bluuebunny/crossref_metadata_2025_split huggingface dataset:
Analysis of the autoantibody response to fibrillarin in human disease and murine models of autoimmunity.
Authors: Takeuchi K , Turley S J , Tan E M , Pollard K M 
Published: December 2022

Abstract
Fibrillarin, a component of the U3 RNP particle, is a target for the spontaneously arising autoantibodies in human scleroderma and a monoclonal autoantibody (72B9) derived from the autoimmune mouse strain (NZB x NZW) F1. Autoantibodies against fibrillarin can also be induced in H-2s mice by treatment with mercuric chloride (HgCl2). The objective of this study was to compare the spontaneously occurring anti-fibrillarin autoantibody response with the autoantibody response induced by HgCl2 treatment. Immunofluorescence microscopy on human HEp2, mouse 3T3, and Xenopus XIK-2 cells, immunoblotting with use of nuclear extract from human MOLT-4, mouse 3T3, and Xenopus XIK-2 cells, and immunoprecipitation with use of in vitro translation products of RNA transcripts from yeast fibrillarin cDNA were used in this analysis. Both spontaneous and induced autoantibodies displayed common reactivity in that, irrespective of the antigenic source, they gave the same nucleolar immunofluorescence pattern and a restricted immunoblotting reactivity targeting predominantly the 34-kDa protein fibrillarin. Immunoprecipitation of N- and C-terminal truncated fibrillarin constructs also demonstrated a common pattern of reactivity. All Abs precipitated a fragment comprising amino acids 1-312 but not a smaller fragment containing amino acids 1-257. The majority of sera could not precipitate an N-terminal truncated molecule consisting of amino acids 157-327. These immunoprecipitation experiments support recognition of a common epitope requiring both N- and C-regions, which may be exemplified by the reactivity of murine monoclonal anti-fibrillarin autoantibody 72B9. Our results indicate that spontaneous human and toxin-induced murine autoantibodies to fibrillarin share common reactivity against this highly conserved nucleolar protein.
The NK1.1 antigen in NK-mediated F1 antiparent killing in vitro.
Authors: Kung S K , Miller R G 
Published: December 2022

Abstract
NK cells in lethally irradiated F1(A x B) hybrid mice can reject parental A or B strain bone marrow cells, a phenomenon called "hybrid resistance." The recognition mechanism used by the NK cells remains unknown. Our laboratory has previously described an in vitro model for hybrid resistance, and we have used it here to test whether the NK surface marker, NK1.1, is involved in such recognition. We found that 1) an anti-NK1.1 mAb (PK136) inhibited F1 lymphokine-activated killer (LAK) antiparent lysis if the LAK expressed NK1.1. Other mAb, even a mAb such as 2B4 that recognizes the same LAK as PK136, did not produce inhibition. 2) The F(ab')2 fragment of PK136 also inhibited lysis. 3) F1 LAK generated from athymic nude mice were as effective antiparent killers as LAK from normal mice and were equally inhibitable by anti-NK1.1 mAb, strengthening the conclusion that killing is mediated by NK cells and not T cells. 4) As previously shown by others, addition of anti-NK1.1 mAb to a mixture of NK1.1+ LAK cells and NK-resistant FcR+ cells allowed lysis of the FcR+ cells via "redirected lysis," in which the anti-NK1.1 mAb binds to NK1.1 on the NK cells and FcR on the target cell. The ability of anti-NK1.1 mAb to inhibit direct lysis and enhance redirected lysis is most consistent with NK1.1 being a receptor involved in NK activation.
Regulation of the production of the RANTES chemokine by endothelial cells. Synergistic induction by IFN-gamma plus TNF-alpha and inhibition by IL-4 and IL-13.
Authors: Marfaing-Koka A , Devergne O , Gorgone G , Portier A , Schall T J , Galanaud P , Emilie D 
Published: December 2022

Abstract
Production by endothelial cells of the regulated on activation normal T expressed and secreted chemokine (RANTES) has recently been evidenced during delayed-type hypersensitivity (DTH) reactions and may contribute to the local accumulation of macrophages and CD4+ memory T lymphocytes. To document the mechanism inducing RANTES production in this condition, we analyzed the effect of cytokines known to influence the formation of DTH granulomas. Little or no RANTES was produced after stimulation of HUVEC with IFN-gamma, IL-1 beta, or TNF-alpha. However, the combination TNF-alpha+IFN-gamma induced a strong RANTES production. In situ hybridization experiments with a RANTES probe showed that this synergy was also observed at the mRNA level and that the effect of the combination was mainly to increase the amount of RANTES mRNA per cell. The expression of the luciferase gene under the control of the RANTES gene regulatory elements was analyzed; TNF-alpha and the combination TNF-alpha+IFN-gamma activated the regulatory elements. Sequential treatment of HUVEC with TNF-alpha and IFN-gamma showed that IFN-gamma sensitized HUVEC to the stimulating effect of TNF-alpha. The production of RANTES induced by TNF-alpha+IFN-gamma was partly but significantly inhibited by the Th2-type cytokines IL-4 and IL-13. In contrast, IL-10 had no effect. These results indicate that the microenvironment of DTH granulomas, containing high levels of both TNF-alpha and IFN-gamma, may be responsible for RANTES production by perigranulomatous endothelial cells. Inhibition of this production by Th2-type cytokines may be a mechanism by which these cytokines interfere with the formation of DTH granulomas.
Ligand binding by the IL-2 receptor is modulated by intracellular determinants of the IL-2 receptor beta-chain.
Authors: Goldsmith M A , Amaral M C , Greene W C 
Published: December 2022

Abstract
The biologic actions of IL-2 are mediated by the IL-2R, a multisubunit receptor complex displayed on the surface of lymphocytes and select other hematopoietic lineages. The IL-2R exhibits multiple affinities for IL-2 that result from the monomeric (alpha), heterodimeric (alpha beta and beta gamma), and heterotrimeric (alpha beta gamma) assembly of different receptor subunits. In the present studies, we have used a series of IL-2R mutants in a transient mammalian expression system to investigate the potential role of intracellular receptor regions in the ligand-binding functions of the IL-2R. Analyses of chimeric and deletion mutants of the IL-2R beta subunit have revealed that its intracellular domain critically and selectively influences high affinity ligand binding mediated through the extracellular domains of the alpha beta-heterodimeric receptor. In contrast, intermediate affinity binding of IL-2 by beta gamma-heterodimeric receptors exhibits no dependence on the cytoplasmic domain of IL-2 R beta. Further, co-expression of either a full-length or severely truncated form of IL-2 R gamma to generate an alpha beta gamma-heterotrimeric complex also overcomes the functional dependence upon the cytoplasmic tail of IL-2 R beta. Collectively, our findings suggest that the cytoplasmic domain of IL-2R beta produces intrasubunit transmembrane conformational changes in this receptor subunit that promote extracellular IL-2 binding in combination with IL-2R alpha. These findings have important implications for the receptor dynamics involved in both ligand binding and signal transduction as well as for clinical applications pertaining to altering IL-2R function.
B cell superstimulatory influenza virus (H2-subtype) induces B cell proliferation by a PKC-activating, Ca(2+)-independent mechanism.
Authors: Rott O , Charreire J , Semichon M , Bismuth G , Cash E 
Published: December 2022

Abstract
The influenza virus hemagglutinin glycoprotein (HA) induces a vigorous B cell proliferation and Ig-synthesis by an unknown activation mechanism, which is susceptible to the inhibitory effects of anti-Ig and anti-class II mAbs. To gain further insight into the activation mode of this T cell-independent, B cell "superstimulatory" virus, we analyzed the sensitivity of H2-subtype virus-mediated B cell activation to the inhibitory effects of various signal transduction-blocking agents and compared it to the well characterized anti-mu-mediated and the LPS-employed pathway. Cyclic-AMP agonists (cAMP-analogues, pentoxifylline, cholera toxin, and forskolin) blocked HA-mediated activation of B cells only at concentrations at least 50-fold higher than required for blocking of anti-mu-induced activation. However, HA-treatment failed to induce an increase in intracellular cAMP levels in responding B cells. The B cell response to HA was highly resistant to calcineurin-inhibitory cyclosporin-A treatment and did not result in a measurable Ca2+ influx. Similarly, HA failed to induce an increase in tyrosine phosphorylations, including phosphorylation of phospholipase C gamma 2. HA-activated B cells showed an increase in membrane-associated protein kinase C activity, and depletion of protein kinase C by pretreatment of B cells with phorbol esters inhibited a subsequent activation by HA. Collectively, our results provide a new example of B cell stimulation by multivalent type-2 Ags, which seems to be mediated by a phosphatidylinositol- and Ca(2+)-independent signaling pathway.
Regulation of lymphocyte pseudopodia formation by triggering the integrin alpha 4/beta 1.
Authors: Szabo M C , Teague T K , McIntyre B W 
Published: December 2022

Abstract
Video microscopy and digital imaging were used to quantitatively analyze lymphocyte adhesion and formation of pseudopodia on the extracellular matrix protein fibronectin (FN). A morphology kinetics assay comparing pseudopodial extension values over a 24-h period showed that HPB-ALL T leukemic cells undergo a wave of morphologic change, returning to a round shape after 8 h. Using anti-alpha 4 and anti-alpha 5 mAbs and a panel of cell types that are single or double positive for expression of the alpha 4/beta 1 and alpha 5/beta 1 FN binding integrins, it was determined that cell adhesion to FN was influenced by both beta 1-integrins, whereas alpha 4/beta 1 was found to be the major FN receptor mediating pseudopodia extension. The protein kinase inhibitor staurosporine, the protein kinase C inhibitors calphostin C and chelerythrine, and the protein tyrosine kinase inhibitor herbimycin A blocked pseudopodial extension in HPB-ALL cells. In contrast, two cAMP-dependent protein kinase inhibitors H8 and H89 did not inhibit. Inhibitors of phospholipase A2, lipoxygenases, and cyclooxygenases could block formation of pseudopodia, yet had little or no effect on cell adhesion to FN. The preincubation of cells with arachidonic acid could prevent the inhibition mediated by the reversible phospholipase A2 inhibitor cibacron blue. We conclude that the formation of lymphocyte pseudopodia in response to FN can utilize the adhesive and signaling activities of the alpha 4/beta 1-integrin and the enzymatic activities of protein kinases and phospholipases.
Transgenic control of perforin gene expression. Functional evidence for two separate control regions.
Authors: Lichtenheld M G , Podack E R , Levy R B 
Published: December 2022

Abstract
Perforin is a pore-forming effector molecule of CTL and NK cells. To characterize perforin gene expression and its transcriptional control mechanisms in vivo, expression of a cell surface tag, i.e., human CD4, was driven by 5.1 kb of the murine perforin 5' flanking and promoter region in transgenic mice. Six out of seven transgenic lines expressed the perforin-tag hybrid gene at low to intermediate levels, depending on the integration site. Tissues not yet reported to contain perforin-expressing lymphocytes were identified. Transgene expression occurred in all cells that physiologically are able to express perforin, i.e., in T cells and NK cells, and in some T cells that normally may express little or no perforin. At the whole organ level, significant amounts of transgenic mRNA and endogenous perforin mRNA were co-expressed in the lymphoid organs, as well as in the lung, the ileum, the oviduct/uterus, and the bone marrow. At the single cell level, the perforin tag was present on NK cells and on CD8+, as well as on CD4+ T cells. Also targeted were Thy-1.2+ gamma delta T cells, but not Thy1.2- gamma delta T cells, B cells, nor monocytes. During thymic T cell development, transgene expression occurred in double negative (CD4-CD8-) thymocytes and was detected at all subsequent stages, but exceeded the expression levels of the endogenous gene in the thymus. In conclusion, the analyzed perforin 5' flanking and promoter region contains important cis-acting sequences that restrict perforin expression to T cells and NK cells, and therefore provides a unique tool for manipulating T cell and/or NK cell-mediated immune responses in transgenic mice. On the other hand, the normal control of perforin gene expression involves at least one additional negative control mechanism that was not mediated by the transgenic promoter and upstream region. This control restricts perforin gene expression in thymically developing T cells and in most resting peripheral T cells, but can be released upon T cell activation.
Binding analysis of antibodies produced by precursor and branchpoint intermediates of an anti-influenza hemagglutinin B cell clone. Parallel replacement mutations do not confer increased avidity for hemagglutinin.
Authors: Rickert R C , Wloch M K , Hahn R L , Clarke S H 
Published: December 2022

Abstract
We have previously described a set of secondary (2 degrees) hybridomas that are specific for the Sb site of influenza hemagglutinin (HA(Sb)) and share the H37-68 ld (68ld). Cells producing 68ld Abs dominated the 2 degree response of one BALB/c mouse, but were virtually absent from the primary (1 degree) and 2 degree responses of other BALB/c mice. To understand the basis for the idiosyncratic nature of this response, we have assessed the functional importance of a conserved DJ junctional sequence. We find that substitutions of conserved residues within this sequence drastically reduced HA(Sb) binding, providing an explanation for the absence of this ld from 1 degree responses. We have also examined intraclonal affinity maturation by generating Abs inferred to have been produced by the unmutated precursors and branchpoint intermediates of the largest 68ld clone. Interestingly, unmutated Abs of this clone bound HA(Sb) well, comparable to that of some 2 degree 68ld hybridoma Abs. Abs produced by branchpoint intermediates of this clone have a lower affinity for HA(Sb) than their unmutated precursor, despite containing parallel amino acid replacement mutations. Thus, expansion of this clone did not follow the paradigm of stepwise increases in affinity. Together, these data underscore the interconnective nature of precursor frequency and somatic mutation in the formation of the 2 degree response to a given Ag.
Addition of a mu-tailpiece to IgG results in polymeric antibodies with enhanced effector functions including complement-mediated cytolysis by IgG4.
Authors: Smith R I , Coloma M J , Morrison S L 
Published: December 2022

Abstract
The 18-amino acid carboxyl-terminal tailpiece from IgM (mutp) has now been added to the carboxyl-termini of IgG1, IgG2, IgG3, and IgG4 constant regions to produce recombinant IgM-like IgGs. Polymeric IgGs obtained by this approach possess up to six Fcs and 12 antigen-combining sites, greatly increasing the avidity of their interactions with other molecules. Not surprisingly, the C activity of normally active IgG1 and IgG3 and somewhat less active IgG2 Abs is shown to be dramatically enhanced upon polymerization. The multiple Fcs present in a single molecule apparently allow for more efficient interactions with the multiple C1q heads present in C1, the first component of the classical C cascade. An unexpected result however, is that IgG4, normally devoid of C activity, when polymerized in the same fashion directs C-mediated lysis of target cells almost as effectively as the other polymers. Interestingly though, IgG4mutp does not deplete C activity in a standard consumption assay using soluble Ag. The other gamma mutp isotypes are capable of depleting 100% of the serum lytic ability even in the absence of Ag, whereas IgG4mutp shows no evidence of activity in this assay under any of the conditions tested. Additionally, we show that, in contrast to monomeric IgG, polymeric IgGs bind with very high affinity to Fc gamma receptor II (Fc gamma RII), a low affinity receptor for wild-type antibodies; however, binding to Fc gamma Rl, the high affinity receptor, appears to be unaltered. Finally, the in vivo t1/2 of the gamma mutp proteins is decreased relative to wild-type IgG, apparently because of rapid clearance of the polymeric fraction.
Secondary deletional recombination of rearranged switch region in Ig isotype-switched B cells. A mechanism for isotype stabilization.
Authors: Zhang K , Cheah H K , Saxon A 
Published: December 2022

Abstract
Analysis of the Ig switch (S) region structure from many stable isotype-switched B cells reveals that the majority of the ultimate switch recombination sites are located at the 5' end, or even upstream of the 5' end of the Smu region. These findings strongly contrast with results from switch circle analysis which reveal that the primary switch recombination sites are preferentially distributed in the middle or at the 3' end of the Smu region. Secondary deletion within recombined chimeric S regions has been proposed as a mechanism to account for the different results. We directly tested whether secondary deletion of rearranged chimeric S regions indeed occurs in human cells. Circular DNAs representing secondary deletion events in gamma switched cells were isolated and characterized by using a specially designed PCR-based approach. Nucleotide sequence analysis revealed that all clones had the S gamma 1-Smu-S gamma 1 structure. Thus, these cloned fragments resulted from secondary deletion/recombination events within a chimeric Smu-S gamma 1 switch region, i.e., rearrangement between its 5' Smu and 3' S gamma 1. Analysis of the Smu region from stimulated B cells also revealed that some Smu regions undergo an internal deletion/rearrangement between the 5' and 3' ends. These results definitively demonstrate that secondary deletion/recombination of chimeric S regions of isotype-switched B cells occurs. Such secondary deletion/recombination events potentially can be responsible for isotype stabilization of switched B cells, as the active Ig gene may have an insufficient amount of retained chimeric switch sequence to serve as a substrate for further S-S recombination.
Induction of tumor-reactive CTL from peripheral blood and tumor-infiltrating lymphocytes of melanoma patients by in vitro stimulation with an immunodominant peptide of the human melanoma antigen MART-1.
Authors: Rivoltini L , Kawakami Y , Sakaguchi K , Southwood S , Sette A , Robbins P F , Marincola F M , Salgaller M L , Yannelli J R , Appella E 
Published: December 2022

Abstract
MART-1 is an Ag expressed on melanomas and melanocytes, and is recognized by the majority of HLA-A2-restricted tumor-specific tumor-infiltrating lymphocytes (TIL) from melanoma patients. In the present study we have analyzed 10 potential 9-mer epitopes containing the HLA-A2.1 binding motifs for their ability to induce melanoma-specific T cell lines. Antimelanoma CTL could be generated only with MART-1(27-35) peptide, which has been previously shown to be recognized by a majority of HLA-A2-restricted TIL. Anti-MART-1(35-43)-specific CTL could also be induced, but these T cells did not recognize melanoma cells. MART-1(27-35)-specific CTL could be effectively generated from a total of 11 of 12 PBL and from 3 of 3 TIL derived from HLA-A2+ melanoma patients, as well as from 2 of 4 PBL from HLA-A2+ healthy donors by in vitro stimulation with autologous PBMC pulsed with the synthetic MART-1(27-35) peptide. These CTL lines specifically lysed and release cytokines (TNF-alpha, IFN-gamma, and GM-CSF) in response to T2 cells pulsed with MART-1(27-35), as well as to HLA-A2+ MART-1+ melanoma cells. CTL generated with MART-1(27-35) also lysed uncultured HLA-A2+ melanoma cells derived from tumor biopsies, indicating that this MART-1 epitope is likely to be expressed in association with HLA-A2 on the surface of tumor cells in vivo. CTL lines generated with MART-1(27-35) mediated 25- to 100-fold higher lytic activity than MART-1-reactive CTL grown from TIL in the presence of high dose IL-2. These results demonstrate that MART-1(27-35) peptide may represent an ideal candidate for Ag-specific immunotherapy in melanoma patients.
Human IL-1 beta processing and secretion in recombinant baculovirus-infected Sf9 cells is blocked by the cowpox virus serpin crmA.
Authors: Howard A D , Palyha O C , Griffin P R , Peterson E P , Lenny A B , Ding G J , Pickup D J , Thornberry N A , Schmidt J A , Tocci M J 
Published: December 2022

Abstract
Biologically active, mature IL-1 beta (mIL-1 beta) is released from activated monocytes after proteolytic processing from an inactive precursor (pIL-1 beta). IL-1 beta converting enzyme (ICE), the first member of a newly discovered family of cysteine proteinases, is required for this processing event. The cleaved cytokine is released from monocytes by an unknown mechanism which does not employ a standard hydrophobic signal sequence. As in mammalian fibroblasts, insect Sf9 cells do not normally process or secrete human IL-1 beta. The expression of active ICE enables Sf9 cells to process 31-kDa pIL-1 beta correctly at Asp27 and Asp116, and to export 17.5-kDa mIL-1 beta. The recombinant heterodimeric human enzyme purified from Sf9 cells possesses a sp. act. of 2.9 +/- 0.5 x 10(6) U/mg and is indistinguishable from native ICE with regard to its subunit composition and catalytic properties. In this system, co-expression of the cowpox virus crmA gene, an extremely potent serpin inhibitor of ICE (Ki &lt; 7 pM), inhibits ICE activation completely and blocks pIL-1 beta processing and mIL-1 beta secretion by approximately 95%. The results indicate that ICE, in addition to its processing function, facilitates the transport of IL-1 beta across the plasma membrane.
Proinflammatory cytokines potentiate thrombospondin-mediated phagocytosis of neutrophils undergoing apoptosis.
Authors: Ren Y , Savill J 
Published: December 2022

Abstract
Apoptosis leads to swift recognition, ingestion, and degradation of intact senescent neutrophils by macrophages. This protects tissues from leakage of noxious contents from dying cells and may promote resolution of inflammation. However, little has been known of the mechanisms that regulate macrophage capacity for apoptotic cells during an inflammatory response. We examined whether proinflammatory cytokines modulated phagocytosis of senescent neutrophils undergoing apoptosis by human monocyte-derived macrophages at 4 days maturity (4d M phi), an in vitro model of neutrophil "disposal" by apoptosis. Pretreatment of 4d M phi with granulocyte-macrophage-CSF increased the proportion of 4d M phi taking up apoptotic PMN in a concentration-dependent fashion by up to approximately 240%. This was by a rapid effect detectable by 4 h and exerted on the M phi, not the PMN. Granulocyte-macrophage-CSF also increased the number of apoptotic PMN taken up by each M phi. IFN-gamma, IL-1 beta, TNF-alpha, and TGF-beta 1 also enhanced phagocytosis, but IL-4 and IL-6 had no effect. In each case, the cytokine-expanded phagocytic subpopulation employed the thrombospondin (TSP)-dependent recognition mechanism defined for mature M phi, in which M phi vitronectin receptor and CD36 cooperate. However, commensurate increases in M phi expression of VnR, TSP, or CD36 were not detectable, indicating that TSP-mediated recognition can be recruited by other mechanisms. Cytokines did not recruit phosphatidylserine-dependent recognition, the other major mechanism by which some macrophage populations ingest apoptotic cells. Thus, M phi phagocytosis of apoptotic neutrophils can be potentiated by proinflammatory cytokines, suggesting a mechanism for negative feedback control of neutrophil number at inflamed sites.