bluuebunny/crossref_metadata_2025 huggingface dataset:
HLA-DR alleles differ in their ability to present staphylococcal enterotoxins to T cells.
Authors: Herman A , Croteau G , Sekaly R P , Kappler J , Marrack P
Published: June 2004

Staphylococcal enterotoxins (SEs) have been shown to bind to major histocompatibility complex (MHC) class II proteins and stimulate T cells in a V beta-specific manner, and these V beta specificities for various SEs have been well documented in mice and humans. This study was undertaken in order to examine the ability of human class II molecules to present SEs to human and murine T cell hybridomas. Using a panel of transfectants expressing individual HLA class II antigens, we have shown that HLA-DR alleles differ in their ability to bind and present SEs. Since the HLA-DR proteins share a common alpha chain, these results indicate that the polymorphic beta chain plays an important role in SE binding and presentation to T cells. In addition, we have shown that human class II isotypes markedly differ in their ability to present SEs. The results of this study should provide information on the region of MHC class II molecules that interacts with foreign, and perhaps self, super-antigens.
Expression of human adenosine deaminase in mice transplanted with hemopoietic stem cells infected with amphotropic retroviruses.
Authors: van Beusechem V W , Kukler A , Einerhand M P , Bakx T A , van der Eb A J , van Bekkum D W , Valerio D
Published: June 2004

Amphotropic recombinant retroviruses were generated carrying sequences encoding human adenosine deaminase (ADA). Transcription of the human ADA gene was under control of a hybrid long terminal repeat in which the enhancer from the Moloney murine leukemia virus was replaced by an enhancer from the F101 host-range mutant of polyoma virus. Hemopoietic stem cells in murine bone marrow were infected with this virus under defined culture conditions. As a result, 59% of day-12 colony forming unit spleen (CFU-S) stem cells became infected without any in vitro selection. Infected CFU-S were shown to express human ADA before transplantation and this expression sustained upon in vivo maturation. Mice transplanted with infected bone marrow exhibited human ADA expression in lymphoid, myeloid, and erythroid cell types. Moreover, human ADA expression persisted in secondary and tertiary transplanted recipients showing that human ADA-expressing cells were derived from pluripotent stem cells. These characteristics of our amphotropic viruses make them promising tools in gene therapy protocols for the treatment of severe combined immunodeficiency caused by ADA deficiency. In this respect it is also relevant that the viral vector that served as backbone for the ADA vector was previously shown to be nonleukemogenic.
The pili of Aeromonas hydrophila: identification of an environmentally regulated "mini pilin".
Authors: Ho A S , Mietzner T A , Smith A J , Schoolnik G K
Published: June 2004

Ultrastructural studies of Aeromonas hydrophila strain AH26 revealed two distinctive pilus types: "straight" pili appear as brittle, rod-like filaments, whereas "flexible" pili are supple and curvilinear. Straight pili are produced constitutively under all tested conditions of growth. In contrast, the expression of flexible pili is regulated by physical and chemical variables, being produced at 22 vs. 37 degrees C, in a liquid vs. a solid medium, and when the availability of free-iron is reduced by the presence of deferoxamine mesylate. Both pilus proteins were purified and biochemically and functionally characterized. The major repeating subunit of the straight pilus is a 17,000-mol wt polypeptide with amino acid sequence homology with Escherichia coli type 1 and Pap pili. The flexible pilus filament is a homopolymer composed of a novel 46 amino acid polypeptide. Resistance of the flexible pilus filament to disaggregation using various chemical treatments was demonstrated; its stability as a polymer and its apparent mechanical strength seem to be conferred by a 20 amino acid hydrophobic, COOH-terminal domain. Purified straight pili lack hemagglutinating function. In contrast, purified flexible pili cause the agglutinin of human, guinea pig, ovine, bovine, and avian erythrocytes, although this property could only be demonstrated in the presence of divalent cations and was most evident at 4 vs. 22 degrees C. Taken together, these results suggest that the pathogenic and ecological roles of the flexible pilus are related to this species' existence as a free-living organism in aquatic environments and its ability to cause infections, both in cold-blooded vertebrates and the human intestine.
Characterization of the ligand(s) responsible for negative selection of V beta 11- and V beta 12-expressing T cells: effects of a new Mls determinant.
Authors: Vacchio M S , Ryan J J , Hodes R J
Published: June 2004

During T cell development, events occur that result in the generation of a T cell population capable of recognizing foreign antigens in association with self major histocompatibility complex (MHC) gene products. However, selective events also occur during thymic education that result in the deletion of T cells expressing alpha/beta T cell receptors with high affinity for self determinants alone, i.e., potentially self-reactive T cells. Both MHC- and non-MHC-encoded self antigens appear to play critical roles in this negative selection of self-reactive T cells. We recently observed that T cells expressing V beta 5, V beta 11, V beta 12, or V beta 16 products are deleted in most strains of H-2k type, but not in congenic H-2b strains. In contrast, the H-2k strain C58/J deleted V beta 5+ and V beta 16+ T cells, but failed to delete T cells expressing V beta 11 or V beta 12. Based upon this observation, in the present study we have analyzed the genetic regulation of the ligands responsible for deletion of V beta 11- and V beta 12-expressing T cells, and have tested the possibility that these ligands can function as strong alloantigens analogous to the known minor lymphocyte stimulatory (Mls)- and MHC-encoded antigens. Two major findings have resulted from these studies. First, the ligands recognized by V beta 11+ and V beta 12+ T cells were regulated by both MHC- and multiple non-MHC-encoded genes. Correlation between expression of these two V beta s in backcross animals suggested that shared, though not necessarily identical, ligands mediate deletion of V beta 11- and V beta 12-expressing T cells. Second, the ligand for deletion of V beta 11- and V beta 12-expressing T cells functions as a newly defined Mls alloantigen that stimulates primary proliferative responses in T cell populations from mice that express V beta 11+ and V beta 12+ T cells.
A single pre-B cell can give rise to antigen-specific B cells that utilize distinct immunoglobulin gene rearrangements.
Authors: Caton A J
Published: June 2004

A group of hybridomas that express antibodies with related specificities for the influenza virus hemagglutinin (HA), that represent B cells that were the clonal progeny of a single pre-B cell, and that utilized distinct L chain gene rearrangements have been characterized. The clonal relationship was established by the sharing of H chain gene rearrangements at both the productive and the nonproductive alleles. Among these hybridomas, one group had rearranged only one of its kappa alleles, having joined a V kappa 24 gene to the J kappa 2 gene segment. The other group utilized the same V kappa 24 gene segment in productive rearrangement to the J kappa 5 gene segment, and shared an aberrant rearrangements among members of the same B cell clone can normally occur, and can contribute to the generation and diversification of the immune repertoire that is available for the recognition of foreign antigens. Mechanisms by which the distinct rearrangements expressed by the hybridomas might have been generated are discussed.
Natural autoantibodies to thymocytes: origin, VH genes, fine specificities, and the role of Thy-1 glycoprotein.
Authors: Hayakawa K , Carmack C E , Hyman R , Hardy R R
Published: June 2004

15 SM/J mouse hybridoma antibodies that show antithymocyte autoantibody (ATA) activity by immunofluorescence staining were studied. Half of these antibodies react with determinants whose expression is associated with Thy-1, as shown by blocking experiments with anti-Thy-1 and loss of reactivity with Thy-1- mutant cell lines. The Thy-1 dependence of three of these ATA is further confirmed by their reexpression on a Thy-1 gene transfectant. However, the remaining antibodies exhibited binding that showed little or no dependence on Thy-1. Furthermore, we find that most ATA derives from the Ly-1 B subpopulation, as demonstrated by lipopolysaccharide-induced ATA secretion in vitro and by comparison of ATA hybridoma frequencies. VH region gene sequence data of 14 monoclonal ATA from Ly-1 B cell-derived hybridomas reveal the utilization of nine VH genes belonging to four different VH families (J558, 3609, Q52, and Vgam3.8). While we find that two of these hybridomas arose from a clonal expansion, we also find four examples of a 3609 family VH gene utilized in clonally independent lines showing similar specificity. Yet another example of identical VH gene usage by clonally unrelated cells is found in two J558 ATA of a distinct fine specificity. These data suggest that the enrichment of ATA B cells in the Ly-1 B subset is primarily due to repeated independent recruitment of B cells by antigen resulting in the expression of a restricted set of VH genes.
Characterization of an immuno-dominant variable surface antigen from pathogenic and nonpathogenic Entamoeba histolytica.
Authors: Edman U , Meraz M A , Rausser S , Agabian N , Meza I
Published: June 2004

A 125-kD surface antigen of Entamoeba histolytica is recognized by 73% of immune sera from patients with amoebic liver abscesses. Using pooled human immune sera a cDNA clone (lambda cM17) encoding this antigen (M17) has been isolated from a lambda gt11 expression library of the virulent stain E. histolytica HM1:IMSS. Monospecific antibodies, purified by binding to phage lysate of lambda cM17, and mAb FA7 reacted exclusively with the 125-kD antigen by Western blot analysis. Surface binding and cap formation are observed with patient sera, purified monospecific antiserum, and mAb FA7. Corresponding genomic clones (pBSgM17-1/2/3) were isolated by hybridization with the cDNA clone. These contained an open-reading frame of 3345 bp, which is in good agreement with the mRNA size of approximately 3.0 kb as revealed by Northern hybridization with lambda cM17. The inferred amino acid sequence predicts a 125,513 dalton protein that contains 17 potential N-linked glycosylation sites and is unusually rich in tyrosine and asparagine residues. A distinctly hydrophobic NH2-terminal region may serve as membrane anchor or signal sequence. In contrast to conservation of an immunodominant epitope recognized in pathogenic and nonpathogenic strains by monoclonal FA7 and human immune sera, amplification and sequence analysis of a 1,4000-bp fragment of this gene from a fresh nonpathogenic isolate by use of the PCR demonstrate regions of significant sequence divergence in this antigen. A 1% sequence variability among different isolates of the pathogenic strain HM1:IMSS and a 12-13% variability between pathogenic and nonpathogenic strains are revealed by comparison to published partial amino acid sequences (Tannich, E., R.D. Horstmann, J. Knobloch, and H.H. Arnold. 1989. Proc. Natl. Acad. Sci. USA. 86:5118). Some restriction enzymes were found that allowed PCR diagnosis of nonpathogenic and pathogenic isolates with the exclusion of E. histolytica-like Laredo, suggesting that a detailed study of nonpathogenic and pathogenic isolates in relation to the M17 antigen sequence will provide a basis of differentiating isolates.
Generation of interleukin 4 (IL-4)-producing cells in vivo and in vitro: IL-2 and IL-4 are required for in vitro generation of IL-4-producing cells.
Authors: Le Gros G , Ben-Sasson S Z , Seder R , Finkelman F D , Paul W E
Published: June 2004

T cell populations derived from naive mice produce very small amounts of interleukin 4 (IL-4) in response to stimulation on anti-CD3-coated dishes. IL-4 production by such cells is mainly found among large- and intermediate-sized T cells and is dependent upon IL-2. Injection of anti-IgD into mice, a stimulus that leads to striking increases in serum levels of IgG1 and IgE, causes a striking increase in the IL-4-producing capacity of T cells. This increase is first observed 4 d after injection of anti-IgD. IL-4 production by T cells from anti-IgD-injected donors is mainly found among large- and intermediate-sized T cells. Small, dense T cells are poor producers of IL-4. The capacity of T cells from anti-IgD-injected donors to produce IL-4 is enhanced by addition of IL-2 and is largely, but not completely, inhibited by neutralization of in situ produced IL-2. These results indicate that the control of IL-4 production in T cells from naive and anti-IgD-injected donors is similar. However, it is possible that a portion of the IL-4-producing activity of T cells from activated donors is IL-2 independent. Although small T cells from naive donors have a very limited capacity to produce IL-4 in response to stimulation with anti-CD3, even in the presence of added IL-2, they can give rise to IL-4-producing cells upon in vitro culture on plates coated with anti-CD3 if both IL-2 and IL-4 are added. This leads to the appearance of IL-4-producing cells within 2 d. When analyzed after 5 d of culture by harvesting and re-exposure to anti-CD3-coated culture wells and IL-2, these cells have increased their IL-4-producing capacity by approximately 100-fold. The development of IL-4-producing cells in response to anti-CD3, IL-2, and IL-4 is not inhibited by interferon gamma (IFN-gamma), nor does IFN-gamma diminish IL-4 production by these cells upon challenge with anti-CD3 plus IL-2.
Apparent lack of MHC restriction in binding of class I HLA molecules to solid-phase peptides.
Authors: Chen B P , Rothbard J , Parham P
Published: June 2004

The specificity of binding of solubilized, purified HLA-A,B molecules to solid-phase peptides has been examined using the assay described by Bouillet et al. [1989. Nature (Lond.). 339:473.] 64 peptides derived from the sequences of viral antigens, HLA-A,B,C heavy chains, and clathrin light chains were tested for binding to HLA-A2.1, Aw68.1, Aw69, B44, and B5, molecules that differ by 5-17 residues of the peptide binding groove. 15 of the peptides, including those known to be T cell epitopes, gave significant binding. The pattern of peptide binding for each of the five HLA-A,B molecules was not significantly different. Binding was demonstrated to be a property of native beta 2m-associated HLA-A,B molecules that preserved conformational antigenic determinants after binding to peptide. In comparison to our previous results from solution-based assays the proportion of HLA-A,B molecules that can bind solid-phase peptides is very high. This accessibility of solid-phase peptides to the binding site of MHC molecules may be directly related to the observed absence of MHC specificity in the binding.
Selective expansion of T cells expressing V beta 2 in toxic shock syndrome.
Authors: Choi Y , Lafferty J A , Clements J R , Todd J K , Gelfand E W , Kappler J , Marrack P , Kotzin B L
Published: June 2004

Infection with Staphylococcus aureus and the production of toxic shock syndrome toxin-1 (TSST-1) have been implicated in the pathogenesis of toxic shock syndrome. Previous in vitro studies have demonstrated that TSST-1 is a powerful but selective stimulator of human T cells, and that the majority of activated cells express the TCR V beta 2 gene segment. We therefore studied patients with toxic shock syndrome using a modification of the PCR to determine if expansion of V beta 2+ T cells is a marker of the in vivo disease process. Five of eight patients studied demonstrated markedly elevated levels of circulating V beta 2+ T cells, whereas none showed significantly elevated levels of T cells expressing other V beta gene segments. The results suggest that toxin-mediated T cell activation, which involves a large fraction of the human T cell repertoire, may be critical in the pathogenesis of this disease.
Intravascular filarial parasites elaborate cyclooxygenase-derived eicosanoids.
Authors: Liu L X , Serhan C N , Weller P F
Published: June 2004

The nematode parasites that cause human lymphatic filariasis survive for long periods in their vascular habitats despite continual exposure to host cells. Since prostanoids formed from arachidonic acid can modulate interactions among platelets, leukocytes, and endothelial cells, we examined whether intravascular nematode parasites can elaborate prostanoids. Microfilariae of Brugia malayi utilize exogenous and endogenous arachidonic acid to generate and release two predominant prostanoids, prostacyclin and prostaglandin E2. Filarial metabolism of host fatty acids to form these vasodilatory, antiaggregatory, and immunomodulatory eicosanoids provides a means by which these helminthic parasites may influence host immune and other cellular responses.
Identification of a 60-kD tumor necrosis factor (TNF) receptor as the major signal transducing component in TNF responses.
Authors: Thoma B , Grell M , Pfizenmaier K , Scheurich P
Published: June 2004

We describe here a monoclonal antibody (H398) that immunoprecipitates a human 60-kD tumor necrosis factor (TNF) membrane receptor (p60) and competes with TNF binding to p60 but not to p85 TNF receptors. Despite partial inhibition of TNF binding capacity of cells coexpressing both TNF receptor molecules, H398 uniformly and completely inhibits very distinct TNF responses on a variety of cell lines. These data suggest a limited structural heterogeneity in those components actually contributing to TNF responsiveness and identify p60 as a common receptor molecule essential for TNF signal transduction. As H398 is a highly effective TNF antagonist in vitro, it might be useful as a therapeutic agent in the treatment of TNF-mediated acute toxicity.
The autoantigen Ku is indistinguishable from NF IV, a protein forming multimeric protein-DNA complexes.
Authors: Stuiver M H , Coenjaerts F E , van der Vliet P C
Published: June 2004

We have isolated a cDNA encoding the 84-kD subunit of NFIV. Tryptic peptide sequences were identified within the coding sequences, confirming its proper identity. The primary sequence of the protein is identical to that of the large subunit of the Ku autoantigen. A missing NFIV peptide sequence was identified within the sequence of the small subunit of Ku. In addition, the proteins are identical in immunological aspects. We suggest that the Ku and NFIV proteins are identical. This connection adds new biochemical data to our knowledge of the Ku autoantigen.
In vivo induction of anergy in peripheral V beta 8+ T cells by staphylococcal enterotoxin B.
Authors: Rellahan B L , Jones L A , Kruisbeek A M , Fry A M , Matis L A
Published: June 2004

We have developed a model of peripheral in vivo T cell tolerance that is induced by administration of the protein superantigen staphylococcal enterotoxin B (SEB). Rather than activating V beta 8+ T cells, in vivo administration of SEB induced in them a profound state of anergy. This was shown by their failure to proliferate to subsequent in vitro restimulation with SEB or to anti-V beta 8 antibodies. This unresponsiveness was V beta 8 specific since T cells from SEB-immunized mice responded normally to other antigens. 8 d after SEB administration, there was no reduction in the number of V beta 8+ T cells or in the intensity of V beta 8 T cell receptor (TCR) expression. Although a portion of the V beta 8+ T cells from SEB-primed mice were able to express interleukin 2 receptors (IL-2Rs), they failed to proliferate in response to exogenous IL-2, indicating they were defective in their IL-2 responsiveness. 2-4 wk after SEB administration, there was a reduction of approximately 50% in the number of V beta 8+ cells in immunized compared with control animals. There was, however, no reduction in the level of TCR expression on the remaining V beta 8+ cells. These data demonstrate that proteins that activate T cells in vitro in a V beta-specific manner can induce a state of anergy in peripheral T cells in vivo and may possibly further mediate clonal deletion in a portion of the tolerized cells.
Increased expression of the interleukin 2 (IL-2) receptor beta chain (p70) on CD56+ natural killer cells after in vivo IL-2 therapy: p70 expression does not alone predict the level of intermediate affinity IL-2 binding.
Authors: Voss S D , Robb R J , Weil-Hillman G , Hank J A , Sugamura K , Tsudo M , Sondel P M
Published: June 2004

The expression of the 70-kD beta subunit of the interleukin 2 receptor (IL-2R) has been examined on peripheral blood lymphocytes (PBL) obtained from patients receiving systemic infusions of IL-2. Using monoclonal antibodies directed against p70, flow cytometric analyses revealed a greater than threefold increase in expression of the IL-2R beta chain on CD56+ natural killer (NK) cells from post-IL-2 therapy PBL relative to pre-therapy cells. The level of p70 expression on the post-therapy cells was three- to fourfold greater (based on fluorescence intensity) than the level of p70 expression on YT cells, an NK-like cell line that expresses approximately 12,000 intermediate affinity IL-2 binding sites/cell. Despite the high level of p70 expression, in 125I-IL-2 binding assays only 790-1,290 intermediate affinity IL-2 binding sites/cell were detected on post-therapy cells from six patients. These data represent the first report of increased p70 expression after in vivo IL-2 administration and suggest a requirement for at least one additional subunit for the formation of functional intermediate affinity IL-2Rs. Furthermore, the presence on the surface of post-therapy NK cells of excess p70 that does not bind IL-2 with intermediate affinity implies that the formation of intermediate affinity IL-2Rs is not solely determined by the level of p70 expression, and that the response of NK cells to IL-2 might be regulated by altering the expression of p70 or some other IL-2R subunit.
Identification and characterization of receptors specific for human pancreatic secretory trypsin inhibitor.
Authors: Niinobu T , Ogawa M , Murata A , Nishijima J , Mori T
Published: June 2004

Specific binding sites for human pancreatic secretory trypsin inhibitor (PSTI) on 3T3 Swiss albino cells were studied using radioiodinated recombinant PSTI. Some ion species, pH, and temperature significantly influenced the binding of 125I-PSTI. Kinetic studies showed that the binding of 125I-PSTI to 3T3 Swiss albino cells reached the maximum level within 120 min at 4 degrees C, with a slow dissociation rate. The half-maximal inhibition (ID50) of 125I-PSTI binding by unlabeled PSTI occurred at 1.0 x 10(-10) M. On Scatchard analysis of the competitive binding data, linear plots indicated a single class of receptors with high affinity (Kd = 5.3 x 10(-10) M) on 3T3 Swiss albino cells, the number of receptors being 5,400 per cell. Treatment of surface-bound radiolabeled PSTI with a chemical crosslinker (disuccinimidyl suberate) led to the identification of a membrane polypeptide of Mr 140,000 to which PSTI was crosslinked. The formation was inhibited by an excess amount of unlabeled PSTI in a dose-dependent manner. The binding of 125I-PSTI to 3T3 Swiss albino cells was competitively inhibited by unlabeled PSTI but not by other peptide hormones, such as epidermal growth factor (EGF), bovine fibroblast growth factor, insulin-like growth factor, transforming growth factor alpha, platelet-derived growth factor, and tumor necrosis factor, indicating the presence of receptors specific for PSTI. Various protease inhibitors had no or only a little effect, and mercaptoethanol and dithiothreitol strongly decreased the binding of 125I-PSTI. Incubation at 37 degrees C resulted in rapid internalization of cell-bound 125I-PSTI, followed by the appearance of trichloroacetic acid-soluble 125I-radioactivity in the culture medium, due to degradation of internalized PSTI. In addition, PSTI stimulated [3H]thymidine incorporation into DNA on 3T3 Swiss albino cells in a dose-dependent manner. The combined addition of PSTI and EGF stimulated [3H]thymidine incorporation to an extent greater than that seen with either agent alone. These results indicated that the biological effect of PSTI was mediated by high affinity plasma membrane receptors, which were not a cell-surface proteinase(s). Specific binding of 125I-PSTI was noted with the following cells: WI-38, 3T3 Swiss albino, HUVE, BDC-1, and H4-II-E-C3.
Antibody raised against soluble CD4-rgp120 complex recognizes the CD4 moiety and blocks membrane fusion without inhibiting CD4-gp120 binding.
Authors: Celada F , Cambiaggi C , Maccari J , Burastero S , Gregory T , Patzer E , Porter J , McDanal C , Matthews T
Published: June 2004

We studied the humoral response of mice immunized with soluble CD4-rgp120 complex, testing polyclonal and monoclonal antibodies (mAbs) with the aim of identifying molecular changes that take place after the first interaction between human immunodeficiency virus and the cell surface. The antisera had a paradoxically high syncytia-blocking titer associated with anti-CD4 specificity, while their capacity to inhibit CD4-gp120 binding was relatively modest. One of the mAbs produced from these responders blocks syncytia formation but does not inhibit CD4 interaction with gp120. Apparently, this mAb interacts with the CD4 moiety of CD4-gp120 complex and prevents a post-binding event necessary for membrane fusion and viral infection.
A cDNA clone expressed in natural killer and T cells that likely encodes a secreted protein.
Authors: Yabe T , McSherry C , Bach F H , Houchins J P
Published: June 2004

We have isolated a series of cross-hybridizing cDNA clones, as a group designated as NKG5, from a human natural killer (NK) cell clone cDNA library. These clones show a high degree of homology with a previously described gene, 519, which was thought to be T cell specific. A comparison of the full-length cDNA sequence of NKG5 and the published sequence of 519 shows that NKG5 lacks a 242-base segment that is found in 519 and that this deletion leads to the use of a different putative translational start codon. Unlike 519, the predicted NKG5 polypeptide has an NH2-terminal sequence that is strongly hydrophobic, characteristic of a signal peptide, and lacks any additional hydrophobic regions in the remainder of the peptide, suggesting that NKG5 encodes a secreted protein. Both NKG5 and 519 are expressed in NK and T cells but not in a variety of other hematopoietic cell lines. NKG5 is an abundant transcript and its level of expression is about 40 times that of 519 in NK and T cells. Southern blot and DNA sequence analyses suggest that NKG5 and 519 mRNAs are transcripts from a single gene that has allelic polymorphism.
Dysregulated expression of the T cell cytokine Eta-1 in CD4-8- lymphocytes during the development of murine autoimmune disease.
Authors: Patarca R , Wei F Y , Singh P , Morasso M I , Cantor H
Published: June 2004

The development of autoimmune disease in the MRL/MpJ-lpr inbred mouse strain depends upon the maturation of a subset of T lymphocytes that may cause sustained activation of immunological effector cells such as B cells and macrophages. We tested the hypothesis that abnormal effector cell activation reflects constitutive overexpression of a T cell cytokine. We found that a newly defined T cell cytokine, Eta-1, is expressed at very high levels in T cells from MRL/l mice but not normal mouse strains and in a CD4-8- 45R+ T cell clone. The Eta-1 gene encodes a secreted protein that binds specifically to macrophages, possibly via a cell adhesion receptor, resulting in alterations in the mobility and activation state of this cell type (Patarca, R., G. J. Freeman, R. P. Singh, et al. 1989. J. Exp. Med. 170:145; Singh, R. P., R. Patarca, J. Schwartz, P. Singh, and H. Cantor. 1990. J. Exp. Med. 171:1931). In addition, recent studies have indicated that Eta-1 can enhance secretion of IgM and IgG by mixtures of macrophages and B cells (Patarca, R., M. A. Lampe, M. V. Iregai, and H. Cantor, manuscript in preparation). Dysregulation of Eta-1 expression begins at the onset of autoimmune disease and continues throughout the course of this disorder. Maximal levels of Eta-1 expression and the development of severe autoimmune disease reflect the combined contribution of the lpr gene and MRL background genes.
VLA-4 mediates CD3-dependent CD4+ T cell activation via the CS1 alternatively spliced domain of fibronectin.
Authors: Nojima Y , Humphries M J , Mould A P , Komoriya A , Yamada K M , Schlossman S F , Morimoto C
Published: June 2004

We previously showed that fibronectin (FN) synergized with anti-CD3 in induction of CD4+ T cell proliferation, and that VLA-5 acted as a functional FN receptor in a serum-free culture system. In the present study, we showed that VLA-4 is also involved in this CD3-dependent CD4 cell activation through its interaction with the alternatively spliced CS1 domain of FN. When highly purified CD4 cells were cultured on plates coated with anti-CD3 plus synthetic CS1 peptide-IgG conjugate, significant proliferation could be observed. Neither CS1 alone nor anti-CD3 alone induced this activation. This proliferation was completely blocked by anti-VLA beta 1 (4B4) and anti-VLA-4 (8F2), while anti-VLA-5 (monoclonal antibody [mAb] 16 and 2H6) had no effect. These data indicate that VLA-4 mediates CD3-dependent CD4 cell proliferation via the CS1 domain of FN. Anti-VLA-4 also partially (10-40%) inhibited CD4 cell proliferation induced by native FN plus anti-CD3, implying that the CS1 domain is active in the native plasma FN. However, this native FN-dependent proliferation was entirely abolished by addition of anti-VLA-5 alone. Moreover, when native FN-coated plates were pretreated with anti-FN (mAb 333), which blocks RGDS sites but not CS1 sites, no CD4 cell activation could be observed. These results strongly suggest that CD4 cell activation induced by plasma FN/anti-CD3 may be dependent on both VLA4/CS1 and VLA5/RGDS interactions, although the latter interaction may be required for function of the former.
Interleukin 2 gene transfer into tumor cells abrogates tumorigenicity and induces protective immunity.
Authors: Gansbacher B , Zier K , Daniels B , Cronin K , Bannerji R , Gilboa E
Published: June 2004

To study the effects of localized secretion of cytokines on tumor progression, the gene for human interleukin 2 (IL-2) was introduced via retroviral vectors into CMS-5 cells, a weakly immunogenic mouse fibrosarcoma cell line of BALB/c origin. Secretion of low levels of IL-2 from the tumor cells abrogated their tumorigenicity and induced a long-lasting protective immune response against a challenge with a tumorigenic dose of parental CMS-5 cells. Co-injection of IL-2-producing CMS-5 cells with unmodified tumor cells inhibited tumor formation even when highly tumorigenic doses of CMS-5 cells were used. Cytolytic activity in mice injected with parental CMS-5 cells was transient and was greatly diminished 3 wk after injection, as commonly observed in tumor-bearing animals. However, in mice injected with IL-2-producing cells, tumor-specific cytolytic activity persisted at high levels for the duration of the observation period (at least 75 d). High levels of tumor-specific cytolytic activity could also be detected in parental CMS-5 tumor-bearing animals 18 d after inoculation with tumor cells, if IL-2-producing CMS-5 cells but not unmodified parental tumor cells were used as targets. These studies highlight the potential advantages of localized secretion of cytokines mediated via gene transfer to induce potent anti-tumor immune responses.
Characterization of thymus-derived lymphocytes expressing Ti alpha-beta CD3 gamma delta epsilon zeta-zeta, Ti alpha-beta CD3 gamma delta epsilon eta-eta or Ti alpha-beta CD3 gamma delta epsilon zeta-zeta/zeta-eta antigen receptor isoforms: analysis by gene transfection.
Authors: Clayton L K , Bauer A , Jin Y J , D'Adamio L , Koyasu S , Reinherz E L
Published: June 2004

To characterize the function of the CD3 eta subunit of the T cell receptor (TCR), we have used cDNAs encoding CD3 zeta, CD3 eta, or both to reconstitute a variant of a cytochrome c-specific, I-Ek-restricted murine T cell hybridoma, termed MA5.8, which lacks CD3 zeta and CD3 eta proteins. We provide direct evidence that assembly and surface expression of TCRs can be mediated by either of these subunits separately or together. However, the level of TCR expression on zeta transfectants is up to one order of magnitude greater than that on eta transfectants, implying that CD3 eta is weakly associated with the pentameric Ti alpha-beta CD3 gamma delta epsilon complex and/or inefficient at salvaging the incomplete TCR from lysosomal degradation. As a component of the TCR, the CD3 eta subunit preferentially forms a heterodimer with CD3 zeta, but is also able to form a CD3 eta-eta homodimer. Crosslinking of Ti alpha-beta CD3 gamma delta epsilon zeta-zeta, Ti alpha-beta CD3 gamma delta epsilon eta-eta, or Ti alpha-beta CD3 gamma delta epsilon zeta-zeta/zeta-eta TCR isotypes with anti-CD3 epsilon monoclonal antibody or a cytochrome c peptide epitope on I-Ek antigen-presenting cells mediates signal transduction resulting in reversible cell-cycle arrest of transfected clones. Given the potential for diversity of signals generated by these functional TCR isotypes and the expression of the CD3 eta gene product in the thymus, CD3 eta is likely to play a role in selection and/or activation of thymocytes during development.
Prooxidant activity of transferrin and lactoferrin.
Authors: Klebanoff S J , Waltersdorph A M
Published: June 2004

Acceleration of the autoxidation of Fe2+ by apotransferrin or apolactoferrin at acid pH is indicated by the disappearance of Fe2+, the uptake of oxygen, and the binding of iron to transferrin or lactoferrin. The product(s) formed oxidize iodide to an iodinating species and are bactericidal to Escherichia coli. Toxicity to E. coli by FeSO4 (10(-5) M) and human apotransferrin (100 micrograms/ml) or human apolactoferrin (25 micrograms/ml) was optimal at acid pH (4.5-5.0) and with logarithmic phase organisms. Both the iodinating and bactericidal activities were inhibited by catalase and the hydroxyl radical (OH.) scavenger mannitol, whereas superoxide dismutase was ineffective. NaCl at 0.1 M inhibited bactericidal activity, but had little or no effect on iodination. Iodide increased the bactericidal activity of Fe2+ and apotransferrin or apolactoferrin. The formation of OH.was suggested by the formation of the OH.spin-trap adduct (5,5-dimethyl-1-pyroline N-oxide [DMPO]/OH)., with the spin trap DMPO and the formation of the methyl radical adduct on the further addition of dimethyl sulfoxide. (DMPO/OH).formation was inhibited by catalase, whereas superoxide dismutase had little or no effect. These findings suggest that Fe2+ and apotransferrin or apolactoferrin can generate OH.via an H2O2 intermediate with toxicity to microorganisms, and raise the possibility that such a mechanism may contribute to the microbicidal activity of phagocytes.
Distinct fates of self-specific T cells developing in irradiation bone marrow chimeras: clonal deletion, clonal anergy, or in vitro responsiveness to self-Mls-1a controlled by hemopoietic cells in the thymus.
Authors: Speiser D E , Chvatchko Y , Zinkernagel R M , MacDonald H R
Published: June 2004

Elimination of potentially self-reactive T lymphocytes during their maturation in the thymus has been shown to be a major mechanism in accomplishing self-tolerance. Previous reports demonstrated that clonal deletion of self-Mls-1a-specific V beta 6+ T lymphocyte is controlled by a radiosensitive I-E+ thymic component. Irradiation chimeras reconstituted with I-E- bone marrow showed substantial numbers of mature V beta 6+ T cells despite host Mls-1a expression. Analysis of the functional properties of such chimeric T cells revealed a surprising variability in their in vitro reactivity to host Mls-1a, depending on the H-2 haplotype of stem cells used for reconstitution. In chimeras reconstituted with B10.S (H-2s) stem cells, mature V beta 6+ lymphocytes were present but functionally anergic to host-type Mls-1a in vitro. In contrast, in chimeras reconstituted with B10.G (H-2q) bone marrow, nondeleted V beta 6+ cells were highly responsive to Mls-1a in vitro. These findings suggest that clonal anergy of V beta 6+ cells to self-Mls-1a may be controlled by the affinity/avidity of T cell receptor interactions with bone marrow-derived cells in the thymus depending on the major histocompatibility complex class II molecules involved. Furthermore, chimeras bearing host (Mls-1a)-reactive V beta 6+ cells did not differ clinically from those with anergic or deleted V beta 6+ cells and survived more than one year without signs of autoimmune disease. Interestingly, their spleen cells had no Mls-1a stimulatory capacity in vitro. Therefore, regulation at the level of antigen presentation may be an alternative mechanism for maintenance of tolerance to certain self-antigens such as Mls-1a.
Suppression of developmental anomalies by maternal macrophages in mice.
Authors: Nomura T , Hata S , Kusafuka T
Published: June 2004

We tested whether nonspecific tumoricidal immune cells can suppress congenital malformations by killing precursor cells destined to cause such defects. Pretreatment of pregnant ICR mice with synthetic (Pyran copolymer) and biological (Bacillus Calmette-Guérin) agents significantly suppressed radiation- and chemical-induced congenital malformations (cleft palate, digit anomalies, tail anomalies, etc.). Such suppressive effects were associated with the activation of maternal macrophages by these agents, but were lost either after the disruption of activated macrophages by supersonic waves or by inhibition of their lysosomal enzyme activity with trypan blue. These results indicate that a live activated macrophage with active lysosomal enzymes can be an effector cell to suppress maldevelopment. A similar reduction by activated macrophages was observed in strain CL/Fr, which has a high spontaneous frequency of cleft lips and palates. Furthermore, Pyran-activated maternal macrophages could pass through the placenta, and enhanced urethane-induced cell killing (but not somatic mutation) in the embryo. It is likely that a maternal immunosurveillance system eliminating preteratogenic cells allows for the replacement with normal totipotent blast cells during the pregnancy to protect abnormal development.
A repetitive peptide of Leishmania can activate T helper type 2 cells and enhance disease progression.
Authors: Liew F Y , Millott S M , Schmidt J A
Published: June 2004

Leishmaniasis provides a biologically relevant model to analyze the heterogeneity of CD4+ T cells and may lead to answering the major question of the mechanism for the preferential induction of T helper type 1 (Th1) and Th2 cells. Using synthetic peptides corresponding to the tandemly repeating regions of Leishmania proteins, we have identified an epitope that can preferentially induce the disease-exacerbating Th2 cells in susceptible BALB/c mice. Lymph node cells from BALB/c mice immunized subcutaneously with the octamer (p183) of the repeating 10-mer peptide EAEEAARLQA proliferated strongly against the peptide as well as the soluble antigen extract (SolAg) of Leishmania major. The proliferative T cells are CD4+, major histocompatibility complex class II restricted, and secrete interleukin 4 (IL-4) but little or no IL-2 and interferon gamma when stimulated with the peptide in vitro. T cells from BALB/c mice with progressive disease, but not from BALB/c mice cured of the infection, recognized this epitope. BALB/c mice injected subcutaneously with p183 developed significantly exacerbated disease when subsequently challenged with L. major. Furthermore, subcutaneous injection with p183 prevented the subsequent induction of resistance against L. major by intravenous immunization with soluble antigen. The T cell response to p183 is H-2d restricted. Immunization of the genetically resistant B10.D2 mice with p183 also produced strong T cell responses and exacerbated disease when challenged with L. major.
Differences defined by bone marrow transplantation suggest that lpr and gld are mutations of genes encoding an interacting pair of molecules.
Authors: Allen R D , Marshall J D , Roths J B , Sidman C L
Published: June 2004

Homozygosity for either of the lymphoproliferation (lpr) or generalized lymphoproliferative disease (gld) mutations of mice causes the development of systemic lupus erythematosus-like autoimmune syndromes that are characterized by severe lymphadenopathy and highly elevated serum immunoglobulin levels. Although the mutations are nonallelic, analysis of homozygous lpr/lpr and gld/gld mice on the same strain background has indicated that the pathology and severity of the autoimmune syndromes induced by these mutations are indistinguishable. To explain this, it has previously been suggested that lpr and gld may represent mutations in molecules involved in sequential steps of an intracellular metabolic pathway of T cells. We have now investigated the behavior of both lpr and gld in a variety of bone marrow chimeras and have found that functional differences between lpr and gld become apparent after bone marrow transfer. Transfer of lpr/lpr bone marrow to irradiated congenic +/+ recipients caused the development of a graft-vs.-host-like lymphoid wasting syndrome, whereas transfer of gld/gld bone marrow to +/+ recipients resulted in development of a gld-like autoimmune syndrome. Additionally, gld/gld hosts behaved like +/+ hosts irrespective of the genotype of the donor bone marrow, whereas lpr/lpr hosts behaved unlike +/+ hosts when reconstituted with either lpr/lpr, gld/gld, or +/+ bone marrow. These are the first clear differences between these two mutations yet described. Our studies indicate that the molecule altered by the gld mutation is expressed only by bone marrow-derived cells, whereas the molecule altered by the lpr mutation is expressed by both bone marrow-derived cells and by one or more peripheral radioresistant cell populations. To reconcile these differences with the fact that homozygous lpr/lpr and gld/gld mice are indistinguishable, we suggest an alternative model for the relationship between the lpr and gld mutations in which the two molecules affected represent an interacting ligand-receptor pair expressed by different cells.
Coding sequences of the tal-1 gene are disrupted by chromosome translocation in human T cell leukemia.
Authors: Chen Q , Yang C Y , Tsan J T , Xia Y , Ragab A H , Peiper S C , Carroll A , Baer R
Published: June 2004

The tal-1 proto-oncogene encodes a helix-loop-helix DNA-binding protein that has been implicated in the formation of T cell acute lymphoblastic leukemia (T-ALL). Patients with T-ALL harbor structural rearrangements of tal-1 that result from either local DNA deletion or t(1;14)(p34;q11) chromosome translocation. By analyzing t(1;14)(p34;q11) chromosomes from a series of patients, we have now identified a discrete region of tal-1 wherein most of the translocation breakpoints occur. Moreover, mapping of tal-1 genomic DNA revealed that coding exons are situated on both sides of the t(1;14)(p34;q11) major breakpoint region. Hence, the translocated allele of tal-1 is truncated in a manner that reduces its amino acid coding potential.
In vitro expansion of CD3/TCR- human thymocyte populations that selectively lack CD3 delta gene expression: a phenotypic and functional analysis.
Authors: Poggi A , Biassoni R , Pella N , Paolieri F , Bellomo R , Bertolini A , Moretta L , Mingari M C
Published: June 2004

Highly purified CD1-3-4-8- human thymocytes were obtained by panning techniques combined with cell depletion with antibody-coated magnetic beads. Most of these cells expressed cytoplasmic CD3 antigen, as assessed by mAbs known to react with the CD3 epsilon chain. After culture with low doses of PMA (0.5 ng/ml) and subsequent addition (at 24 h) of recombinant interleukin 2 (rIL-2; 100 U/ml) cells underwent extensive proliferation (40-60-fold of the initial cell input after 2 wk). The majority of the proliferating cells were CD3-TCR-. The remaining cells (5-40%) were represented by CD3+ TCR gamma/delta+ (BB3- A13+) cells. Further removal of CD3+ TCR-gamma/delta+ cells resulted in highly purified CD3- populations that further proliferated in culture with no substantial phenotypic changes. When CD3+ thymocytes were cultured under the same experimental conditions, only CD3+ TCR-alpha/beta+ cells could be detected, thus indicating that PMA did not affect the surface expression of the CD3/TCR complex, but rather induced preferential growth of CD3- thymocytes. Surface marker analysis of cultured CD3- thymocytes showed that they were homogeneously CD7+, whereas low proportions of cells expressed CD2 and CD8 antigens. Among the natural killer (NK) cell markers, CD56 was highly expressed by all cells, whereas CD16, CD57, CD11b, NKH2, and GL183 were absent. Importantly, these cells were different from peripheral NK cells, as 80-95% of them expressed cytoplasmic CD3 antigen. Functional analysis revealed a strong cytolytic activity against both NK-sensitive (K562) and NK-resistant (M14, Daudi) human target cells. In a redirected killing assay against the Fc gamma R+ P815 cells, mAbs specific for triggering molecules including CD3, CD2, and CD16 failed to augment target cell lysis, while a strong cytolytic effect was induced by PHA. In addition, PHA alone or in combination with PMA induced tumor necrosis factor alpha (TNF-alpha) and interferon gamma (IFN-gamma) (but not IL-2) production by CD3- thymocytes. Cloning of fresh CD1-3-4-8-thymocytes in the presence of PMA and rIL-2 resulted in CD3-CD56+ clones that displayed a pattern of cytolytic activity and lymphokine production similar to that of the polyclonal populations. Northern blot analysis of transcripts coding for CD3/TCR molecules revealed the presence of CD3 zeta, epsilon, and gamma transcripts, while CD3 delta was undetectable. Mature transcripts for both gamma and delta TCR chains could be detected, whereas no TCR-alpha mRNA and only a truncated (1.0 kb) form of TCR-beta mRNA were revealed.(ABSTRACT TRUNCATED AT 400 WORDS)