bluuebunny/crossref_metadata_2025_split huggingface dataset:
Alteration of a model antigen by Au(III) leads to T cell sensitization to cryptic peptides
Authors: Griem Peter , Panthel Klaus , Kalbacher Hubert , Gleichmann Ernst 
Published: March 2007

AbstractCertain metal ions are known to be potent sensitizers, but the self proteins modified by metal ions and the self peptides recognized by ‘metal‐specific’ T cells are unknown. In humans and mice treatment with gold anti‐rheumatic drugs, containing Au(I), may lead to allergic and autoimmune side effects. Human and murine T cells do not react to Au(I), however, but to the reactive metabolite Au(III). Here we show that alteration by Au(III) of a model antigen, bovine ribonuclease (RNase) A, results in T cell sensitization to cryptic peptides of this protein. Upon immunization of mice with Au(III)‐pretreated RNase [RNase/Au(III)], CD4+ T cell hybridomas specific for RNase/Au(III) were obtained in addition to those recognizing the immunodominant peptide RNase 74–88; the latter also were obtained after immunization with native RNase. RNase/Au(III)‐specific T cell hybridomas reacted against RNase/Au(III) and RNase denatured by S‐sulfonation of cysteine residues, but not against native RNase, or RNase pretreated with Au(I), Al(III), Cu(II), Fe(II), Fe(III), Ni(II), Mn(II), or Zn(II). Using a panel of overlapping, synthetic RNase peptides which were devoid of gold or gold‐induced modifications, epitope mapping revealed that RNase/Au(III)‐specific T cell hybridomas recognized the cryptic peptides 7–21 and 94–108, respectively. Comparison of the proliferative response of bulk CD4+ T cells, prepared from splenocytes after immunization with either RNase/Au(III) or native RNase, revealed that Au(III) pretreatment of RNase led to a markedly enhanced response to the two cryptic peptides while it did not influence the response to the immunodominant peptide. The cryptic peptides were also presented after preincubation of bone marrow‐derived macrophages with RNase and Au(I), but not with RNase alone, suggesting that oxidation of Au(I) to Au(III) and subsequent protein alteration by Au(III) can happen in mononuclear phagocytes. We conclude that Au(III) alteration of proteins alters antigen processing and, thus, leads to presentation of cryptic peptides. This mechanism may shed light on the development of allergic and autoimmune side effects of Au(I) anti‐rheumatic drugs. In addition, it might provide a general mechanism of how metal ions act as T cell sensitizers.
The murine homolog of human Ep‐CAM, a homotypic adhesion molecule, is expressed by thymocytes and thymic epithelial cells
Authors: Nelson Andrew J. , Dunn Robert J. , Peach Robert , Aruffo Alejandro , Farr Andrew G. 
Published: March 2007

AbstractIn this report, we demonstrate that gp40, a molecule previously shown to be expressed by thymic epithelial cell lines in vitro and by thymic epithelial cells in vivo, is the murine homolog of human Ep‐CAM, a calcium‐independent homotypic adhesion molecule. gp40 is also expressed at low levels by thymocytes and peripheral T cells. In the adult thymus, gp40 expression was inversely related to the state of thymocyte maturation, with the highest levels associated with CD4− CD8− and CD4+CD8+ thymocyte populations. Ultrastructural immunohistochemistry revealed gp40 localization to areas of thymocyte/epithelial contact and demonstrated that gp40 is also expressed by thymic dendritic cells. During fetal development, thymocytes at days 14–16 of gestation expressed high levels of gp40. At later stages, the observed decline in the frequency of gp40+ cells and levels of expression correlated with the emergence of αβ+ thymocytes by day 18 of gestation. In short‐term cultures, stimulation of unfractionated adult thymocytes with concanavalin A increased gp40 expression, particularly among CD3hi and CD3int thymocyte populations. This demonstration that Ep‐CAM, initially considered to be expressed primarily by epithelial cells, is also expressed by thymocytes, T cells and antigen‐presenting cells, raises the possibility that Ep‐CAM may contribute to adhesive interactions between thymocytes and epithelial cells or dendritic cells, either in the context of thymocyte development or peripheral T cell trafficking and function.
Identification of overlapping epitopes in mutant ras oncogene peptides that activate CD4+ and CD8+ T cell responses
Authors: Abrams Scott I. , Stanziale Stephen F. , Lunin Scott D. , Zaremba Sam , Schlom Jeffrey 
Published: March 2007

AbstractMutant ras p21 proteins contain sequences which distinguish them from normal endogenous ras and, thus, may represent unique epitopes for T cell recognition of antigen bearing tumor cells. Here, we examined the capacity of a mutant K‐ras 9‐mer peptide to induce in vivo CD8+ cytotoxic T lymphocytes (CTL). The peptide chosen reflected positions 4–12 of the point‐mutated sequence of the K‐ras oncogene encoding the Gly to Val substitution at codon 12. The overall rationale for selecting this particular 9‐mer sequence was threefold: the mutant peptide contained a putative major histocompatibility complex (MHC) class I consensus anchor motif for murine H‐2Kd; specific binding to MHC class I may then create an immunogenic complex for the induction of anti‐ras CD8+ CTL; and finally, the mutant sequence overlapped with a newly characterized anti‐ras CD4+ T helper type 1 epitope, which may have implications for the coordination and activation of both anti‐ras immune mechanisms against the same target cell antigenic determinant. A functional interaction with H‐2Kd was demonstrated with the mutant ras4–12(V12) peptide, but not the normal ras4–12(G12) peptide, which specifically inhibited an H‐2Kd‐restricted, anti‐nucleoprotein NP147–155 CTL response in a dose‐dependent fashion. An anti‐ras CD8+ T cell line was then established from immune splenocytes of BALB/c (H‐2d) mice injected with ras4–12(V12) in adjuvant, which mediated peptide‐specific lysis of syngeneic P815 tumor targets. Cytotoxicity was restricted by H‐2Kd and strongly specific for the mutant ras peptide. Importantly, these anti‐ras CTL specifically lysed a syngeneic tumor line (i.e. A20 lymphoma) transduced with the corresponding point‐mutated ras oncogene, suggesting T cell receptor recognition of endogenously derived antigen. Overall, these data demonstrated that mutant ras p21 at codon 12 (Gly → Val) contained a peptide sequence which exhibited specific functional binding to a murine MHC class I molecule; the ability of the mutant, but not the normal sequence to bind selectively to murine MHC class I likely reflected the generation of a C‐terminal anchor residue; and the ras 4–12(V12) peptide was immunogenic for the production of antigen‐specific CD8+ CTL, which lysed in vitro a syngeneic tumor cell line harboring the mutant K‐ras oncogene.
Expression levels of stress protein gp96 are not limiting for major histocompatibility complex class I‐restricted antigen presentation
Authors: Lammert Eckhard , Arnold Danièle , Rammensee Hans‐Georg , Schild Hansjörg 
Published: March 2007

AbstractImmunization of mice with gp96 induces CTL with specificity for proteins that are expressed in the cells from which gp96 was isolated (Arnold et al., J. Exp. Med. 1995. 182: 885, Udono et al., Proc. Natl. Acad. Sci. USA 1994. 91: 3077). Recently, it has been shown that gp96 from cells transfected with vesicular stomatitis virus (VSV) nucleocapsid protein as well as gp96 loaded in vitro with peptides containing an epitope of this protein are taken up by phagocytic cells which obtain thereby the capacity for stimulating VSV‐specific cytotoxic T lymphocytes (Suto and Srivastava, Science 1995. 269: 1585). The immunization experiments together with the peptide transfer from gp96/peptide complexes to major histocompatibility complex (MHC) class I molecules of phagocytic cells are consistent with the hypothesis that the endoplasmic reticulum‐resident protein gp96 plays a crucial role in the antigen presentation of a cell (Srivastava et al., Immunogenetics 1994. 39: 93). To examine the involvement of gp96 in class I‐restricted antigen presentation, we reduced gp96 RNA and protein levels by transfecting P13.1 cells with a vector containing part of gp96 cDNA in antisense orientation to the promoter. We found that antisense clones expressing strongly reduced levels of gp96 mRNA and gp96 protein show normal levels of MHC class I molecules on the cell surface and are recognized by T cells to the same extent as wild‐type cells. Thus, our results show that normal levels of gp96 expression in a cell are not limiting for class I‐restricted antigen presentation.
Regions of RAG1 protein critical for V(D)J recombination
Authors: Kirch Susan A. , Sudarsanam Priya , Oettinger Marjorie A. 
Published: March 2007

AbstractThe products of the recombination activating genes RAG1 and RAG2 are essential for activating V(D)J recombination, and thus are indispensable for the production of functional and diverse antigen receptors. To investigate the function of RAG1, we have tested a series of insertion and substitution mutations for their ability to induce V(D)J rearrangement on both deletional and inversional plasmid substrates. With these substrates we were also able to assess the effects of these mutations on both coding and signal joint formation, and to show that any one mutant affected all these reactions similarly. As defined previously, the core active regions of RAG1 and RAG2 permit the deletion of 40% and 25%, respectively, of well‐conserved sequence. We show here that this “dispensable” region of RAG1 is not necessary for coding joint formation or for recombination of an integrated substrate, and that this portion is not functionally redundant with the “dispensable” region of RAG2. Recombination with these core regions is also still subject to the 12/23 joining rule. Further, the minimal essential core region of RAG1 can be located within an even smaller portion of the gene.
The murine VpreB1 and VpreB2 genes both encode a protein of the surrogate light chain and are co‐expressed during B cell development
Authors: Dul Jeanne L. , Argon Yair , Winkler Thomas , ten Boekel Edwin , Melchers Fritz , Mårtensson Inga‐Lill 
Published: March 2007

AbstractThe surrogate light chain is composed of two polypeptides, VpreB and λ5. In the mouse there are two VpreB genes which are 99% identical within the coding regions. Extensive restriction enzyme mapping and sequencing of these two genes showed that only the coding region and immediate 5′ and 3′ flanking sequences exhibited such high homology. More distal sequences have diverged considerably. The region 5′ of the respective gene directed transcription of a reporter gene in a pre‐B cell line, indicating that it contained promoter, and perhaps enhancer function. The VpreB2 gene is functional, as it directed the production in COS cells of a 16‐kDa protein that assembled with λ5 and was recognized by a VpreB‐specific monoclonal antibody. Using transfected COS cells expressing either VpreB1 or VpreB2, a PCR assay was developed to examine the steady state level of transcripts from each gene. When this assay was applied to a number of cell lines representing early stages of B cell differentiation, co‐expression of the two genes was observed in every case. VpreB1 and VpreB2 were co‐expressed in the fetal liver of CB17 mice, where peak expression of each gene occurred at days 16–17 of gestation. Similarly, adult bone marrow from several strains of mice expressed both genes. In sorted bone marrow cells expression of both VpreB genes was detected in pro‐B/pre‐BI and large pre‐BII cells, while the RNA steady state levels were at least 100‐fold lower in small pre‐BII and immature/mature B cells. Finally, single‐cell reverse transcriptase‐polymerase chain reaction on such sorted bone marrow cells detected VpreB1 and VpreB2 expression in at least 30% of all pro‐B/pre‐BI cells and large Ig heavy chain, surrogate light chain (pre‐B receptor) expressing pre‐BII cells. These results demonstrate that the control of expression of the two VpreB genes overlaps during development. They suggest that both VpreB1 and VpreB2 polypeptides can assemble with λ5 and μ to form pre‐B cell receptor complexes.
Tolerance towards resident intestinal flora in mice is abrogated in experimental colitis and restored by treatment with interleukin‐10 or antibodies to interleukin‐12
Authors: Duchmann Rainer , Schmitt Edgar , Knolle Percy , Zum Büschenfelde Karl‐Hermann Meyer , Neurath Markus 
Published: March 2007

AbstractThere is now increasing evidence that hyperresponsiveness towards intestinal flora is a crucial event in the pathogenesis of inflammatory bowel disease (IBD). In support of this hypothesis, we recently described in humans that tolerance exists towards indigenous intestinal flora but is broken in active IBD lesions. In the present study, we have attempted to transfer this model into mice from different genetic backgrounds (BALB/c, SJL/J, C3H/HeJ). We found that mononuclear cells from spleen, small bowel and large bowel of mice do not proliferate, i.e. are tolerant when exposed to bacterial sonicates derived from autologous intestine (BsA) but do proliferate, i.e. are immune when exposed to bacterial sonicates derived from the heterologous intestine of syngenic littermates (BsH). Furthermore, we demonstrate that both local and systemic tolerance to BsA is broken in a murine model of chronic intestinal inflammation induced by the hapten reagent 2,4,6‐trinitrobenzene sulfonic acid (TNBS), which mimics several important characteristics of Crohn's disease. Tolerance to BsA was restored and TNBS‐induced colitis was abrogated in mice systemically treated with interleukin (IL)‐10 or antibodies to IL‐12. Treatment specifically restored tolerance to BsA, but did not suppress proliferation to BsH. In summary, we here report a new mouse model for the study of immunity and tolerance towards bacterial products. Our data suggest that tolerance to BsA is an important protective mechanism and that restoration of tolerance to resident intestinal flora by IL‐10 and antibodies to IL‐12 may be of potential therapeutic utility in patients with inflammatory bowel disease.
Human CD4 and human major histocompatibility complex class II (DQ6) transgenic mice: supersensitivity to superantigen‐induced septic shock
Authors: Yeung Rae S. M. , Penninger Josef M. , Kündig Thomas , Khoo Wilson , Ohashi Pamela S. , Kroemer Guido , Mak Tak W. 
Published: March 2007

AbstractRodents are significantly less sensitive to enterotoxin‐induced shock, and are thus not valid human disease models. Here, we describe a mouse strain carrying the human CD4 and human major histocompatibility complex (MHC) class II (DQ6) transgenes in an endogenous CD4‐ and CD8‐deficient background. T lymphocytes from these animals react to minute amounts (10–100 times less than control mice) of staphylococcal enterotoxin B (SEB) in vitro, similar to concentrations to which human cells react. In vivo, these double‐transgenic, double‐knockout mice succumb to normally sublethal amounts of SEB. This sensitivity is not due to a biased T cell receptor Vβ repertoire, increased T cell reactivity, or increased sensitivity to macrophage‐derived cytokines. Rather, tumor necrosis factor (TNF)‐α production by T cells and serum levels of TNF‐α correlate precisely with the clinical syndrome, showing a biphasic T cell‐dependent response. These data show that both human CD4 and MHC class II molecules can render mice supersensitive to superantigen‐induced septic shock syndrome. This animal model mimics the progression of septic shock in man by transforming normally resistant mice into hypersensitive SEB responders, a trait that is characteristic of humans. Mice that have been humanized by exchanging autochthonous superantigen ligands by their human equivalents may be useful to decipher superantigen responses in vivo and to assess the pathogenesis of superantigen‐associated diseases.
The macrophage cell surface glycoprotein F4/80 is a highly glycosylated proteoglycan
Authors: Haidl Ian D. , Jefferies Wilfred A. 
Published: March 2007

AbstractMolecules whose expression is limited to particular leukocyte populations are of interest since they may perform unique functions for these cells. We therefore examined the biochemical nature of the F4/80 molecule, which is expressed solely on macrophage and dendritic cell subpopulations. Our study clearly indicates that post‐translational modifications, which can influence both a protein's structural and functional features, constitute a major component of the 160‐kDa cell‐surface F4/80 molecule. The F4/80 molecule is synthesized as a single polypeptide chain which acquires numerous intramolecular disulfide bonds and requires an extended time period (T1/2 = 60 min) for transport to an endoglycosidase H‐resistant form. The F4/80 molecule contains extensive N‐linked glycosylation which contributes approximately 40 kDa to the mature molecule. The N‐linked carbohydrates are of the branched, complex type, containing repeating N‐acetylglycosamine or N‐acetyllactosamine units which mediate the reactivity of the F4/80 molecule with Datura stramonium lectin. O‐linked glycosylation is also present and contributes approximately 10 kDa to the F4/80 molecule. Furthermore, the sialic acid modifications of the F4/80 molecule are primarily through α2‐6 linkages to galactose. Finally, we demonstrate that the F4/80 molecule is a proteoglycan modified by chondroitin sulfate glycosaminoglycans. In addition to clarifying the nature of the F4/80 molecule biochemically, these posttranslational modifications have specific implications for molecular recognition processes. We conclude that the modifications of the F4/80 molecule may mediate cell‐cell recognition, cell adhesion, or ligand binding independently of the F4/80 molecule protein core.
Divergent effects of FcγRIIIA ligands on the functional activities of human natural killer cells in vitro
Authors: Sulica Andrei , Metes Diana , Gherman Maria , Whiteside Theresa L. , Herberman Ronald B. 
Published: March 2007

AbstractThe Fcγ receptor (R)IIIA (CD16) plays an important role in regulating the cytotoxic and non‐cytotoxic functions of human natural killer (NK) cells. Some anti‐CD 16 monoclonal antibodies (mAb) have been shown to stimulate NK activity, while human monomeric (m) IgG induces dose‐dependent inhibition of NK activity. To explore further these interactions mediated via FcγRIIIA, purified NK cells were cultured for 2–3 days in the presence of mIgG, 3G8 mAb, interleukin‐2 (IL‐2) or a combination of mIgG or 3G8 with IL‐2. Binding of mIgG or 3G8 to FcγRIIIA induced divergent effects of functions of cultured NK cells: 3G8 mAb + IL‐2 induced dose‐dependent inhibition of proliferation attributable to apoptosis; in contrast, mIgG + IL‐2 significantly increased NK cell proliferation. Incubation of NK cells in the presence of mIgG up‐regulated expression of surface activation markers (CD69, IL‐2Rα, ICAM‐1), cytotoxicity, cytokine production (IL‐1β, IFN‐γ and TNF‐α) and release of soluble IL‐2R. Thus, mIgG binding to FcγRIIIA induced stimulatory signals in human NK cells, leading to up‐regulation of IL‐2Rα expression, cell proliferation and cytokine release.
Early message expression of interleukin‐4 and interferon‐γ, but not of interleukin‐2 and interleukin‐10, reflects later polarization of primary CD4+ T cell cultures
Authors: Gollob Kenneth J. , Dutra Walderez O. , Coffman Robert L. 
Published: March 2007

AbstractThe CD4+ T cell subpopulation identified by the Mel‐14high phenotype represents naive T cells that develop into T helper 1 (Th1) cells upon stimulation with anti‐CD3 and interleukin (IL)‐2; however, the addition of IL‐4 induces the development of high IL‐4, IL‐5 and IL‐10‐secreting cells (IL‐4 SC). Here, we investigate the timing of cytokine gene expression in purified naive, Mel‐14high CD4+ T cells after stimulation under conditions that induce one of three dramatically different populations following secondary stimulation: anti‐CD3 + IL‐2 (Th1), anti‐CD3 + IL‐2 and IL‐4 (IL‐4 SC) and anti‐CD3 + anti‐interferon (IFN)‐γ/anti‐IL‐4 (null population). The skewing toward IL‐4 SC development induced by IL‐4 in the primary stimulation was clearly visible by 40 h. The message for IL‐4 was absent at 20 h after stimulation, but was induced 30–100‐fold by 40 h and was accompanied by a decrease in IFN‐γ message. The message for IL‐5 was undetectable. The development of Mel‐14high T cells into a Th1 population follows kinetics similar to those of the development of IL‐4 SC in that IFN‐γ gene transcription was undetectable at 20 h and induced 100‐fold by 40 h post‐stimulation. This induction was inhibited by the inclusion of anti‐IFN‐γ/anti‐IL‐4 during the primary stimulation. Finally, IL‐10 message is present in both cultures that developed into IL‐4 SC and those that developed into Th1 populations, in contrast to the differentiated cultures where there is a segregation between IL‐10 and IFN‐γ expression. Undetectable levels of message for both IL‐4 and IFN‐γ were seen in the null cultures, which expressed both IL‐2 and IL‐10 message. T cells identified by the Mel‐14low phenotype develop into a population that secretes high levels of all cytokines tested, and this phenotype was revealed within 20 h after stimulation as determined by message analysis. These results demonstrate an early correlation of gene transcription for IL‐4 and IFN‐γ, with the development of IL‐4 SC and Th1 cells, and a lack of a correlation at these early time points for expression of IL‐2, IL‐5 and IL‐10.
Antigen‐specific B cells are required for the secondary response of T cells but not for their priming
Authors: Chowdhury Mohammed Golam Mowla , Maeda Ken‐Ichi , Yasutomo Koji , Maekawa Yoichi , Furukawa Atsuko , Azuma Miyuki , Nagasawa Hideyuki , Himeno Kunisuke 
Published: March 2007

AbstractWe studied the potential role of B cells in T cell responses using severe‐combined immunodeficient (SCID) mice grafted with the thymus from fetal C.B‐17 mice (TG mice). These mice developed both CD4+ and CD8+ T cells, but not B cells within 2 months after transplantation. TG mice showed normal delayed‐type hypersensitivity responses against the immunizing antigen ovalbumin (OVA). Lymph node (LN) cells of TG mice proliferated well in response to concanavalin A (Con A). Further, Con A stimulation induced the production of interleukin (IL)‐2, IL‐6 and interferon (IFN)‐γ and the expression of IL‐4 mRNA. Thus, TG mice were reconstituted without remarkable immunodeficiency. However, these T cells failed to proliferate to OVA stimulation. Response to OVA was also inhibited in SCID mice grafted with fetal C.B‐17 liver cells when B cells were depleted in the proliferation assay. Unresponsiveness against immunizing antigen was restored by the addition of antigen‐primed B cells, but not by naive B cells, lipopolysaccharide‐activated B cells or B cells primed with sheep red blood cells. Next, we examined whether antigen‐primed B cells could induce T cell responses without professional antigen‐presenting cells (APC). T and B cells were purified from OVA‐immunized mice by cell sorter. These T cells proliferated in response to OVA and produced IFN‐γ in the absence of non‐B APC. When anti‐CD80 or anti‐CD86 was added in the assay, proliferation and IFN‐γ production was inhibited. These results indicate that B cells activated specifically with antigen are required for the secondary response of T cells, but not for their priming.