bluuebunny/crossref_metadata_2025_split huggingface dataset:
Bone marrow can function as a lymphoid organ during a primary immune response under conditions of disrupted lymphocyte trafficking.
Authors: Tripp R A , Topham D J , Watson S R , Doherty P C 
Published: December 2022

Abstract
In this study we sought to better understand lymphocyte trafficking patterns and the function of secondary lymphoid organs, such as the spleen, during the generation of virus-specific T cell precursors. Treatment of mice with the Mel-14 mAb to CD62L, the lymph node homing receptor, limits trafficking of naive T cells into lymph nodes through high endothelial venules. Administering Mel-14 following respiratory infection with influenza virus forced the generation of primary virus-specific CD4+ and CD8+ T cell precursors from the mediastinal lymph nodes to the spleen. However, splenectomy did not seriously impede virus clearance from the lung and, despite a substantial reduction of the total lymphocyte pool, the acute T cell responses in the regional lymph nodes were largely normal. Mel-14 treatment of splenectomized mice did not affect clonal expansion of the virus-specific T cells in the MLN, while the response in the cervical lymph nodes was still greatly inhibited. More surprisingly, virus-specific T cell precursors were now detected from days 5 to 6 after infection in the bone marrow (BM) of the splenectomized, Mel-14-treated mice. This was not due to contamination with circulating T cells or infection of BM cells because the distribution profiles of precursor T cells for PBL and BM diverged and PCR analysis showed no evidence of virus replication in the BM. It appears that, under these conditions of disrupted lymphocyte trafficking, the BM can supplant the secondary lymphoid tissue either as a site of primary immune response or as a cache for excess T cell precursors.
Modifications of peptide ligands enhancing T cell responsiveness imply large numbers of stimulatory ligands for autoreactive T cells.
Authors: Vergelli M , Hemmer B , Kalbus M , Vogt A B , Ling N , Conlon P , Coligan J E , McFarland H , Martin R 
Published: December 2022

Abstract
In this report, we demonstrate for autoreactive T cell clones that single amino acid modifications of the antigenic ligand can result in not only abrogated, decreased, or unmodified, but also increased, T cell responsiveness (superagonist ligands). We further studied the effects of combinations of multiple substitutions with different effects in single peptides. Experiments with peptides carrying multiple amino acid exchanges revealed that the final outcome of TCR ligation by a given ligand is the integration of negative, neutral, and positive effects of each single residue. In addition, the introduction of superagonist substitutions together with nonconservative modifications of primary and secondary TCR contacts resulted in stimulatory ligands. These findings indicate that: 1) the specificity of a single TCR is highly degenerate; 2) ligands exist for autoreactive T cells that have higher agonist activity than the autoantigen itself; 3) the rules to search for cross-reactive epitopes in autoimmunity should take into account that amino acids at certain positions within an antigenic peptide may exert superagonist activity and compensate for the negative effects of residues at other positions that would otherwise not be tolerated.
Carbohydrate and peptide specificity of MHC class II-restricted T cell hybridomas raised against an O-glycosylated self peptide.
Authors: Jensen T , Hansen P , Galli-Stampino L , Mouritsen S , Frische K , Meinjohanns E , Meldal M , Werdelin O 
Published: December 2022

Abstract
MHC class II E(k)-restricted, IL-2 secreting T cell hybridomas were raised against the synthetic glycopeptide Hb(67-76)-alpha-GalNAc, (T72(Tn)), in CBA/J mice (H-2(k)). The fine specificity of the hybridomas against the glycan moiety was investigated by testing their response against a panel of Hb(67-76)-derived glycopeptides, all with a glycan attached to serine or threonine at the position 72 in the peptide, but with different glycans. The hybridomas showed a high degree of specificity for the alpha-GalNAc moiety with few and faint cross-responses to the glycopeptides having other glycans attached even though some of these were structurally very similar to alpha-GalNAc. The fine specificity of the hybridomas for the peptide moiety was investigated by testing their responses to a panel of Hb(67-76)-alpha-GalNAc glycopeptides with alanine substitutions at all positions except at the two MHC binding anchor positions, I68 and K76, and the T72 to which the alpha-GalNAc was attached. Glycopeptides substituted with alanine at positions where the amino acid side chain pointed toward the TCR did not stimulate the hybridomas, whereas glycopeptides substituted with alanine at positions orientated down into the MHC binding groove stimulated many of the hybridomas. These results indicate that the glycan attached to the peptide as well as solvent-accessible parts of the peptide are recognized with a high degree of specificity by the T cells, whereas the parts of the peptide buried in the MHC binding site are less important or totally ignored by the T cells.
Diversity of the cytotoxic melanoma-specific immune response: some CTL clones recognize autologous fresh tumor cells and not tumor cell lines.
Authors: Dufour E , Carcelain G , Gaudin C , Flament C , Avril M F , Faure F 
Published: December 2022

Abstract
In the present work, to analyze the heterogeneity of the tumor-specific cytotoxic immune response, a large number of T cell clones were generated from the infiltrate of a tumor-proximal invaded lymph node, and two kinds of melanoma-specific CD8+ CTL clones were derived. The majority of T cell clones (about a hundred) are characterized by a specific lysis of the autologous tumor cell lines. Among 34 of the latter clones, HLA-A2 molecule and MART-1(27-35) peptide have been shown to play a predominant role in tumor recognition. However, no significant amplification at the tumor site was observed for 3 of these CTL. The other kind of tumor-specific CTL (1 oligoclonal and 2 clonal cell lines) did not lyse the autologous melanoma cell lines but lysed the "fresh" autologous tumor cells in a MHC class I-dependent manner. Functional analysis of the two different CTL clones have shown that they did not lyse NK targets, autologous peripheral monocytes, activated T cells, and transformed B cells or any of the few allogeneic cultured and uncultured melanoma cells we tested. TCR repertoire analysis has shown that one of these CTL clones was significantly detectable "in situ" among tumor-infiltrating lymphocytes, while not detectable among PBMC. Such melanoma-specific lymphocytes, which could not have been picked out through conventional screening procedures using tumor cell lines, could potentially play a role in tumor rejection. These results suggest that the immune response analyzed toward melanoma cell lines does not totally reflect the in situ immune status.
Intracellular localization of the human receptor for the globular domains of C1q.
Authors: van den Berg R H , Prins F , Faber-Krol M C , Lynch N J , Schwaeble W , van Es L A , Daha M R 
Published: December 2022

Abstract
This study was performed to determine the localization of the recently described receptor for the globular domain of C1q, gC1qR. In contrast to previous reports, we were not able to detect significant surface expression of gC1qR on Raji cells, monocytes, neutrophils, human or rat mesangial cells, the endothelial cell line EA.hy 926, or HUVEC using FACS analysis. Only by using digoxigenin-conjugated Abs could some surface staining of gC1qR be observed on rat mesangial cells and neutrophils. However, after permeabilizing these cells with saponin, a strong positive intracellular staining for gC1qR was observed by FACS, fluorescence microscopy on coverslips, and confocal laser scanning microscopic analysis. By reflection contrast microscopy and electron microscopy on ultrathin sections of permeabilized Raji cells, it was shown that gC1qR is present in double membranous cytoplasmic vesicles located in the proximity of the plasma membrane. To determine whether certain conditions could induce surface expression of gC1qR, Raji cells were either stimulated with T cell growth factor, LPS, or driven to apoptosis by incubation with fenretinide or by serum depletion. None of the conditions resulted in significant surface expression of gC1qR. Our hypothesis that gC1qR is not a surface molecule but a soluble molecule that is secreted by cells is supported by the observation that gC1qR is found in significant concentrations in supernatants of several cultured cells and in normal human and rat sera. Our results suggest that the recently described gC1qR is not a cell surface receptor, but a soluble binding protein with affinity for the globular heads of C1q. Excreted gC1qR might act as a potential fluid phase regulator of complement activation.
Initiation of autoimmune diabetes in NOD/Lt mice is MHC class I-dependent.
Authors: Serreze D V , Chapman H D , Varnum D S , Gerling I , Leiter E H , Shultz L D 
Published: December 2022

Abstract
MHC class II alleles clearly contribute a primary genetic component of susceptibility to autoimmune insulin-dependent diabetes mellitus (IDDM) in nonobese diabetic (NOD) mice. However, IDDM does not occur in NOD mice made MHC class I-deficient by a functionally inactivated beta2-microglobulin allele (beta2m(null)). In the present study the beta2m(null) mutation was used to examine the relative contributions of MHC class I and class II-dependent T cell responses for initiating autoimmune pancreatic beta cell destruction in NOD mice. Splenocytes from diabetic NOD donors transferred IDDM to both lymphocyte-deficient NOD-scid (class I+) and NOD-scid.beta2m(null) mice (class I-). In contrast, splenocytes from young prediabetic NOD donors only transferred IDDM to class I+, but not class I- NOD-scid recipients. However, splenocytes from prediabetic NOD donors did transfer IDDM to NOD-scid.beta2m(null) recipients previously engrafted with class I+, but not class I-, pancreatic islets. CD4+ T cell lines reactive against some syngeneic class I+ targets could be isolated from NOD.beta2m(null) mice. However, NOD.beta2m(null) T cells underwent activation-driven deletion when transferred into class I+ NOD-scid recipients. Hence, the class I autoreactive T cells present in NOD.beta2m(null) donors did not elicit IDDM when transferred into class I+ NOD-scid recipients. Collectively, these results show that autoimmune IDDM in NOD mice is initiated by MHC class I-dependent T cell responses, but this leads to the subsequent activation of additional T cell populations that can mediate pancreatic beta cell destruction in a MHC class I-independent manner.
Differential regulation of T cell activation for primary versus secondary proliferative responses.
Authors: Gupta M , Satyaraj E , Durdik J M , Rath S , Bal V 
Published: December 2022

Abstract
T cell activation in vivo results in proliferation and generation of effector cytokine-secreting cells, as also in development of memory cells that mount enhanced responses upon restimulation. However, differences in the signals promoting generation of effector vs memory T cells are not yet characterized. In this study, using various strategies to modulate an allorecognition system for priming human T cells in vitro, we show that there are indeed differences between the signaling requirements for a first proliferative response and those for priming T cells for enhanced recall proliferative responses. Using APCs fixed with varying concentrations of paraformaldehyde, we show that the loss of ability of these APCs to generate a first response is not matched by a similar loss in their ability to prime responder T cells for recall responses. Prevention of DNA replication during T cell priming with aphidicolin, a DNA polymerase inhibitor, is not inimical to successful T cell priming. Thus, clonal expansion during priming is less crucial than the primed activation status of T cells for the enhanced recall response. We also show that pentoxifylline, a phosphodiesterase inhibitor, inhibits the primary proliferative response, but its presence during priming enhances the recall response capabilities of T cells. On the other hand, the presence of the calcineurin inhibitor cyclosporin A during priming reduces the efficiency of priming, but at low concentrations it induces, like pentoxifylline, enhancement in recall response capability. These findings have significant implications in designing immunosuppressive therapy and in the analysis of signals for T cell memory commitment.
Role of CD3gamma and CD3delta cytoplasmic domains in cytolytic T lymphocyte functions and TCR/CD3 down-modulation.
Authors: Luton F , Buferne M , Legendre V , Chauvet E , Boyer C , Schmitt-Verhulst A M 
Published: December 2022

Abstract
TCR engagement leads to down-modulation of TCR/CD3 complexes from the T cell surface. The importance of this effect in T cell physiology is unknown. Here, we characterized a CTL clone deficient in TCR/CD3 surface expression that had lost both CD3delta and CD3gamma mRNA, allowing us to address the role of these chains in the assembly, signaling, and dynamics of the TCR/CD3 complex. Expression of either CD3delta or CD3gamma alone failed to reconstitute surface expression of the TCR/CD3 complex, but reconstitution with a cytoplasmically truncated CD3delta (delta t) and a native (gamma) or cytoplasmically truncated (gamma t) human CD3gamma led to reexpression of TCR/CD3 complexes in both cases. This indicated that CD3delta and CD3gamma assume specific functions in TCR/CD3 assembly independently of their cytoplasmic domains. The delta t gamma t variant specifically killed target cells, expressed the IFN-gamma gene in response to Ag, and produced TNF-alpha in response to anti-CD3 mAb, but it was affected in CD3 ligand-induced TCR/CD3 down-modulation. Both PMA- and CD3 ligand-induced TCR/CD3 down-modulation were defective in the delta t gamma t variant, whereas the delta t gamma variants were unaffected, and previously described delta gamma t variants were affected only in PMA-induced down-modulation. Specific protein kinase C (PKC) inhibitors indicated that PMA- but not CD3 ligand-induced down-modulation was dependent on PKC activity. Thus, amino acid sequences present in either the CD3delta or CD3gamma cytoplasmic domain control ligand-induced TCR/CD3 down-modulation, and neither these sequences nor this property are required for cytolysis and IFN-gamma gene expression in response to Ag.
Regulation of human T lymphocyte coactivation with an alpha4 integrin antagonist peptide.
Authors: McIntyre B W , Woodside D G , Caruso D A , Wooten D K , Simon S I , Neelamegham S , Revelle J K , Vanderslice P 
Published: December 2022

Abstract
The cyclic hexapeptide CWLDVC (TBC 772) is an antagonist of alpha4 integrins and a potent inhibitor of lymphocyte interactions with fibronectin, vascular cell adhesion molecule-1, and muscosal vascular addressin cell adhesion molecule-1 (MAdCAM-1). As such, peptide TBC 772 effectively inhibits the activation of freshly isolated human T lymphocytes stimulated with purified vascular cell adhesion molecule-1 coimmobilized with anti-CD3 mAb. The influence of peptide binding on distinct sites of the alpha4beta1 complex was determined by flow cytometry and cellular adhesion assays employing a panel of mAbs. Binding of the alpha4-specific mAb L25 and the beta1-specific mAb 33B6 was not altered by the peptide; however, binding of mAb 19H8, which is specific for a combinatorial epitope of alpha4beta1, was dramatically inhibited. Treatment of lymphocytes with the peptide caused an increase in a ligand-induced epitope on beta1 integrin defined by mAb 15/7. In T cell activation studies using coimmobilized anti-CD3 mAb and the anti-integrin mAbs, the peptide had broader inhibitory activity, suppressing costimulation induced by all the integrin mAbs. The peptide was not generally toxic and was integrin selective in its suppressive activity, as coactivation by ligation of CD3 in conjunction with CD28 or CD26 was not affected. These results suggest that the antagonist peptide CWLDVC can effectively neutralize integrin coactivation systems by a mechanism independent of competitive binding.
Differential effects of Flk-2/Flt-3 ligand and stem cell factor on murine thymic progenitor cells.
Authors: Moore T A , Zlotnik A 
Published: December 2022

Abstract
Most primitive thymic progenitors, termed CD4(low) cells (CD25- CD44+ CD117+), retain the ability to generate multiple lymphoid lineages. T cell lineage commitment occurs as CD4(low) cells differentiate into pro-T cells (CD25+ CD44+ CD117+). We previously reported that the in vitro cytokine responses of CD4(low) and pro-T cells differ. While Flt-3 ligand (Flt-3L) has been shown to be involved in early bone marrow hemopoiesis, its role in thymopoiesis has not been thoroughly examined. Here, we report that Flt-3L has no significant effect on pro-T cells, either by in vitro proliferation or in fetal thymic organ culture repopulation. In contrast, CD4(low) cells cultured in vitro for 3 days in IL-3 + IL-6 + IL-7 + Flt-3L generated a twofold increase in cell number 21 days after transfer into fetal thymic organ culture that increased to sixfold by day 35 when compared with the corresponding CD4(low) cells cultured in IL-3 + IL-6 + IL-7 + stem cell factor. Additionally, the Flt-3L-cultured CD4(low) cells displayed fetal thymic organ culture repopulation kinetics that more closely approximated those seen with freshly isolated CD4(low) cells. These data suggest that Flt-3L serves as a self-renewal or proliferation/expansion signal for CD4(low) cells, while the effect of stem cell factor is more likely to transduce a differentiation signal, resulting in more rapid repopulation at the expense of cell expansion.
Induction of IL-4-producing CD4+ T cells by antigenic peptides altered for TCR binding.
Authors: Tao X , Grant C , Constant S , Bottomly K 
Published: December 2022

Abstract
The adaptive immune responses to foreign Ags are primarily regulated by the cytokines produced by CD4 T cells. The generation of distinct cytokine-producing T cell subsets has been shown to be influenced by a number of factors, including cytokines, different types of APCs, and the amounts of priming Ag. We have previously reported that the affinity of an antigenic peptide for its presenting MHC class II molecules and that different doses of Ag peptide affect the outcome of the functional CD4 T cell response. In the current study, we further examined the impact of the affinity of an antigenic peptide for its TCR on CD4 T cell priming. We generated a panel of Ag peptide variants mutated at positions known to be critical for binding to a well-characterized TCR (known as altered peptide ligands, or APLs). Compared with the WT peptide, these APLs are defective in stimulating the proliferative responses of T cells. However, they can effectively prime in vitro naive CD4 T cells for differentiation into both Th1-like and Th2-like cells. In contrast, the WT peptide primes only for IFN-gamma-producing Th1-like cells. Using highly purified dendritic cells as APCs to present the APL or WT peptide leads to the same pattern of priming as using total splenic APCs. These results indicate that priming by APLs for both IL-4 production and IFN-gamma production does not require two different types of APCs. In summary, our data indicate that APL can directly stimulate naive CD4 T cells to become Th2 effector cells.
Functional characterization of the interaction between the superantigen staphylococcal enterotoxin A and the TCR.
Authors: Antonsson P , Wingren A G , Hansson J , Kalland T , Varga M , Dohlsten M 
Published: December 2022

Abstract
In this report, we show that despite an overall amino acid residue identity of more than 80% between the staphylococcal enterotoxins (SE) A and E, these proteins markedly differ in their absolute requirement for the MHC class II during T cell activation. The superantigens were produced as C215Fab-SE fusion proteins and analyzed for their ability to activate T cells in a MHC class II-independent manner, using C215 Ag expressing cell lines as pseudo super-APCs. C215Fab-SEA, but not C215Fab-SEE, induced T cell cytotoxicity and proliferation in these MHC class II-independent systems. Introduction of a region from SEA, comprising amino acids 20-27, to SEE transferred the ability to engage T cells in the absence of MHC class II. Analysis of the Vbeta specificity of the chimeric SEA/SEE molecules and a panel of SEA mutants demonstrated that the site for TCR interaction covers the edge surrounding the shallow cavity on top of the SEA molecule.
Two-hybrid cloning of a gene encoding TNF receptor-associated protein 2, a protein that interacts with the intracellular domain of the type 1 TNF receptor: identity with subunit 2 of the 26S protease.
Authors: Dunbar J D , Song H Y , Guo D , Wu L W , Donner D B 
Published: December 2022

Abstract
A protein that binds the intracellular domain of the type 1 TNFR (TNFR-1IC) has been identified by two-hybrid cloning. The 97-kDa TNFR-associated protein, TRAP2, shows sequence identity with internal amino acid sequences from subunit 2 of the 26S protease. TRAP2 antiserum recognizes subunit 2 of the 26S protease, which is consistent with the identity of these proteins. TRAP2 antiserum interacted with the 97-kDa protein in HeLa cell lysates and cytosol, the latter observation showing that TRAP2 resides in the same cellular compartment as TNFR-1IC. A fusion of glutathione-S-transferase and TNFR-1IC (GST-TNFR-1IC) precipitated TRAP2 from a HeLa cell lysate; conversely, GST-TRAP2 precipitated TNFR-1 from such a lysate. These observations show that the proteins interact in the cellular milieu. After in vitro transcription/translation and 35S labeling, TRAP2 was precipitated from a cellfree system by GST-TNFR-1IC, showing that TNFR-1IC and TRAP2 interact directly. TRAP2 was also precipitated from the cellfree translation system by a GST fusion containing the N-terminal half of TNFR-1IC, but not by a GST fusion containing the C-terminal half of TNFR-1IC that contains a "death domain" that plays an obligatory role in signaling cytotoxicity. The ability of deletion mutants of TNFR-1IC to interact with TRAP2 was tested using the two-hybrid system. This also showed that the amino acid sequences that mediate binding reside outside of the death domain in TNFR-1IC. The demonstration that a subunit of the 26S protease binds TNFR-1 may identify a novel TNF-signaling pathway.