bluuebunny/crossref_metadata_2025 huggingface dataset:
Low molecular weight B cell growth factor-responsive cloned human B cell lines. I. Phenotypic differences and lack of requirement for CD23 (Fc epsilon RII).
Authors: Warrington R J , Rutherford W J , Wong S K , Cook J M , Rector E S
Published: December 2022

The EBV-transformed B cell line JR-2 proliferates in response to partially purified preparations of low m.w. B cell growth factor (LMW-BCGF). Two clones of JR-2 were generated that retained this LMW-BCGF responsiveness, exhibiting similar dose/response characteristics but differing phenotypically. The B10 clone grows as single, discrete, small round cells, whereas D3 grows in aggregates. The clones also differ in the expression of cell surface Ag, D3 being weakly DR+ and strongly CD23+, whereas B10 lacks these Ag. The CD23 on D3 cells binds IgE. Both clones are T9+, 4F2+, B1-, B2- and CALLA-. D3 expresses surface IgG and differentiates in the presence of LMW-BCGF, to secrete IgG. B10 lacks surface and cytoplasmic Ig and fails to differentiate in response to LMW-BCGF. CD23 cannot be induced on B10 by incubation with either LMW-BCGF or IL-4. B10 does not shed CD23 and shed CD23 is not a growth factor for either cloned line. Expression of CD23 on D3 cells is not affected by preincubation with LMW-BCGF. Neither B10 or D3 cells respond to rIL-1, rIL-2, rIL-4, rIL-6, rTNF-alpha/beta, rIFN-gamma, or to high m.w. BCGF (Namalwa), alone or in combination. Both clones absorb BCGF activity and crossover absorptions indicate that the clones remove growth factors required by each other. D3 and B10 both appear therefore to respond selectively to LMW-BCGF. These data suggest that the loss of CD23 from a cloned derivative of the cell line JR-2, although accompanied by considerable phenotypic change, is not associated with the disappearance of LMW-BCGF responsiveness, indicating that CD23 is not the essential receptor for LMW-BCGF.
C-reactive protein reacts with the U1 small nuclear ribonucleoprotein.
Authors: Du Clos T W
Published: December 2022

C-reactive protein (CRP) was found to produce a small, discrete, speckled fluorescence pattern in the nucleus of HEp-2 cells. Double staining with anti-RNP serum and CRP produced very similar staining patterns. By counterimmunoelectrophoresis CRP was bound to extractable nuclear antigens found in rabbit thymus extract. The reactive components of the extract were only partially sensitive to treatment with RNase. CRP immunoprecipitated the U1 RNA species from [32P]labeled HeLa cells and the protein bands of the Sm/RNP complex from [35S]-methionine-labeled HeLa cells. By blotting, CRP bound to several discrete bands in a calcium-dependent, PC-inhibitable manner. Two of the bands comigrated with the 70K protein band associated with the U1 snRNP, and its major breakdown product. Binding to these bands was inhibited by both EDTA and PC indicating that CRP binds these proteins through the PC-binding site. Binding to the 70K protein of the U1 snRNP was confirmed by reactivity with the recombinant 70K protein in a dot blot. These findings indicate the CRP binds to the U1-RNP snRNP particle. Considering the ability of CRP to inhibit antibody responses to its ligands and its ability to activate C and promote phagocytosis it is suggested that CRP may play a role in the regulation of autoantibody responses to nuclear Ag.
Bactericidal defect of neutrophils in a guinea pig model of thermal injury is related to elevation of intracellular cyclic-3',5'-adenosine monophosphate.
Authors: Bjornson A B , Knippenberg R W , Bjornson H S
Published: December 2022

PG of the E series inhibit major effector functions of polymorphonuclear leukocytes (PMN) by elevating intracellular cAMP. The present study investigated the involvement of this mechanism in the bactericidal defect of PMN induced by thermal injury in a guinea pig model. Peripheral and peritoneal exudate PMN harvested from thermally injured guinea pigs at 1 or 2 days postburn had decreased bactericidal activity against Pseudomonas aeruginosa and a marked increase in cAMP content. Production of PGE1 by these cells in the absence of exogenous PMN activators was also increased. Treatment of PMN in vitro or in vivo with nonsteroidal anti-inflammatory drugs (indomethacin, ibuprofen, and piroxicam) restored bactericidal activity to normal and concomitantly reduced cAMP content and PGE1 production. A concomitant reduction in cAMP content and PGE1 production was also observed as bactericidal activity of PMN returned to normal under natural conditions during 4 to 7 days postburn. The enhancement of PMN bactericidal activity mediated by NSAID was fully counteracted by purified PGE1, theophylline, and by cAMP itself. These results suggest that the bactericidal defect of PMN induced by thermal injury is related to elevation of cAMP and that PGE1 plays a significant role in this phenomenon.
The rise in concentration of free Ca2+ and of pH provides sequential, synergistic signals for secretion in antigen-stimulated rat basophilic leukemia (RBL-2H3) cells.
Authors: Ali H , Collado-Escobar D M , Beaven M A
Published: December 2022

Ag stimulation of rat basophilic leukemia (RBL-2H3) cells results in hydrolysis of inositol phospholipids, a transient increase in concentration of cytosol Ca2+ [( Ca2+]i), a gradual increase in cytosolic pH (pHi) and the activation of protein kinase C. To determine whether all these changes serve as signals for secretion, studies were conducted with cells permeabilized with streptolysin O in which pHi and [Ca2+]i could be varied independently of each other and enzyme activities could be manipulated. At resting pHi (approximately 7.0) and [Ca2+]i (0.1 microM), the permeabilized cells showed little secretory response to Ag. At resting pHi, elevated levels of Ca2+ (0.33 microM) were required for maximal secretory response to Ag. At a pHi of 7.4, however, 0.1 microM [Ca2+]i was sufficient to sustain near maximal responses to Ag. Therefore, a small increase of [Ca2+]i to 0.33 microM was required to initiate secretion, but once the pHi was elevated secretion could be sustained at near basal levels of [Ca2+]i. Since elevating the [Ca2+]i and pHi, by themselves promoted little secretion, another potentiating signal must have been generated by antigen stimulation. This signal was possibly transduced via hydrolysis of inositol phospholipids and protein kinase C. Even with an elevated [Ca2+]i (0.33 microM) the hydrolysis of the phospholipids and secretion stimulated by Ag were inhibited by guanosine 5'(2-O-thio)diphosphate and neomycin. Furthermore, both protein-kinase C and the secretory response to Ag were lost after permeabilized cells were washed but both were retained if cells were exposed to PMA before permeabilization.
Characterization of T cell antigens associated with the cell wall protein-peptidoglycan complex of Mycobacterium tuberculosis.
Authors: Barnes P F , Mehra V , Hirschfield G R , Fong S J , Abou-Zeid C , Rook G A , Hunter S W , Brennan P J , Modlin R L
Published: December 2022

Mycobacterium tuberculosis cell walls are likely to contain critical T cell Ag capable of inducing protective immunity against the development of tuberculosis in animal models. Therefore, we characterized cell wall-associated Ag that stimulate T lymphocytes in tuberculosis patients and clinically well tuberculin-positive individuals. A protein-peptidoglycan complex isolated from the M. tuberculosis cell wall had potent immunologic activity, evoking PBMC proliferative responses similar to those induced by sonicated whole M. tuberculosis. In order to characterize the immunoreactive protein determinants associated with the protein-peptidoglycan complex, T cell lines were established to cell wall Ag and used to probe M. tuberculosis proteins separated by SDS-PAGE. These T cell lines proliferated primarily to protein Ag of 10, 19, 23, 28, 30, 40 to 50, and 65 kDa. Cell wall-reactive T cell clones that recognized the 10-, 23-, 28-, and 30-kDa proteins as single bands on SDS-PAGE did so under reducing and nonreducing conditions, suggesting that these are not proteolytic fragments or subunits of larger protein aggregates. We propose that these protein monomers, when post-translationally complexed with peptidoglycan, are the key ingredients of the immunogenic protein-peptidoglycan complex. In order to assess the relationship of the cell wall-associated Ag to those secreted proteins from "early culture filtrates" of actively growing M. tuberculosis recently implicated in eliciting protective immunity, cell wall-reactive T cell clones were tested for their ability to recognize early culture filtrates. Results revealed that at least three proteins shared with the cell wall complex are contained within early culture filtrates. Our data indicate that antigenic determinants associated with the protein-peptidoglycan complex of the M. tuberculosis cell wall may be involved in protective immunity and hence are potential candidates for inclusion in an effective antituberculosis vaccine.
Assessment of human natural killer and lymphokine-activated killer cell cytotoxicity against Toxoplasma gondii trophozoites and brain cysts.
Authors: Dannemann B R , Morris V A , Araujo F G , Remington J S
Published: December 2022

Because previous work has suggested that NK cells may be important in host resistance against the intracellular parasite Toxoplasma gondii we examined whether human NK cells and lymphokine-activated killer (LAK) cells have activity against trophozoites and cysts of this organism in vitro. A method to radiolabel Toxoplasma trophozoites with 51Cr was developed and direct cytotoxic activity was determined by using modifications of the standard 51Cr release assay. Viability of 51Cr-labeled trophozoites assessed by both methylene blue staining and trypan blue exclusion was greater than 90%. Significantly more 51Cr was released by anti-Toxoplasma antibody and C than by antibody in the absence of C. Incubation of trophozoites with freshly isolated human NK cells or NK cells activated with either rIL-2 or rIFN-alpha did not result in significant release of 51Cr (specific lysis was 0 to 2.3%). In contrast, the average specific lysis of radiolabeled trophozoites by LAK cells was significant (specific lysis was 7.8% +/- 1.1, p less than 0.01). In a series of separate experiments, preincubation of radiolabeled trophozoites with heat-inactivated normal or Toxoplasma antibody-positive human serum increased the cytotoxicity of LAK cells from a mean specific lysis of 15% +/- 4.5 to 39% +/- 8.5, respectively (p less than 0.05), as assessed by 51Cr release. Because previous work has shown that radioisotope release from parasites may be nonspecific, separate experiments were performed to determine the cytotoxicity of LAK cells against antibody-coated trophozoites by using ethidium bromide-acridine orange staining to assess effector cell damage. LAK cells had a mean specific lysis of 51% against antibody-coated trophozoites by ethidium bromide-acridine orange staining. Preincubation with heat-inactivated Toxoplasma-antibody positive human serum did not increase activity of rIL-2-activated NK cells against 51CR-labeled trophozoites. Neither human NK cells (freshly isolated or activated by rIL-2 or rIFN-alpha) nor LAK cells were cytotoxic for purified preparations of cysts of Toxoplasma isolated from the brains of chronically infected mice.
Analysis of an immunodominant region of infectious bronchitis virus.
Authors: Kusters J G , Jager E J , Lenstra J A , Koch G , Posthumus W P , Meloen R H , van der Zeijst B A
Published: December 2022

We analyzed the antigenic fine-structure of an immunodominant region in the peplomer protein of infectious bronchitis virus. This region near the N-terminus of the S2 subunit is recognized by polyclonal antisera and by the majority of mAb that cross-react with denatured protein. Despite their involvement in neutralization, epitopes in this region were conserved in different serotypes. Epitopes of four mAb and two chicken antisera were localized by using prokaryotic expression of cDNA fragments, and overlapping peptides with lengths increasing from 3 to 12 residues (PEPSCAN). We found overlapping epitopes with lengths of 6, 9, 11, and more than 17 residues. The results indicate that the expression products are antigenically equivalent to denatured protein fragments. This suggests a general strategy for the localization of sequential epitopes in large proteins. We propose that the immunodominance of the N-terminal region of S2 is explained by features of the protein structure. Presumably, this region is a protruding protein segment of about 20 residues with a high local mobility, as indicated by the antigenicity of the peptides. The conservation of the sequence points to an involvement in a molecular recognition process during infection.
B7, a new member of the Ig superfamily with unique expression on activated and neoplastic B cells.
Authors: Freeman G J , Freedman A S , Segil J M , Lee G , Whitman J F , Nadler L M
Published: December 2022

The human B cell restricted activation antigen B7 identifies an in vivo primed subpopulation of B cells that demonstrate an accelerated response to triggers of B cell activation and proliferation. The cDNA encoding B7 was molecularly cloned by cDNA expression. The identity of the B7 cDNA was confirmed by indirect immunofluorescence and immunoprecipitation of a 44/54-kDa protein from B7 transfected COS cells. The sequence of the B7 polypeptide predicts a type I membrane protein of 262 amino acids with eight potential N-linked glycosylation sites in the extracellular region and a short, highly positively charged cytoplasmic tail. The extracellular region is homologous to the Ig gene superfamily and consists of two contiguous Ig-like domains. The first Ig domain has the characteristics of a variable domain and the second that of a constant region domain. B7 mRNA was detected only on anti-Ig activated B lymphocytes and not in other hematopoietic cells. After in vitro activation of B cells with anti-Ig, B7 mRNA was maximally expressed between 4 and 12 h with four RNA transcripts of 1.7, 2.9, 4.2, and 10 kb. The 2.9-kb mRNA predominated in in vitro-activated B cells whereas the 1.7-kb mRNA was most abundant in tumor cells of B cell origin. B7 expression was confined to several histologically defined subgroups of B cell malignancies. The majority of non-Hodgkin's lymphomas were B7+ whereas the B cell leukemias and circulating non-Hodgkin's lymphomas were generally negative. These results demonstrate that the B7 gene encodes a unique molecule belonging to the Ig superfamily and that B7 expression is limited to normal activated B cells and noncirculating B cell malignancies.
A distinct subset of human CD4+ cells with a limited alloreactive T cell receptor repertoire.
Authors: Morishita Y , Sao H , Hansen J A , Martin P J
Published: December 2022

We describe a subset of CD4+/CD3+ human T lymphocytes that demonstrated a remarkably limited TCR repertoire responding to alloantigen stimulation. These cells have been characterized previously by their granular morphology and expression of CD11b but not CD28. Whereas multiple CD28+/CD4+ alloproliferative cloned cell lines generated by culture at limiting dilution immediately after isolation from peripheral blood each had a unique TCR-beta gene rearrangement, 19 of 21 CD11b+/CD4+ clones showed identical TCR-beta, and gamma gene rearrangements. In conventional MLR, the CD11b+/CD4+ cells responded poorly after stimulation with some HLA-class II Ag, and staining with a TCR Id-specific antibody and DNA blot hybridization suggested that the responding CD11b+/CD4+ cells typically contained predominant clonal populations. Clones of CD11b+/CD4+ cells with different TCR gene rearrangements showed closely similar patterns of responses when stimulated by a panel of allogeneic PBMC, but the response pattern did not correspond to that of any known HLA-class II Ag. These findings indicate that CD11b+/CD4+ cells have a limited alloproliferative repertoire characterized by predominant recognition of a limited number of undefined determinants that appear to be expressed in association with multiple distinct HLA-class II Ag. Our results suggest that CD11b+/CD4+ cells are selected for clonal reactivity by processes distinct from those for CD28+/CD4+ cells.
Cytotoxic T lymphocyte unresponsiveness induced by prolonged treatment with immobilized anti-CD3 antibody. Association of impairment of cytolytic activity with temporary depletion of intracellular protein kinase C.
Authors: Sano S , Kiyotaki C , Tatsumi Y , Fujiwara H , Hamaoka T
Published: December 2022

In our study we investigated the effect of pretreatment of bulk CTL and CTL clones with immobilized anti-CD3 antibody (Ab) or PMA. Primary CTL and CTL clones were cultured in dishes coated with anti-CD3 Ab or in medium containing PMA (5 nM) and assayed for Ag-specific or Ag-nonspecific "redirected" cytolysis using FcR+ P815 cells as targets. Cytotoxic activity of bulk CTL and five of six CTL clones tested in this study were inhibited by prolonged (longer than 6 h) pretreatment with immobilized anti-CD3 Ab or PMA, whereas proliferation of CTL clones or expression of surface CD3 molecules were not. The intracellular granule enzyme (N-alpha-benzyloxycarbonyl-L-lysine thiobenzyl ester esterase) activity of CTL clones was not reduced under these suppressive conditions, indicating that the incompetence of CTL is not merely due to depletion of cytolytic granules by chronic stimulation. The suppressed cytotoxicity could be recovered by culturing CTL without perturbation of CD3 molecules for 24 h. In one exceptional clone, BM10-37, pretreatment with immobilized anti-CD3 Ab or PMA did not suppress the cytotoxic activity. Immunostaining of intracellular protein kinase C (PKC) revealed that PKC was depleted after prolonged treatment with immobilized anti-CD3 Ab or PMA in those susceptible CTL clones but not in the resistant BM10-37. These findings lead us to conclude that prolonged stimulation of CD3 of CTL results in depletion of PKC and that PKC may be essential for signal transduction to deliver a lethal hit to the target cells.
T lymphocytes from healthy individuals with specificity to self-epitopes shared by the mycobacterial and human 65-kilodalton heat shock protein.
Authors: Munk M E , Schoel B , Modrow S , Karr R W , Young R A , Kaufmann S H
Published: December 2022

The immune response to mycobacterial pathogens comprises a significant percentage of T cells with specificity for a 65-kDa heat shock protein (hsp) which is highly conserved in bacteria and man. PBMC were activated in vitro with killed Mycobacterium tuberculosis and afterward tested for CTL activity on autologous target cells primed with 1) killed M. tuberculosis, 2) intact recombinant 65-kDa hsp of Mycobacterium bovis/M. tuberculosis; or 3) tryptic fragments of the recombinant 65-kDa hsp. Strong CTL activity was observed on targets primed with killed M. tuberculosis or with tryptic fragments of the 65-kDa hsp, but not on those primed with the intact 65-kDa hsp. M. tuberculosis activated T cells from 2/13 donors tested exerted killer activity against unprimed targets. To assess whether T cell responses were directed against self-epitopes shared by the mycobacterial and human 65-kDa hsp, four peptides of at least 10 amino acids length were synthesized corresponding to fully or almost identical regions of these molecules. Peripheral blood T cells from 8/9 individuals tested, after activation with killed M. tuberculosis, expressed strong CTL activity toward autologous targets primed with one or more of these synthetic peptides. By using HLA-DR transfected murine L cells we found that the epitopes were recognized in the context of histocompatible HLA-DR (class II) molecules. We conclude that the demonstration of T cells with specificity to self-epitopes in vitro is not indicative for autoimmune disease. However, if at certain stages of infection such T cells are activated by crossreactive microbial epitopes they could cause autoimmune responses.
Characterization of IL-6 receptor expression by monoclonal and polyclonal antibodies.
Authors: Hirata Y , Taga T , Hibi M , Nakano N , Hirano T , Kishimoto T
Published: December 2022

mAb and polyclonal antibodies against human IL-6R were prepared by using a murine transfectant cell line expressing the human IL-6R and a synthetic oligopeptide made on the basis of the deduced amino acid sequence as immunogens. Immunoprecipitation of radiolabeled IL-6R with these antibodies showed that the Mr of a mature IL-6R was 80 kDa and its value was reduced to 50K after treatment with O- and N-glycanase and neuraminidase, indicating that IL-6R is a glycoprotein. Two mAb recognizing different epitopes were prepared. One, PM1 inhibited the binding of 125I-IL-6 to the receptor and blocked the IL-6-dependent growth of a T lymphoma line, KT3. PM1 could not bind to IL-6R when it was saturated with IL-6, indicating that this antibody recognizes the IL-6 binding or the adjacent site on IL-6R. The other, MT18 was not inhibited by IL-6 for its recognition of IL-6R, therefore, this could be used for cytofluorometric staining of normal cells. Nonstimulated B cells expressed undetectable amount of IL-6R regardless of the expression of surface IgD. However, after the stimulation with PWM, IL-6R was observed on IgD- B cells with a relatively large size, but subtly on IgD- small B cells and not on IgD+ B cells, fitting the function of IL-6 which acts on activated B cells to induce Ig production. In contrast, IL-6R was detected on non-stimulated CD4+/CD8- and CD4-/CD8+ T cells. The level of IL-6R on both T cell subpopulations was not significantly changed after stimulation with phytohemagglutinin.
A novel polypeptide secreted by activated human T lymphocytes.
Authors: Miller M D , Hata S , De Waal Malefyt R , Krangel M S
Published: December 2022

We have identified two cDNA clones, I-309 and G-26, which define genes expressed abundantly in activated human PBMC, but at low or undetectable levels in resting PBMC. Based upon nucleotide sequence analysis, both clones are predicted to encode small, structurally related polypeptides, each containing a hydrophobic leader sequence characteristic of secreted proteins and a motif of four conserved cysteine residues. Further, I-309 and G-26 are structurally related to a growing family of genes that apparently encode small polypeptides whose secretion is induced upon cell activation. I-309 represents a previously undescribed human gene. We have generated an anti-peptide antiserum to the I-309 gene product which recognizes proteins in culture supernatants of an activated T cell clone and of COS cells transfected with the I-309 cDNA, supporting the idea that I-309 encodes a secreted protein. Because I-309 encodes a small protein secreted by activated T cells that displays structural features similar to other cytokines, we believe that it defines a novel cytokine with as yet unknown function.
The physical separation of Lps and Ifa loci in BXH recombinant inbred mice.
Authors: Fultz M J , Vogel S N
Published: December 2022

Several reports in the literature suggest that many of the phenotypic defects of LPS-hyporesponsive C3H/HeJ mice may be attributed to decreased IFN production by their macrophages. The physical proximity on chromosome 4 of the gene which encodes sensitivity to LPS (Lps) and the structural gene cluster which encodes IFN-alpha (Ifa), suggests the possibility that the Lps gene, whose product is unknown, may actually be a part of the Ifa gene cluster. The C57BL/6J and C3H/HeJ mouse strains carry distinct alleles at both the LPs and the Ifa loci. In this study, we have analyzed these parental strains, as well as 12 recombinant inbred strains derived from these parental strains (e.g., BXH strains), for inheritance of these distinct alleles. The results show the segregation of these two loci: in 5 of 12 BXH RI strains, the IFN-alpha restriction fragment length polymorphism characteristic of one parental strain was discordant with the predicted LPS response phenotype. Therefore, we conclude that the Lps and the Ifa genes are physically distinct despite the apparent cause and effect relationship which is observed phenotypically.
IL-4 inhibits H2O2 production and antileishmanial capacity of human cultured monocytes mediated by IFN-gamma.
Authors: Lehn M , Weiser W Y , Engelhorn S , Gillis S , Remold H G
Published: December 2022

The effect of IL-4 on the IFN-gamma-induced state of activation of cultured human monocytes was investigated with regard to their ability to produce hydrogen peroxide and their antileishmanial capacity towards the intracellular parasite Leishmania donovani. IL-4 was found to inhibit the IFN-gamma-dependent hydrogen peroxide production of monocytes. Treatment of monocytes with IFN-gamma (200 to 600 U/ml) for 48 h increased the hydrogen peroxide production fourfold above background. Coincubation of the monocytes with IL-4 (1 to 1000 U/ml) and IFN-gamma (200 to 600 U/ml) inhibited this increase by 50 to 100%. IL-4 alone did not modulate the hydrogen peroxide production of monocytes. Pretreatment of monocytes with IL-4 for 20 min to 3 h was already effective in preventing the IFN-gamma response. Addition of IL-4 not later than 6 h after the start of incubation with IFN-gamma was necessary for an optimal inhibitory effect. IL-4 also inhibited the IFN-gamma-induced antileishmanial capacity of monocytes: IFN-gamma (1000 U/ml) induced a 54 +/- 10% reduction in the number of parasites. Monocytes treated with combinations of IL-4 (100 to 1000 U/ml) and IFN-gamma (1000 U/ml) were unable to reduce the parasite numbers. IL-4 alone did not alter the uptake of Leishmania donovani nor induce antileishmanial activity. These results demonstrate that IL-4 disables human cultured monocytes to respond to IFN-gamma activation.
Characterization of a polysaccharide antigen from Bacteroides gingivalis.
Authors: Schifferle R E , Reddy M S , Zambon J J , Genco R J , Levine M J
Published: December 2022

A polysaccharide Ag (PS) was isolated from the phenol-water extract of Bacteroides gingivalis strain A7A1-28 and separated from LPS by Sephacryl S-400 HR chromatography. The PS was composed of glucose, glucosamine, galactosamine, and galactosaminuronic acid, while the LPS contained rhamnose, mannose, galactose, glucose, glucosamine, galactosamine, phosphate, and lipid, but not galactosaminuronic acid. The PS and LPS were immunochemically distinct by immunoelectrophoresis in agarose with homologous rabbit antiserum. The phenol-water extract from strain A7A1-28 was immunoreactive by immunoelectrophoresis against antisera to three additional strains of B. gingivalis, however, the PS was only reactive with homologous serum. Immunochemical characterization of decarboxylated and deacetylated PS derivatives suggest that the acetylation of the amino sugars, but not the presence of the carboxylate residue on galactosaminuronic acid contributes to major immunodeterminant expression.
Regulation of the IL-2 receptor alpha-gene. Interaction of a kappa B binding protein with cell-specific transcription factors.
Authors: Freimuth W W , Depper J M , Nabel G J
Published: December 2022

The IL-2R alpha enhancer contains an 11 bp sequence that resembles kappa B, a regulatory element associated with several genes, including Ig kappa-L chain and human immunodeficiency virus. Although nuclear factor of the kappa-enhancer in B cells (NF-kappa B) binding is activated by PMA, TNF-alpha, and IL-1, activation of the IL-2R alpha enhancer does not consistently correlate with NF-kappa B induction. In this report, we show that TNF-alpha activates NF-kappa B and the human immunodeficiency virus enhancer in the Jurkat T leukemia but does not stimulate the IL-2R alpha enhancer. In contrast, this cytokine, and IL-1, increased IL-2R alpha gene expression in YT-1 cells. Comparing YT-1 and Jurkat T leukemias, we find that the IL-2R kappa B site is required for TNF-alpha and IL-1 stimulation in YT-1 cells, but that plasmids containing this site linked to a heterologous promoter do not respond to these cytokines. These data suggest that upstream regulatory elements in addition to IL-2R kappa B are needed to mediate this cytokine effect. TNF-alpha also synergized with PMA and other cytokines in the stimulation of the IL-2R alpha enhancer in YT-1. Because these effects are not observed in Jurkat cells, the function of the IL-2R kappa B site is cell-specific and likely mediated by different associated transcription factors present in each cell type.
Identification of a novel 110-kilodalton structure expressed on a subset of T cell receptor-gamma delta-bearing cloned lymphocytes.
Authors: David V , Bachelez H , Leca G , Degos L , Boumsell L , Bensussan A
Published: December 2022

A small percentage of circulating CD3+ cells express a heterodimeric gamma delta receptor. Most of these cells do not express the surface marker CD4 and only a fraction of them bear CD8 molecules. The specificity and function of TCR-gamma delta are unclear. We obtained a murine mAb produced against an IL-2-dependent human T cell clone defining a novel molecule sTA which is not expressed on resting human peripheral blood CD3+ cells but strongly expressed on a fraction of TCR-gamma delta-bearing clones. Like receptors for growth factors such as IL-2, the sTA Ag is present on clones and cell lines according on the cell cycle. SDS-PAGE analysis of sTA immunoprecipitates from 125I-labeled sTA+ clone lysate demonstrated a single band of molecular mass 110 kDa under reducing conditions. Triggering with anti-sTA mAb did not result in [Ca2+]i mobilization of sTA+ clones. Additionally, the presence of anti-sTA did not alter the cytotoxicity of these sTA+ clones neither against tumor target cells nor against specific PHA blast cells. Interestingly, due to the fact that most sTA+ clones fail to proliferate in response to CD3 triggering, it appears that sTA may serve as a useful marker to study the functional heterogeneity of TCR-gamma delta expressing cells.
The gag-specific cytotoxic T lymphocytes in rhesus monkeys infected with the simian immunodeficiency virus of macaques.
Authors: Miller M D , Lord C I , Stallard V , Mazzara G P , Letvin N L
Published: December 2022

The simian immunodeficiency virus of macaques (SIVmac) is a lentivirus which induces an AIDS-like disease in rhesus monkeys. We have explored the virus-specific cellular immune response in SIVmac-infected rhesus monkeys. Con A-activated, IL-2 expanded PBL of some SIVmac-infected rhesus monkeys lyse autologous B lymphoblastoid cell lines infected with a recombinant vaccinia virus that carries the SIVmac gag gene. This lysis is mediated by CD8+ lymphocytes and is MHC class I restricted. Moreover, these effector lymphocytes do not express the NK cell-associated molecules NKH1 or CD16. These cells are, therefore, CTL. In a limited prospective study of SIVmac-infected rhesus monkeys, the presence of the SIVmac gag-specific CTL activity in PBL correlated with both a reduced efficiency in isolating SIVmac from PBL of these monkeys and their extended survival. This method for assessing SIVmac gag-specific cellular immunity in rhesus monkeys will be important not only in investigating the immunopathogenesis of SIVmac-induced disease, but also in evaluating the capacity of candidate AIDS vaccines to elicit a cell-mediated immune response in this animal model.
Glucocorticoid regulation of the humoral immune system. I. In vivo effects of dexamethasone on IgA and IgG in serum and at mucosal surfaces.
Authors: Wira C R , Sandoe C P , Steele M G
Published: December 2022

The present studies were undertaken to determine whether glucocorticoids influence the levels of Ig in serum, saliva, and vaginal secretions. When measured by RIA, IgA levels in serum were elevated when increasing doses of dexamethasone, a potent synthetic glucocorticoid, were administered to intact- and adrenalectomized-ovariectomized rats. In contrast, IgA levels decreased in saliva and vaginal secretions over the same dose range. Time course studies indicated that the decline in salivary IgA, observed at 24 h after a single injection of dexamethasone, preceded a rise in serum IgA detected at 24 h after the second hormone treatment. Both responses were maximal at day 2 and did not change with further hormone exposure. After immunization and boosting with SRBC at two mucosal sites (intestinal Peyer's patch and uterine lumen), dexamethasone increased anti-SRBC IgA antibody levels in serum and reduced their presence in vaginal secretions. In contrast, anti-SRBC IgG-antibody levels in serum and vaginal secretions were reduced with hormone treatment. In the absence of hormone treatment, pooled sera from nonimmunized animals, when analyzed by HPLC, contained polymeric and dimeric IgA that was present in roughly equal proportion. In response to dexamethasone, polymeric IgA increased to a greater extent than did monomeric IgA. In summary, these studies demonstrate that dexamethasone alters the levels of IgA as well as specifically directed IgA and IgG antibodies in secretions and serum. Further, it suggests that glucocorticoid controlled IgA increases in serum and decreases in vaginal and salivary secretions may be due, in part, to a redistribution of polymeric IgA from mucosal surfaces to serum.
Induction of IL-1 release through stimulation of the C3b/C4b complement receptor type one (CR1, CD35) on human monocytes.
Authors: Bacle F , Haeffner-Cavaillon N , Laude M , Couturier C , Kazatchkine M D
Published: December 2022

Soluble polymerized human C3b and C3b bound to Sepharose induced the production of cell-associated IL-1 and the extracellular release of IL-1 activity from cultured human adherent monocytes. Cultures were performed in serum-free conditions in the presence of polymyxin B and of the PG synthesis inhibitor indomethacin. Induction of intracellular IL-1 activity was associated with that of IL-1 alpha and IL-1 beta Ag. Biologically active released IL-1 was IL-1 beta. Monomeric C3b induced cell-associated IL-1 but not the release of IL-1 from monocytes. IL-1 production was also induced by stimulation of CR1 on adherent monocytes with anti-C3b receptor (CR1) antibody. The ability of multivalent C3b to induce IL-1 production could be dissociated from that of possibly contaminating LPS by several criteria including: the lack of detectable LPS in purified C3b; functional inhibition of LPS in cultures with polymyxin B; the lack of induction of extracellular IL-1 by C3b monomer; a correlation between the expression of CR1 on monocytes and the monocytic response to C3b; a dissociation between the expression of CR1, and the expression of LPS binding sites and the IL-1 response of monocytes to endotoxin. Induction of IL-1 represents a novel pathway by which C3b-CR1 interactions may modulate the immune response.
Modulation of platelet responses to collagen by Clq receptors.
Authors: Peerschke E I , Ghebrehiwet B
Published: December 2022

We recently described specific binding sites for Clq on human blood platelets that cross-react with antibodies against Clq receptors (ClqR) on lymphoblastoid cells. Inasmuch as Clq inhibits collagen-induced platelet aggregation, we compared the effects of ClqR occupancy by purified Clq, monoclonal (IIl/B5) and polyclonal anti-ClqR antibodies on collagen-induced platelet adhesion, release, and aggregation. Washed platelets in buffered Tyrode's solution containing 2 mM magnesium were preincubated (30 min, 22 degrees C) with antibodies, Clq, or appropriate control buffers and antiserum. Platelet aggregation and release measurements were made using 14C-serotonin-labeled platelets stimulated with type I collagen. Adhesion assays were performed in the presence of magnesium under static conditions at 22 degrees C with 51Cr-labeled platelets and collagen-coated microtiter wells. Concentrations of IIl/B5, polyclonal anti-Clq R antiserum, or Clq causing 82 to 92% (n = 7) inhibition of collagen-induced release and 67 to 98% inhibition of aggregation, failed to inhibit magnesium-dependent platelet adhesion to collagen. Inasmuch as divalent cation-independent platelet-collagen interactions have also been described, further studies were performed to compare the divalent cation requirement of ClqR occupancy by Clq and inhibition of platelet-collagen interactions. Whereas, Clq binding to platelets was divalent cation independent, neither Clq nor anti-ClqR antibodies prevented platelet adhesion to collagen in the presence of EDTA. These data suggest that under defined in vitro conditions, ClqR modulate collagen-induced platelet aggregation and secretion, but not platelet adhesion.
Expression of two distinct cytolytic mechanisms among murine CD4 subsets.
Authors: Ju S T , Ruddle N H , Strack P , Dorf M E , DeKruyff R H
Published: December 2022

A TNF (TNF-alpha and TNF-beta)-sensitive target, L929, and two TNF-resistant targets, P815 and LK were used to compare the cytolytic activity among subsets of CD4+ (Th) clones. Cytolytic activity was induced with either Con A, CD3-mAb, or Ag-pulsed LK cells. Six Th1 clones are strongly cytolytic against all three targets. In contrast, Th2 clones are either noncytolytic or weakly cytolytic. Although there is an apparent correlation between TNF production, killing of L929 cells, and the killing of TNF-resistant targets, an anti-TNF serum (capable of neutralizing both TNF-alpha and TNF-beta) selectively inhibits CD4 clones to lyse L929 cells, whereas the lysis of P815 or LK cells was unaffected. The continuous presence of noncytotoxic levels of Actinomycin D (AcD) and cycloheximide, but not mitomycin C, cyclosporin A (CsA), or cholera toxin (ChT) inhibits the lysis of Ag-pulsed, Ia-bearing LK cells; indicating a requirement for de novo synthesis of RNA and protein for cytolytic activity. Although pretreatment with AcD, CsA, or ChT strongly inhibits production of IL-2, TNF and IFN-gamma, only clones pretreated with AcD lose cytolytic activity against Ag-pulsed, Ia-bearing LK cells. These observations support a model of TNF-independent killing of TNF-resistant targets. The TNF-independent cytolytic activity does not correlate with serine esterase activity released into media upon activation of CD4 clones. Moreover, the effects of metabolic inhibitors on serine esterase release do not correlate with their effects on cytolytic activity. Collectively, the data demonstrate that activated CD4 cells express two distinct cytolytic activities; a TNF (and IFN-gamma)-mediated cytotoxicity and a TNF (and IFN-gamma)-independent cytolytic activity. Both pathways require de novo synthesis of RNA and protein and appear to be independent of granule enzyme release. Only the TNF-independent cytolytic activity is resistant to CsA and ChT inhibition.
Distinct mechanisms of suppression of murine T cell activation by the related macrolides FK-506 and rapamycin.
Authors: Dumont F J , Staruch M J , Koprak S L , Melino M R , Sigal N H
Published: December 2022

FK-506 and the structurally related macrolide rapamycin (RAP) were investigated in comparison with cyclosporin A (CsA) for their immunosuppressive effects on murine T cells. All three agents suppressed the proliferation of splenic T cells triggered by lectins or antibodies to CD3 and Ly-6C. FK-506 or CsA also inhibited proliferation, IL-2 production, and IL-2R expression in splenic T cells activated with ionomycin + PMA. However, RAP minimally affected IL-2 production and IL-2R expression in these cells, although it reduced proliferation. Similarly, FK-506 and CsA, but not RAP, suppressed IL-2 production by activated DO.11.10 T hybridoma cells. In such a system, as well as in normal T cells stimulated with high ionomycin concentrations, FK-506 and CsA enhanced proliferation, indicating that they both abrogate negative signals associated with T cell activation. On the contrary, RAP diminished the autonomous proliferation of hybridoma cells, whereas FK-506 and CsA had little effect. The proliferative response induced in D10.G4 cells by IL-1 + ionomycin but not that induced by IL-1 + PMA was sensitive to inhibition by FK-506 and CsA. In contrast, RAP inhibited equally well both types of stimulation. Finally, T cell proliferation driven by IL-2 or IL-4 was found to be relatively resistant to FK-506 or CsA but sensitive to RAP. Altogether, these data demonstrate that FK-506 and CsA alter similar calcium-associated events of T cell activation and block T cell proliferation primarily by suppressing lymphokine production. RAP interferes with a different set of events and inhibits T cells by impairing their response to growth-promoting lymphokines.
Necessity of IgE antibodies and mast cells for manifestation of resistance against larval Haemaphysalis longicornis ticks in mice.
Authors: Matsuda H , Watanabe N , Kiso Y , Hirota S , Ushio H , Kannan Y , Azuma M , Koyama H , Kitamura Y
Published: December 2022

Genetically mast cell-deficient WBB6F1-W/Wv mice showed an apparent defect in manifestation of the resistance against larval Haemaphysalis longicornis ticks, but their serum IgE levels increased more than 100-fold after the second tick infestation. Immune sera obtained from the WBB6F1-W/Wv mice were adoptively transferred to the other WBB6F1-W/Wv mice which had received intracutaneous injections of WBB6F1-+/+ mouse-derived cultured mast cells. Because the resistance against ticks was detectable only when both mast cells and IgE antibodies were available, immediate hypersensitivity reaction appeared to have a physiologic role in the manifestation of the resistance against H. longicornis ticks.
Alterations in the pattern of arachidonate metabolism accompany rat macrophage differentiation in the lung.
Authors: Peters-Golden M , McNish R W , Hyzy R , Shelly C , Toews G B
Published: December 2022

The present study was undertaken to investigate the changes in arachidonic acid (AA) metabolism which accompany rat macrophage (m phi) differentiation in the lung in order to determine whether these changes occur in the alveolar space or in the pulmonary interstitium, as well as the mechanisms responsible for such changes. Metabolism of endogenous and exogenous AA by cultured m phi obtained from the peritoneum (PM), the pulmonary interstitium (IM), and the alveolar spaces (AM) was examined by using HPLC and RIA. Although PM and AM released similar amounts of endogenous AA in response to both ionophore A23187 and the particulate zymosan, PM metabolized AA predominantly to cyclooxygenase (CO) products, whereas AM produced predominantly 5-lypoxygenase (5-LO) metabolites. IM synthesized a profile of eicosanoids which more closely resembled that of PM. Studies of the metabolism of exogenously supplied AA demonstrated that AM indeed had less CO activity than did PM. PM, but not AM, CO activity decreased during prolonged culture in air, suggesting the possibility that oxidative inactivation of CO plays a role in the decline in CO capacity which accompanies m phi differentiation in the lung. In contrast, the greater expression of 5-LO metabolism in AM than PM did not reflect mere differences in enzyme capacity, since upon activation of protein kinase C with PMA or oleoylacetylglycerol, ionophore-stimulated PM produced amounts of 5-LO products which were comparable to the amounts produced by AM stimulated with A23187 alone. These results indicate that increases in 5-LO metabolism and decreases in CO metabolism accompany rat m phi differentiation in the lung, that these changes occur largely in the alveolar space, and that the increased 5-LO capacity and decreased CO capacity are independently regulated by different mechanisms.
Activated macrophages destroy intracellular Leishmania major amastigotes by an L-arginine-dependent killing mechanism.
Authors: Green S J , Meltzer M S , Hibbs J B , Nacy C A
Published: December 2022

Macrophages infected with amastigotes of Leishmania major and treated with IFN-gamma in vitro develop potent antimicrobial activities that eliminate the intracellular parasite. This antileishmanial activity was suppressed in a dose dependent fashion by NG-monomethyl-L-arginine (NGMMLA), a competitive inhibitor of nitrite, nitrate, nitric oxide and L-citrulline synthesis from L-arginine. Excess L-arginine added to infected macrophage cultures reversed the inhibitory effects of NGMMLA. Addition of arginase to culture media inhibited intracellular killing by IFN-gamma-treated cells. Similar effects were seen with macrophages obtained from BCG-infected C3H/HeN mice. Increased levels of nitrite, an oxidative product of the L-arginine-dependent effector mechanism, was measured in cultures of infected IFN gamma-treated macrophages as well as infected BCG-activated macrophages. Nitrite production correlated with development of antileishmanial activity. Nitrite production and microbicidal activity both decreased when in vivo or in vitro-activated macrophages were cultured in the presence of either arginase or NGMMLA. Nitric oxide synthesized from a terminal guanidino nitrogen atom of L-arginine and a precursor of the nitrite measured, may disrupt Fe-dependent enzymatic pathways vital to the survival of amastigotes within macrophages.
Cross-antigenicity between the major surface proteins (ospA and ospB) and other proteins of Borrelia burgdorferi.
Authors: Jiang W , Luft B J , Munoz P , Dattwyler R J , Gorevic P D
Published: December 2022

Two of the major surface Ag of Borrelia burgdorferi, the 31-kDa OspA and 34-kDa OspB proteins, are encoded by a 49-kb plasmid. In this study, mAb and monospecific polyclonal antibodies were used to define cross-antigenicity of the OspA and OspB protein to each other and to other lower molecular mass proteins by Western blot analysis. Two mAb studied, 105.5 and 184.1, were directed predominantly against the 31-kDa OspA protein. However, each also reacted with other minor bands, though with different specificities. Using V8 protease digestion and cleavage by cyanogen bromide, we demonstrated that each mAb reacted to the 31-kDa protein differently. Monospecific polyclonal rabbit and human antibodies directed against the 34-, 31-, 22-, and 20-kDa proteins were eluted from blots and used to further corroborate the cross-reactivity among these Ag. Rabbit antibodies to the 31- and 22-kDa Ag gave remarkably similar peptide maps after V8 protease digestion of the 31-kDa OspA protein, as did mAb 184.1, suggesting that this mAb recognized an immunodominant epitope common to the 22- and 31-kDa proteins. It seems likely therefore that the humoral immune response to Borrelia surface Ag may be due to a limited number of cross-reactive epitopes on distinct, but related, gene products.
Elevated natural killer cell responses in mice infected with recombinant vaccinia virus encoding murine IL-2.
Authors: Karupiah G , Coupar B E , Andrew M E , Boyle D B , Phillips S M , Müllbacher A , Blanden R V , Ramshaw I A
Published: December 2022

The role of cytotoxic T cells and NK cells in the recovery of immunodeficient, athymic, nude mice infected with a recombinant vaccinia virus (VV) encoding murine IL-2 was investigated. Kinetic studies with the IL-2-encoding recombinant (VV-HA-IL2) and control (VV-HA-TK) viruses excluded a role for cytotoxic T cells but suggested the possible involvement of NK cells. In athymic nude mice given VV-HA-IL2, NK activity was at least threefold higher than mice infected with VV-HA-TK and this activity persisted for at least 6 days after infection. The effectors mediating the NK-like activity were asialo-GM1+ (as-GM1+), Thy1.2+/-, CD4- and CD8-, the phenotype of conventional NK cells. Elevated NK activity coincided with the rapid clearance of VV-HA-IL2 from ovaries of infected normal CBA/H mice but not from ovaries of CBA beige mice which had no detectable NK activity in spleens or ovaries. The expression of IL-2 in recombinant VV infection probably induces a cascade of immunologic effects of which elevated NK activity is one. We speculate that the chemoattractant and NK activity augmenting effects of IL-2 may contribute to recovery from VV-infection.