence of anti-LFA-1 or anti-CD2 monoclonal antibodies, implicating these molecules in the triggering process. A proportion of the CD3+, Leu-7+ CTL expressed HLA-DR antigens, indicating possible in vivo activation. Because previous clinical studies have indicated that lymphocytes with this phenotype may be elevated in clinical situations associated with immunosuppression and chronic viral infection, this unique subset of CD3+ T lymphocytes may represent a population of in vivo primed CTL possibly against viral antigens.
Induction of Fc epsilon receptors on mouse macrophages and lymphocytes by homologous IgE.
Authors: Daeron M , Ishizaka K
Published: December 2022

Normal mouse peritoneal macrophages express Fc gamma 2aR, Fc gamma 1/2bR, and Fc epsilon R, whereas B lymphocytes in mesenteric lymph nodes (MLN) bear Fc gamma 2bR, Fc gamma 1R, and Fc epsilon R. Rosette formation of Fc epsilon R+ macrophages and lymphocytes with IgE-coated ox erythrocytes was inhibited by rodent IgE but not by any other isotype of mouse immunoglobulins. In contrast, IgG1 rosettes and IgG2b rosettes of both macrophages and lymphocytes were inhibited not only by homologous isotype but also by IgE, suggesting that IgE has some affinity for Fc gamma 1/2bR on macrophages and for both Fc gamma 1R and Fc gamma 2bR on lymphocytes. Incubation of normal mouse macrophages with mouse IgE for 24 hr resulted in a twofold increase in the proportion of Fc epsilon R+ cells. Mouse IgE can induce Fc epsilon R on B cells as well. Incubation of MLN cells with mouse IgE for 2 to 4 hr, followed by culture of the cells in the absence of IgE, resulted in a 1.8- to 2.9-fold increase in Fc epsilon R+ cells. Determination of Fc gamma R+ cells in the same MLN cells revealed that induction of Fc epsilon R by IgE was accompanied by a substantial decrease in the expression of Fc gamma 1R and Fc gamma 2bR. Induction of Fc epsilon R by IgE on macrophages and lymphocytes requires protein synthesis. In MLN cells, cycloheximide inhibited not only the IgE-induced increase in Fc epsilon R+ cells but also the decrease in Fc gamma 1R+ cells and Fc gamma 2bR+ cells. It was also found that induction of Fc epsilon R by IgE on macrophages was completely inhibited if IgG1 or IgG2b was added to the cells together with IgE. In contrast, IgG2a did not affect the IgE-induced expression of Fc epsilon R on macrophages. In MLN cells, IgG2b but not IgG1 inhibited both IgE-induced increase in Fc epsilon R and decrease in Fc gamma 1R and Fc gamma 2bR. The results indicate that expression of various Fc receptors on lymphocytes is mutually regulated.
Human monocytes and U937 cells bear two distinct Fc receptors for IgG.
Authors: Looney R J , Abraham G N , Anderson C L
Published: December 2022

Several convergent lines of evidence have led us to propose that human monocytes and the related cell line U937 possess a second class of IgG Fc receptor (FcR) in addition to the 72-Kd high affinity FcR previously described. IgG affinity purification from detergent lysates of surface radiolabeled U937 cells has yielded both a 40-Kd IgG-binding membrane protein (p40) and the 72-Kd FcR protein. By the same procedure, only the p40 was isolated from the erythroblast cell line K562 and from the B cell lines, Daudi and Raji. Serologic cross-reactivity between the 40-Kd FcR on U937 and Daudi cells was demonstrated using a goat anti-FcR antiserum. A murine (m) monoclonal antibody, raised against the FcR of K562 cells, precipitated the 40-Kd FcR from lysates of U937 and K562 cells but not from Daudi or Raji cells. This antibody, referred to as anti-p40 (IV.3), selectively inhibited the binding of murine IgG1-coated erythrocytes to U937 cells, whereas monomeric human IgG selectively inhibited binding of human anti-Rh(D)-coated erythrocytes to U937 cells. Both Daudi and U937 cells mediated mIgG1 anti-T3 (Leu-4)-induced stimulation of T lymphocytes. In contrast, mIgG2a anti-T3 (OKT3)-induced stimulation was supported effectively by U937 cells but only modestly by Daudi cells. Intact IgG or Fab fragments of anti-p40 (IV.3) blocked mIgG1 anti-T3 (Leu-4) stimulation but not mIgG2a anti-T3 (OKT3) stimulation of T cells; monomeric human IgG blocked only OKT3-induced stimulation. The simplest interpretation of these results is that human monocytes and U937 cells bear two classes of IgG FcR, one of 72 Kd and the other, as described above, of 40 Kd. We propose that the 72-Kd FcR mediates rosette formation with red cells coated by human anti-Rh IgG as well as T cell stimulation by mIgG2a anti-T3 (OKT3) and that the 40-Kd FcR mediates rosette formation with erythrocytes bearing mIgG1 as well as T cell stimulation by mIgG1 anti-T3 (Leu-4). Furthermore, we suggest that these two FcR are the human homologues of the murine macrophage FcRI (binding mIgG2a) and FcRII (binding mIgG2b/1).