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1.
Eur J Immunol ; 29(11): 3466-77, 1999 11.
Article in English | MEDLINE | ID: mdl-10556801

ABSTRACT

Using the monoclonal antibody C1.7, which recognizes a signaling, membrane-bound molecule on human NK and a proportion of CD8(+) T cells, we cloned a novel molecule we refer to as NK cell activation-inducing ligand (NAIL). It is a 365-amino acid protein that belongs to the immunoglobulin-like superfamily with closest homology to murine 2B4, and human CD84 and CD48. Using a soluble NAIL-Fc fusion protein, we determined the counterstructure for NAIL, CD48, which it binds with high affinity. Stimulation of human B cells with recombinant NAIL in the presence of a suboptimal concentration of human CD40 ligand or IL-4 resulted in increased proliferation. Treatment of human dendritic cells with soluble NAIL-leucine zipper protein resulted in an increased release of IL-12 and TNF-alpha. Using recombinant CD48 protein, we demonstrated the ability of this molecule to increase NK cell cytotoxicity and induce IFN-gamma production. We also showed that 2B4 binds to mouse CD48, suggesting that interaction of these receptors may play a similar role in both species. Taken together these results indicate that the NAIL-CD48 interaction may be an important mechanism regulating a variety of immune responses.


Subject(s)
Antigens, CD/metabolism , Killer Cells, Natural/metabolism , Membrane Proteins/metabolism , Amino Acid Sequence , Animals , B-Lymphocytes/cytology , CD48 Antigen , Cell Division , Cloning, Molecular , Cytokines/biosynthesis , Cytotoxicity, Immunologic , Humans , Immunoglobulin Fc Fragments/metabolism , Interferon-gamma/biosynthesis , Jurkat Cells , K562 Cells , Ligands , Membrane Glycoproteins/metabolism , Membrane Proteins/genetics , Membrane Proteins/immunology , Mice , Molecular Sequence Data , Phosphorylation , RNA, Messenger , Receptors, Immunologic/metabolism , Recombinant Fusion Proteins/metabolism , Signaling Lymphocytic Activation Molecule Family , Tyrosine/metabolism , U937 Cells , Up-Regulation
2.
J Biol Chem ; 274(1): 418-23, 1999 Jan 01.
Article in English | MEDLINE | ID: mdl-9867859

ABSTRACT

Recent progress in the understanding of immune function indicates that the interaction of CD40L with its receptor, CD40, plays a pivotal role in both humoral immunity and cell-mediated defense against pathogens. Functional studies of this interaction on both dendritic cells and malignant cells have demonstrated that CD40L also plays an important role in immune surveillance and anti-tumor immunity. CD40L exists in nature predominantly as a membrane-anchored molecule. To develop CD40L as a potential therapeutic, it is important to optimize soluble forms of this molecule that could be used in a clinical setting. Several reports have shown that soluble forms of CD40L, like CD40 antibodies, are biologically active. In the present report we demonstrate that the incorporation of an isoleucine zipper trimerization motif significantly enhances the biological activity of soluble CD40L.


Subject(s)
Isoleucine/chemistry , Leucine Zippers , Membrane Glycoproteins/metabolism , Animals , Biopolymers , CD40 Antigens/metabolism , CD40 Ligand , CHO Cells , Calorimetry, Differential Scanning , Cricetinae , Electrophoresis, Polyacrylamide Gel , Membrane Glycoproteins/chemistry , Membrane Glycoproteins/isolation & purification , Protein Conformation , Thermodynamics
3.
J Virol ; 70(8): 5557-63, 1996 Aug.
Article in English | MEDLINE | ID: mdl-8764069

ABSTRACT

The Epstein-Barr virus BZLF2 gene encodes a glycoprotein that associates with gH and gL and facilitates the infection of B lymphocytes. In order to determine whether the BZLF2 protein recognizes a B-cell-specific surface antigen, a soluble protein containing the extracellular portion of the BZLF2 protein linked to the Fc portion of human immunoglobulin G1 (BZLF2.Fc) was expressed from mammalian cells. BZLF2.Fc was used in an expression cloning system and found to bind to a beta-chain allele of the HLA-DR locus of the class II major histocompatibility complex (MHC). Analysis of amino- and carboxy-terminal deletion mutants of the BZLF2.Fc protein indicated that the first 90 amino acids of BZLF2.Fc are not required for HLA-DR beta-chain recognition. Site-directed mutagenesis of an HLA-DR beta-chain cDNA and subsequent immunoprecipitation of expressed mutant beta-chain proteins using BZLF2.Fc indicated that the beta1 domain, which participates in the formation of peptide binding pockets, is required for BZLF2.Fc recognition. The addition of BZLF2.Fc to sensitized peripheral blood mononuclear cells in vitro abolished their proliferative response to antigen and inhibited cytokine-dependent cytotoxic T-cell generation in mixed lymphocyte cultures. Flow-cytometric analysis of Akata cells induced to express late Epstein-Barr virus antigens indicated that expression of BZLF2 did not result in reduced surface expression levels of MHC class II. The ability of BZLF2.Fc to bind to the HLA-DR beta chain suggests that the BZLF2 protein may interact with MHC class II on the surfaces of B cells.


Subject(s)
Glycoproteins , HLA-DR Antigens/immunology , Herpesviridae Infections/immunology , Herpesvirus 4, Human/immunology , Lymphocytes/virology , Tumor Virus Infections/immunology , Viral Proteins/immunology , Amino Acid Sequence , Antigen Presentation , Cell Line , Herpesviridae Infections/virology , Humans , Lymphocytes/immunology , Molecular Sequence Data , Protein Binding , Tumor Virus Infections/virology , Viral Proteins/genetics
4.
Int Hist Nurs J ; 1(3): 55-60, 1996.
Article in English | MEDLINE | ID: mdl-11619078

ABSTRACT

In the concluding part of her war diary, Brenda MacDuff, a nurse with the Colonial Nursing Service in Malaya, tells of her final incarceration, eventual freedom and reunion with her husband.


Subject(s)
Colonialism , Concentration Camps , Nursing , Prisoners , Warfare , History, 20th Century , Japan , Malaysia , United Kingdom
5.
J Immunol ; 155(12): 5483-6, 1995 Dec 15.
Article in English | MEDLINE | ID: mdl-7499828

ABSTRACT

A cDNA encoding human IL-17 (hIL-17) was cloned from a CD4+ T cell library. The predicted 155-amino acids sequence contains an N-terminal signal peptide and exhibits 72% amino acid identity with HVS13, an open reading frame from a T-lymphotropic Herpesvirus saimiri, and 63% with murine CTLA8. High levels of hIL-17 were induced from primary peripheral blood CD4+ T cells upon stimulation. When expressed in CV1/EBNA cells, recombinant hIL-17 was secreted in both glycosylated and nonglycosylated forms. A hIL-17.Fc fusion protein and supernatants from cells transfected with hIL-17 induced IL-6 and IL-8 production and enhanced the surface expression of the intracellular adhesion molecule-1 (ICAM-1) in human fibroblasts.


Subject(s)
DNA, Complementary/analysis , Interleukins/biosynthesis , Interleukins/genetics , RNA, Messenger/analysis , Amino Acid Sequence , Base Sequence , Blotting, Northern , CD4-Positive T-Lymphocytes/metabolism , Cells, Cultured , Cloning, Molecular , Fibroblasts/drug effects , Humans , Intercellular Adhesion Molecule-1/biosynthesis , Interleukin-17 , Interleukin-6/biosynthesis , Interleukin-8/biosynthesis , Interleukins/pharmacology , Molecular Sequence Data
6.
Int Hist Nurs J ; 1(2): 83-96, 1995.
Article in English | MEDLINE | ID: mdl-11619073

ABSTRACT

In Part 1 of her war diary, Brenda MacDuff, a nurse with the Colonial Nursing Service in Malaya, tells of her early experiences in the country at the outbreak of war in the East and of her subsequent capture by the Japanese Army.


Subject(s)
Colonialism , Nursing , Prisoners , Warfare , History, 20th Century , Japan , Malaysia , United Kingdom
7.
J Immunol ; 154(2): 483-90, 1995 Jan 15.
Article in English | MEDLINE | ID: mdl-7814861

ABSTRACT

The identification and cloning of the novel cytokine IL-15 were recently described. IL-15 is produced by a wide range of cell types, with the highest levels of IL-15 mRNA being detected in epithelial lines, monocytes, muscle, and placenta. Although it has no sequence identity with IL-2, IL-15 shares many of the T cell-stimulatory activities described for IL-2. We have examined IL-15 for its ability to stimulate B cells and have compared its activity with that of IL-2. IL-15 costimulates proliferation of B cells activated with immobilized anti-human IgM or phorbol ester, but has no stimulatory effect on resting B cells. In combination with recombinant CD40L, IL-15 is a potent inducer of polyclonal IgM, IgG1, and IgA secretion, but does not cause production of IgG4 or IgE. The activity of IL-15 in B cell proliferation and differentiation assays is comparable with that of IL-2. Studies that used neutralizing Abs have demonstrated that, for signal transduction in B cells, IL-15 uses the beta-chain of the IL-2R complex but, unlike IL-2, does not require the alpha-chain. IL-2 is required for the generation of a human primary Ag-specific in vitro response using sheep erythrocytes as Ag. Of all cytokines examined, only IL-15 has the capacity to replace IL-2 in this system, although only partially. In summary, IL-15 has comparable activity with IL-2 for the induction of B cell proliferation and differentiation and uses at least some of the components of the IL-2R complex to mediate its effects.


Subject(s)
B-Lymphocytes/immunology , Cell Differentiation/immunology , Interleukins/physiology , Lymphocyte Activation/immunology , Antibody Formation/immunology , Cells, Cultured , Humans , Interleukin-15 , Interleukin-2/physiology
8.
Eur J Immunol ; 23(9): 2326-31, 1993 Sep.
Article in English | MEDLINE | ID: mdl-7690330

ABSTRACT

CD40 ligand (CD40L) is a 33-kDa type II membrane glycoprotein induced on T cells upon activation. CD40L has previously been shown to induce proliferation of resting B cells, immunoglobulin (Ig) secretion from B cells cultured with cytokines and cytokine secretion and tumoricidal activity from monocytes. In this report CD40L is shown to be stimulatory for human T cells, inducing CD25 (p55 IL-2R) and CD40L expression on resting peripheral blood T cells, enhanced expression of these molecules and CD69 on CD3-activated cells and secretion of interferon-gamma, tumor necrosis factor-alpha and interleukin (IL)-2 from T cells cultured in the presence of a sub-mitogenic concentration of phytohemagglutinin A (PHA). Furthermore, stimulation with CD40L induces proliferation of CD3- or PHA-activated T cells of blood, tonsillar or thymic origin. A similar proliferative response is observed with CD4+ and CD8+ T cells and this effect is largely IL-2 independent. A soluble construct of the extracellular domain of the CD40L has similar activity to that of membrane-expressed ligand in the induction of T cell surface antigens and proliferation. The results presented here taken together with the various activities ascribed for CD40L on B cells and monocytes demonstrate that CD40L has pleiotropic biological activity for cells of the hemopoietic lineage.


Subject(s)
Interleukin-2/pharmacology , Lymphocyte Activation/drug effects , Membrane Glycoproteins/pharmacology , T-Lymphocytes/drug effects , Antibodies, Monoclonal/immunology , Antigens, CD/physiology , Antigens, Differentiation, B-Lymphocyte/physiology , CD3 Complex/immunology , CD40 Antigens , CD40 Ligand , Cells, Cultured , Humans , Interleukin-2/biosynthesis , Ligands
9.
Blood ; 81(11): 2998-3005, 1993 Jun 01.
Article in English | MEDLINE | ID: mdl-8499634

ABSTRACT

Biotinylated interleukin-4 (IL-4) was used to examine IL-4 receptor (IL-4R) expression on a range of human B-cell lines by flow cytometry. Using high concentrations of biotinylated IL-4, we have identified a novel low-affinity IL-4 receptor expressed at high levels on pre-B lines. Expression of this low-affinity receptor did not correlate with detected mRNA levels for the previously cloned receptor or with reactivity of two anti-human IL-4R monoclonal antibodies (MoAb). Radiolabeled IL-4 cross-linking studies using pre-B lines showed a doublet of 65 to 75 Kd in contrast to the 110- to 130-Kd molecule detected on cells expressing the cloned IL-4R. A soluble IL-4 binding protein (IL-4bp) was purified from the supernatants of three pre-B lines expressing the low-affinity receptor on their surface. IL-4bp could block both IL-4-mediated CD23 induction on tonsil B cells and IL-4-induced inhibition of proliferation of the pre-B line JM1. Partial N-terminal amino acid sequence was obtained from purified IL-4bp that confirmed this protein to be novel. A 12 amino acid peptide based on the IL-4bp sequence was used to produce a polyclonal antiserum that was reactive with purified IL-4bp, and also bound to the surface of pre-B cells but not to murine CTLL cells transfected with the human IL-4R. Blocking MoAb against the previously characterized high-affinity receptor inhibited IL-4-mediated proliferation of hIL-4R+ CTLL cells but had no effect on IL-4-induced inhibition of JM1 cell proliferation, and only partially inhibited IL-4-mediated CD23 and sIgM induction and proliferation of tonsil B cells. The data presented here provide evidence for a novel cell-surface expressed low-affinity IL-4R that also exists as a biologically active soluble IL-4 binding protein.


Subject(s)
B-Lymphocytes/chemistry , Interleukin-4/metabolism , Receptors, Mitogen/chemistry , Amino Acid Sequence , Antibodies, Monoclonal/immunology , Cross-Linking Reagents , Gene Expression , Humans , Molecular Sequence Data , RNA, Messenger/genetics , Receptors, Interleukin-4 , Receptors, Mitogen/immunology , Solubility
10.
J Immunol ; 150(9): 3671-80, 1993 May 01.
Article in English | MEDLINE | ID: mdl-8097223

ABSTRACT

Recombinant human CD40 ligand (hCD40L) was expressed on the surface of CV1/EBNA cells and examined for its ability to induce proliferation and Ig secretion from human B cells in the presence or absence of soluble cytokines. hCD40L was directly mitogenic in a dose-dependent fashion for purified tonsil B cells with maximal proliferation occurring at days 5 to 7. Proliferation induced by CD40L was significantly enhanced in the presence of IL-2, IL-4, or IL-10 and strongly suppressed by transforming growth factor-beta. Although IL-5, TNF-alpha, and IFN-gamma had no stimulatory effect in the presence of hCD40L alone, if IL-4 was also present in cultures, these cytokines enhanced the proliferative response above that seen with IL-4 alone. Interestingly, in the absence of IL-4, IFN gamma had an inhibitory effect on hCD40L-induced proliferation. Although CD40L alone did not enhance Ig secretion, addition of IL-2 or IL-10 to the cultures significantly elevated the levels of IgM, IgG1, and IgA that were observed. Addition of IL-4 to the cultures did not enhance secretion of these isotypes but had a weak inhibitory effect. However, CD40L-mediated induction of IgG4 and IgE was dependent on the presence of IL-4. Of the cytokines examined, only IL-10 enhanced IgE secretion under these conditions. Although transforming growth factor-beta only partially inhibited secretion of IgM, IgG1, and IgA, it was strongly suppressive for IgG4 and IgE production. Our data demonstrate that proliferation and Ig secretion induced in the presence of CD40L can be modulated in a positive and negative fashion by soluble cytokines. IL-2 and IL-10 specifically enhance IgM, IgG1, and IgA production although IL-4, despite costimulating B cell proliferation, does not augment secretion of these isotypes but provided an essential cosignal with CD40L for the production of IgG4 and IgE.


Subject(s)
B-Lymphocytes/immunology , Cytokines/pharmacology , Immunoglobulins/biosynthesis , Lymphocyte Activation/drug effects , Membrane Glycoproteins/pharmacology , Antigens, CD/pharmacology , B-Lymphocytes/drug effects , CD40 Ligand , Cells, Cultured , Humans , Interferon-gamma/pharmacology , Interleukin-10/pharmacology , Interleukin-4/pharmacology , Ligands , Membrane Glycoproteins/immunology , Recombinant Proteins/pharmacology , Transforming Growth Factor beta/pharmacology
11.
J Exp Med ; 176(6): 1543-50, 1992 Dec 01.
Article in English | MEDLINE | ID: mdl-1281209

ABSTRACT

Signaling through the cell surface molecule, CD40, is known to play an important role in the proliferation and differentiation of B lymphocytes. Using the thymoma cell line EL4, we recently identified and cloned a cDNA encoding a murine ligand for the CD40 molecule (mCD40-L) and showed that it has biological activity in vitro. A cDNA encoding a human homologue of the mCD40-L was isolated using crosshybridization techniques from an activated peripheral blood T cell library. The predicted amino acid sequence indicates that this human ligand for CD40 (hCD40-L) is a 261 amino acid type II membrane protein that exhibits 78% amino acid identity with its murine counterpart. Northern blot and FACS analyses suggest that the hCD40-L is restricted in its expression to T lymphocytes, and that it is most abundant on the CD4+ T cell subpopulation. Cells transfected with hCD40-L caused the proliferation of human tonsil B cells in the absence of costimuli and, in the presence of interleukin 4, induced immunoglobulin E secretion from purified human B cells. A comparison of the efficacy of the hCD40-L and mCD40-L in these assays is presented.


Subject(s)
Antigens, CD/physiology , Antigens, Differentiation, B-Lymphocyte/physiology , B-Lymphocytes/immunology , Immunoglobulin E/metabolism , Lymphocyte Activation , Membrane Glycoproteins/physiology , Amino Acid Sequence , Animals , Antigens, CD/metabolism , Base Sequence , CD40 Antigens , CD40 Ligand , Cells, Cultured , Cloning, Molecular , Humans , Immunoglobulin E/biosynthesis , Interleukin-4/pharmacology , Kinetics , Lymphoma , Membrane Glycoproteins/genetics , Mice , Molecular Sequence Data , Recombinant Proteins/pharmacology , Sequence Homology, Amino Acid , T-Lymphocytes/immunology , Tumor Cells, Cultured
12.
Cytokine ; 4(6): 461-9, 1992 Nov.
Article in English | MEDLINE | ID: mdl-1292630

ABSTRACT

The induction of cytokine secretion by human peripheral blood (PB) T cells was examined. Highly purified T cells stimulated with interleukin 7 (IL-7), in the absence of co-mitogen, secreted IL-2, IL-4, IL-6 and interferon gamma (IFN-gamma) upon restimulation with phorbol ester and ionomycin. In contrast, induction of T-cell cultures initiated with IL-2 or IL-4 yielded only low levels of IL-6 and virtually undetectable levels of IL-4 or IFN-gamma, while IL-2 secretion was reduced. No difference was seen in the ability of CD4+ and CD8+ subpopulations, grown in IL-7, to produce cytokines. In contrast, subdivision of T cells into memory and naive populations using the CD45RO monoclonal antibody (mAb) UCHL1, revealed that almost all of the potential to secrete IL-4 and IL-6 in response to IL-7 resided in the CD45RO+ memory population. Stimulation of cytokine-secreting cells appeared to be a direct effect of IL-7 as neutralizing antibodies directed against IL-2 and IL-4 had no effect on the levels of cytokines produced. The differences observed in the ability of IL-2, IL-4, and IL-7 to potentiate cytokine production was supported by measurement of cytokine mRNA levels by PCR. The elevated levels of cytokine secretion seen in cells cultured with IL-7 was not due simply to increased viability in these cultures compared with those containing IL-2 or IL-4, as these populations showed comparable cloning frequencies in phytohemagglutinin (PHA) + IL-2. These results demonstrate that IL-7, in the absence of co-mitogen, is a potent initial stimulus for multiple cytokine production by human T cells upon restimulation.


Subject(s)
Cytokines/metabolism , Interleukin-7/pharmacology , T-Lymphocytes/immunology , Cytokines/genetics , DNA/genetics , DNA Probes , Gene Expression , Humans , In Vitro Techniques , Interferon-gamma/metabolism , Interleukin-2/metabolism , Interleukin-2/pharmacology , Interleukin-4/metabolism , Interleukin-4/pharmacology , Interleukin-6/metabolism , Kinetics , Molecular Sequence Data , T-Lymphocyte Subsets/drug effects , T-Lymphocyte Subsets/immunology , T-Lymphocyte Subsets/metabolism , T-Lymphocytes/drug effects , T-Lymphocytes/metabolism
13.
Eur J Immunol ; 22(8): 2071-6, 1992 Aug.
Article in English | MEDLINE | ID: mdl-1379186

ABSTRACT

We have identified the murine thymoma line EL4 as a source of biologically active CD40 ligand. Using a biotin-labeled soluble CD40.Fc fusion protein, consisting of the extracellular domain of human CD40 and the Fc region of human IgG1, EL4 cells were subjected to repeated flow cytometric cell sorting to select for cells with enhanced biotinylated CD40.Fc binding. After nine rounds of sorting, the number of CD40.Fc binding sites/cell had risen from 450 on the unsorted parental EL4 cells to 15,000 on EL40.9 cells (EL4 cells sorted with biotinylated CD40.Fc for nine rounds). Scatchard analysis of radiolabeled CD40.Fc binding revealed that the surface-expressed CD40 ligand on parental EL4 and EL40.9 cells bound its receptor with a single class of high-affinity sites (Kd = 0.5 nM). Supernatant (SN) from the sorted EL40.9 cells was found to contain human and murine B cell stimulatory activity which could be removed by preclearing with immobilized CD40.Fc, confirming the presence of soluble CD40 ligand in the preparations. EL40.9 supernatant enhanced soluble CD23 (sCD23) release and induced IgE secretion from interleukin 4-stimulated human B cells. In addition, EL40.9 SN contained proliferative activity for anti-IgM-activated murine B cells which could be removed by treatment with immobilized CD40.Fc. However, the same SN had no demonstrable activity on the proliferation of human B cells. The results presented here describe, for the first time, a source of membrane-bound and soluble CD40 ligand. The soluble form of this murine ligand has activity on murine and human B cells and induces some of the functional responses predicted for the ligand based on the action of stimulatory antibodies directed against the human CD40 surface molecule.


Subject(s)
Antigens, CD/metabolism , Antigens, Differentiation, B-Lymphocyte/metabolism , Animals , Antigens, Differentiation, B-Lymphocyte/biosynthesis , B-Lymphocytes/immunology , CD40 Antigens , Flow Cytometry , Humans , Immunoglobulin E/metabolism , Interleukin-4/pharmacology , Ligands , Lymphocyte Activation , Mice , Receptors, Fc/biosynthesis , Receptors, IgE
14.
Nature ; 357(6373): 80-2, 1992 May 07.
Article in English | MEDLINE | ID: mdl-1374165

ABSTRACT

The CD40 surface molecule is a 277-amino-acid glycoprotein expressed on B lymphocytes, epithelial cells and some carcinoma cell lines. Monoclonal antibodies against CD40 mediate a variety of effects on B lymphocytes, including induction of intercellular adhesion, short- and long-term proliferation, differentiation and enhanced tyrosine phosphorylation of proteins. In addition, germinal centre centrocytes are prevented from undergoing apoptosis by activation through CD40 and receptor for antigen. These data indicate that CD40 could be a receptor for an unknown ligand with important functions in B-cell development and activation. This hypothesis is strengthened by the homology of the extracellular region of the CD40 molecule with a family of cell-surface glycoproteins that includes the receptors for nerve growth factor and tumour necrosis factor. Here we report the cloning of a ligand for CD40 that is expressed on the cell surface of activated T cells and mediates B-cell proliferation in the absence of co-stimulus, as well as IgE production in the presence of interleukin-4.


Subject(s)
Antigens, CD/metabolism , Antigens, Differentiation, B-Lymphocyte/metabolism , Membrane Glycoproteins/genetics , Amino Acid Sequence , Animals , B-Lymphocytes/metabolism , Base Sequence , CD40 Antigens , CD40 Ligand , Humans , Immunoglobulin E/metabolism , Ligands , Membrane Glycoproteins/physiology , Mice , Molecular Sequence Data , Protein Binding , T-Lymphocytes/metabolism , Tumor Cells, Cultured
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