bluuebunny/crossref_metadata_2025_split huggingface dataset:
Unique cytokine production profile of anergic human T cells in SCID-hu mice after staphylococcal enterotoxin B administration.
Authors: Schols D , Jones D , Roncarolo M G 
Published: December 2022

Abstract
Severe combined immunodeficient mice transplanted with human organs (SCID-hu mice), provide a unique in vivo model for studying human intrathymic T cell selection and development of tolerance. In vivo administration of staphylococcal enterotoxin B (SEB) to SCID-hu mice causes intrathymic clonal deletion of SEB-specific V beta+ T cells that occurs already at the immature CD4+8+ double positive stage. The expression of activation markers such as CD25, CD71, and HLA-DR was specifically increased on V beta+ T cells responding to SEB. The remaining SEB-specific human T cells that had not been deleted in vivo failed to proliferate when rechallenged with SEB in vitro. These SEB-specific T cells that were rendered anergic in vivo had a unique cytokine production profile. They failed to produce IL-2, which correlated with the lack of proliferation of these cells. In addition, they failed to produce TNF-alpha. However, the anergized T cells synthesized considerable amounts of IFN-gamma, granulocyte-macrophage CSF and IL-10 after SEB stimulation. This clonal anergy can be completely reversed in vitro by stimulating the SEB-specific cells in the presence of exogenous IL-2 or by triggering of the CD28/CTLA-4 activation pathway. Under these stimulation conditions, anergic T cells produced levels of IL-2 and TNF-alpha that were comparable to their non-anergized counterparts, whereas the levels of granulocyte-macrophage CSF, IL-10 and IFN-gamma production were even higher. Collectively, these data demonstrate that in vivo administration of SEB to SCID-hu mice leads to activation, deletion, and anergy of SEB-specific human thymocytes and that the production of IL-2 and TNF-alpha is selectively switched off in these anergic T cells.
In vitro antibody maturation. Improvement of a high affinity, neutralizing antibody against IL-1 beta.
Authors: Jackson J R , Sathe G , Rosenberg M , Sweet R 
Published: December 2022

Abstract
Abs as therapeutic antagonists should be of relatively high affinity to effectively neutralize their target Ag. Typically, Abs with nanomolar affinities to protein Ags can be obtained in vivo, however, this may not be the upper limit of affinity because the biologic process attempts to optimize Ab function, of which affinity is only one component. SK48, a high affinity neutralizing murine Fab against human IL-1 beta was used to explore the nature of Ab-Ag interactions and the potential for further affinity improvement in vitro using mutagenesis and selection via phage display. The codons of six amino acids in the third complementarity-determining region of the heavy chain (CDR3-H) were both individually and combinatorialy randomized and the resultant libraries were screened for IL-1 binding phenotype. Mutations that reduced affinity suggested that both the backbone conformation of the CDR3 loop and certain side chains are essential for binding, yet alterations to the canonical salt bridge residues at the base of the loop had minimal effect on affinity. Four rounds of selection of the phage Ab libraries on immobilized IL-1 beta gave predominantly the wild-type sequence, indicating efficient affinity maturation of this CDR in vivo. However, a twofold improvement in affinity was observed for both single and double amino acid changes at two positions in the middle of the CDR. Moreover, a 10-fold increase in affinity for IL-1 beta was achieved by combining two of the phage-selected single amino acid substitutions in CDR3-H, thereby demonstrating that a significant improvement in affinity can be achieved through CDR mutagenesis, even in a matured Ab. The increased affinity of this Fab did not, however, enhance its neutralizing activity in vitro.
Elicitation of a systemic and protective anti-melanoma immune response by an IL-2-based vaccine. Assessment of critical cellular and molecular parameters.
Authors: Zatloukal K , Schneeberger A , Berger M , Schmidt W , Koszik F , Kutil R , Cotten M , Wagner E , Buschle M , Maass G 
Published: December 2022

Abstract
We have established a model for the immunologic rejection of melanoma cells. Using a receptor-mediated, adenovirus-augmented gene delivery system (transferrinfection) we have shown that, upon transfection with an IL-2 gene construct, MHC class I+/class II- murine M-3 cells lose their tumorigenicity in both athymic and euthymic mice. More importantly, we found that these melanoma cells, which produce high levels of IL-2, can be used to induce a long-lasting anti-tumor immune response in syngeneic euthymic DBA/2 mice but not in athymic animals. This immune response, which can also be elicited by coadministration of nonmodified, irradiated M-3 cells and IL-2-transduced fibroblasts, results in the rejection of a subsequent challenge with M-3 cells or, in the elimination of preexisting M-3 cancer cell deposits. We found that transfer of T cell-enriched, but not of T cell-depleted, splenocytes from immunized mice conferred protection against M-3 cells, but not against unrelated KLN 205 cancer cells. Transfer of either CD4+ or CD8+ T cells led to only partial protection against challenge with wild-type M-3 cells. Our further observations that T cell-enriched, but not T cell-depleted splenocytes of immunized animals are capable of tumor-specific lytic activity and that this activity resides in the CD8+ cell population are compatible with the assumption that MHC class I-restricted T cell cytotoxicity is a biologically relevant effector mechanism in this model. That other mechanisms also contribute to melanoma cell destruction is evidenced by the presence of large numbers of macrophages and granulocytes in addition to T cells at the challenge sites of immunized mice.
Differences in binding of glucocorticoid receptor to DNA in steroid-resistant asthma.
Authors: Adcock I M , Lane S J , Brown C R , Peters M J , Lee T H , Barnes P J 
Published: December 2022

Abstract
Although glucocorticosteroids are a very effective treatment for asthma and other chronic inflammatory diseases, a small proportion of patients are resistant to their therapeutic effects. The molecular mechanism for this steroid resistance is unclear. Steroid resistance cannot be explained by pharmacokinetic mechanisms, by a defect in the binding of steroids to glucocorticoid receptors, nor by defective nuclear translocation of this receptor, thereby suggesting that the molecular abnormality lies distal to nuclear translocation. We examined the ability of nuclear translocated glucocorticoid receptors to bind to their DNA binding sites (GRE) using electrophoretic mobility shift assays in PBMC from patients with steroid-sensitive and steroid-resistant asthma. The binding of the glucocorticoid receptor to DNA in these patients was also studied using Scatchard analysis. Dexamethasone induced a significant rapid and sustained twofold increase in GRE binding in PBMCs from steroid-sensitive asthmatic patients and nonasthmatic individuals, but this was markedly reduced in steroid-resistant asthmatic patients. Scatchard analysis of glucocorticoid receptor-GRE binding showed no change in binding affinity but did show a reduced number of receptors available for DNA binding in the steroid-resistant patients. These results suggest that the ability of the glucocorticoid receptor to bind to GRE is impaired in steroid-resistant patients because of a reduced number of receptors available for binding to DNA.
A regulatory role for astrocytes in HIV-1 encephalitis. An overexpression of eicosanoids, platelet-activating factor, and tumor necrosis factor-alpha by activated HIV-1-infected monocytes is attenuated by primary human astrocytes.
Authors: Nottet H S , Jett M , Flanagan C R , Zhai Q H , Persidsky Y , Rizzino A , Bernton E W , Genis P , Baldwin T , Schwartz J 
Published: December 2022

Abstract
HIV-1-infected brain macrophages participate in neurologic dysfunction through their continual secretion of neurotoxins. We previously demonstrated that astroglial cells activate HIV-1-infected monocytes to produce such neurotoxic activities. In this study, the mechanism underlying these monocyte secretory activities was unraveled and found dependent on HIV-1's ability to prime monocytes for activation. LPS stimulation of HIV-1-infected monocytes resulted in an overexpression of eicosanoids, platelet-activating factor (PAF), and TNF-alpha. This was dependent on the level of HIV-1 infection and monocyte stimulation. Cell to cell interactions between activated virus-infected monocytes and primary human astrocytes reduced monocyte secretions. The capacity of astrocytes to deactivate monocytes was, notably, TGF-beta independent. Although astrocytes constitutively produced latent TGF-beta 2, HIV-1-infected monocytes neither affected TGF-beta 2 production nor converted it into a bioactive molecule. Furthermore, addition of rTGF-beta 1 or rTGF-beta 2 or its Abs to LPS-stimulated monocyte-astrocyte mixtures had no effect on monokine production. In contrast, addition of rIL-10 to LPS-stimulated monocytes produced a dose-dependent decrease in TNF-alpha. IL-10 mRNAs were detected in monocytes, but not astrocytes, following LPS treatment. These results suggest that macrophage activation, a major component of HIV-1 infection in the brain, precipitates neuronal injury by causing virus-infected cells to synthesize neurotoxins. The neurotoxins produced by monocytes are then regulated by astrocytes. Astrocytes therefore, can play either positive or negative roles for disease depending on prior macrophage activation. These findings begin to unravel the cellular control mechanisms that influence cognitive and motor dysfunctions in HIV-1-infected individuals.
TCR gene usage in experimental autoimmune myasthenia gravis pathogenesis. Usage of multiple TCRBV genes in the H-2b strains.
Authors: Wu B , Shenoy M , Goluszko E , Kaul R , Christadoss P 
Published: December 2022

Abstract
Experimental autoimmune myasthenia gravis (EAMG) is an Ab-mediated autoimmune disease. The pathogenic auto-antibody production depends on the activation of CD4+ cells after their TCR interact with dominant T cell epitopes within acetylcholine receptor (AChR) in the context of the MHC class II molecule. In vitro analysis suggested that the TCRBV6 was the predominant TCR that recognized AChR and one of the dominant epitopes, alpha 146-162, in C57BL6 (B6, H-2b) mice. However, in vivo depletion of TCRBV6 cells in H-2b mice by anti-TCRBV6 mAb neither suppressed the in vitro immune response to AChR nor prevented development of EAMG. Moreover, B10.TCRc (H-2b) strain with a genomic deletion of TCRBV genes including TCRBV6, and B10.V beta 8.2 transgenic mice with a restricted TCRBV8S2 T cell repertoire, responded to AChR, alpha 146-162, and developed EAMG after immunizations with AChR/CFA. These data suggest that more than one TCRBV-bearing cell having the affinity for AChR-dominant peptides is involved in pathogenesis. Therefore, depletion of a single TCRBV (e.g., TCRBV6) with mAb may not be sufficient to completely suppress the response to AChR and development of EAMG. However, if a similar amino acid sequence in the TCR-VDJ (e.g., CDR3) region among different TCRBV gene(s) could be involved in recognizing the dominant AChR epitope(s), then motif-specific mAb reactive to the common motif within the VDJ region of different TCR could be used to eliminate the T cell clones involved in EAMG.
T cell receptor V alpha 4 is expressed by a subpopulation of V beta 6 T cells that respond to the bacterial superantigen staphylococcal enterotoxin B.
Authors: Borrero H , Donson D , Cervera C , Rexer C , Macphail S 
Published: December 2022

Abstract
A subpopulation of murine minor lymphocyte-stimulating locus Ag (Mls)-1a-responsive, TCR V beta 6-expressing T hybrids was responsive to the superantigenic bacterial toxin, staphylococcal enterotoxin B (SEB), presented by murine MHC class II molecules. Comparative functional and surface marker analyses showed that this heterogeneity was not caused by nonspecific effects. It seemed, therefore, that the ability of the TCR V beta 6 T hybrids to respond to SEB was regulated by non-V beta TCR elements. cDNA sequencing analyses of the TCR beta- and alpha-chains expressed by the V beta 6 T hybrids and a beta-chain cDNA gene transfer experiment indicated that although J beta, CDR3 beta, J alpha, and CDR3 alpha were not the relevant elements, SEB responsiveness did correlate with the expression of two different members of the V alpha 4 family. A functional analysis of a separate panel of V alpha 4 T hybrids expressing different V beta elements, including V beta 6, specifically associated SEB responsiveness with the V alpha 4, V beta 6 combination. Overall, the data obtained with both panels of T hybrids showed SEB responsiveness in five out of five V beta 6, V alpha 4 T hybrids, and not in either 11 V beta 6 T hybrids expressing a variety of other V alpha elements or four V alpha 4 T hybrids expressing V beta 11, 14, and 15 elements. Thus, our experiments show for the first time a specific, positive qualitative effect of a defined V alpha element on responsiveness to a bacterial superantigen. Finally, this study has identified four new V alpha elements.
Antagonistic interactions among T cell subsets of old mice revealed by limiting dilution analysis.
Authors: Dozmorov I M , Kalinichenko V V , Sidorov I A , Miller R A 
Published: December 2022

Abstract
When CD4 spleen cells from old (but not young) mice are tested for Con A-induced proliferation in limiting dilution assays, the dose response curve shows a nonlinear relationship. We interpret these observations using a two-cell model, in which proliferation of one cell type (LPC1) can be blocked by a second cell type (LPC2), which can itself generate detectable proliferation only at high multiplicities. The two-cell model accounts for several observations: 1) the variation in curve shape as a function of incubation time; 2) the skewed distribution of wells scored as "negative" in cultures of old splenocytes; and 3) the initially antagonistic effects of old splenocytes titrated into cultures containing fixed numbers of young responders. To provide a further test of the two-cell model, ionomycin-resistant (CaR) and ionomycin-sensitive (CaS) cells were separated using a Percoll/ionomycin gradient. The CaR preparation, shown previously to consist largely of memory T cells, showed the dose curve predicted for the LPC2 cell type, whereas the CaS (naive) cells showed the single-hit kinetics postulated for LPC1 cells. Furthermore, mixtures of CaR and CaS cells from young mice reproduced the zigzag dose curve characteristically produced by unseparated cells from old mice. These data suggest that the spleens of both young and old mice contain two kinds of Con A-responsive CD4 cell: one that proliferates vigorously, and a second, calcium ionophore-resistant type that proliferates less well, that can interfere with proliferation of the first cell type, and whose frequency increases with age.
In situ detection and characterization of apoptotic thymocytes in human thymus. Expression of bcl-2 in vivo does not prevent apoptosis.
Authors: Le P T , Maecker H T , Cook J E 
Published: December 2022

Abstract
Apoptosis plays a crucial role in shaping the T cell repertoire during T cell development in the thymus. The observed disappearance in the thymus of CD4+ CD8+ thymocytes with a specific TCR, and the lack of CD4+ or CD8+ single positive mature cells expressing the same TCR specificity in the periphery have led to the conclusion that deletion occurs at the CD4+ CD8+ double positive stage; however, there is no direct evidence demonstrating apoptotic CD4+ CD8+ cells in situ. Apoptosis of thymocytes in situ at other stages of T cell development has also not been reported. Using three-color immunofluorescence and flow cytometric assays on frozen human thymic tissue and freshly isolated human thymocytes respectively, we directly identify CD4+ CD8+ and CD4- CD8- thymocytes in newborn human thymus that contain intracellular fragmented DNA and are therefore apoptotic. We determine that 75% of the apoptotic thymocytes are CD4+ CD8+ double positive apoptotic thymocytes, and interestingly, that 13% are CD4- CD8- double negative thymocytes. The majority of apoptotic thymocytes in situ are detected at the cortical-medullary junction; however, apoptotic thymocytes are also found scattered throughout the cortex. Furthermore, we determine that within the apoptotic thymocyte population, 54% express the apoptotic regulatory protein bcl-2 in vivo, whereas 32% are bcl-2 negative. Thus, our in vivo data directly demonstrate that both CD4+ CD8+ and CD4- CD8- human thymocytes die in situ via an apoptotic process, and that expression of the bcl-2 protein in situ does not prevent immature thymocytes from apoptosis.
Engagement of the antigen-receptor on immature murine B lymphocytes results in death by apoptosis.
Authors: Norvell A , Mandik L , Monroe J G 
Published: December 2022

Abstract
During their development B lymphocytes pass through a maturational stage in which encounter with Ag leads to tolerance rather than activation. At least four mechanisms for achieving B cell tolerance have been reported: deletion, anergy, receptor editing, and competition for follicular niches. Although turnover rates for immature B cells in the adult mouse bone marrow and several transgenic model systems suggest that a major process contributing to negative selection of B cells is deletion, a detailed study of the negative effect of Ag-receptor engagement on primary, immature B cell survival has never been undertaken. We have utilized an in vitro culture system to determine whether cross-linking sIgM on tolerance-susceptible sIgM+IgD- B cells results in deletion by apoptosis. In contrast to the effect of sIgM cross-linking on mature splenic B cells, treatment of immature, bone marrow-derived B cells results in significant levels of apoptotic death. Ag receptor-mediated apoptosis is detectable by 14 h after sIgM engagement. Moreover, IL-4 and cycloheximide, which have previously been shown to prevent B cell tolerance induction, specifically block the sIgM-induced apoptosis observed in the immature B cells. Similarly, immature B cells from the neonatal spleen are also susceptible to apoptosis after sIgM cross-linking, although they manifest somewhat higher levels of unstimulated apoptosis as compared with bone marrow-derived B cells. These studies are the first detailed demonstration of Ag receptor-mediated apoptosis of primary immature stage B lymphocytes.
Antigen processing in vivo and the elicitation of primary CTL responses.
Authors: Restifo N P , Bacík I , Irvine K R , Yewdell J W , McCabe B J , Anderson R W , Eisenlohr L C , Rosenberg S A , Bennink J R 
Published: December 2022

Abstract
CD8+ T lymphocytes (TCD8+) play an important role in cellular immune responses. TCD8+ recognize MHC class I molecules complexed to peptides of 8 to 10 residues derived largely from cytosolic proteins. Proteins are generally thought to be fragmented in the cytoplasm and delivered to nascent class I molecules in the endoplasmic reticulum (ER) by a peptide transporter encoded by the MHC. To explore the extent to which TCD8+ induction in vivo is limited by proteolysis or peptide transport into the ER, mice were immunized with recombinant vaccinia viruses containing mini-genes encoding antigenic peptides (bypassing the need for proteolysis), or these peptides with a NH2-terminal ER insertion sequence (bypassing the requirements for both proteolysis and transport). Additionally, mice were immunized with recombinant vaccinia viruses encoding rapidly degraded fragments of proteins. We report that limitations in induction of TCD8+ responses vary among Ags: for some, full length proteins are as immunogenic as other forms tested; for others, maximal responses are induced by peptides or by peptides targeted to the ER. Most importantly, in every circumstance examined, targeting peptides to the ER never diminished, and in some cases greatly enhanced, the TCD8+ immune response and provide an important alternative strategy in the design of live viral or naked DNA vaccines for the treatment of cancer and infectious diseases.
Identification and analysis of the expression of CD8 alpha beta and CD8 alpha alpha isoforms in chickens reveals a major TCR-gamma delta CD8 alpha beta subset of intestinal intraepithelial lymphocytes.
Authors: Tregaskes C A , Kong F K , Paramithiotis E , Chen C L , Ratcliffe M J , Davison T F , Young J R 
Published: December 2022

Abstract
Expression screening has been used to clone cDNAs encoding the alpha- and beta-chains of chicken CD8. Amino acid sequence similarities with the mammalian sequences were about 30%. Many amino acid residues of structural or functional importance were more highly conserved, as were the overall structures of both chains. Like human CD8 alpha, the chicken alpha-chain lacked sites for N-linked glycosylation, but the beta-chain contained three such sites. In COS cells transfected with CD8 beta cDNA, surface expression of the beta-chain was dependent on co-transfection of the alpha-chain cDNA, indicating that, as in mammals, chicken CD8 can be expressed as a CD8 alpha alpha homodimer or as a CD8 alpha beta heterodimer. Immunofluorescence analysis with mAbs that were shown to identify the CD8 alpha- and CD8 beta-chains revealed that the vast majority of the CD8+ cells in the thymus, spleen, and blood of adult chickens express both CD8 alpha- and CD8 beta-chains. However, a relatively large proportion of the CD8+ TCR-gamma delta cells in the spleens of embryos and young chicks express only the alpha-chain of CD8. Among intestinal epithelial lymphocytes the major CD8+ T cell populations present in mice are conserved, but there is a population of TCR-gamma delta CD8 alpha beta cells that is not found in rodents. This observation is important in interpretation of experiments examining the pathways of development of intestinal intraepithelial lymphocytes in chickens.
Vaccination routes that fail to elicit protective immunity against Schistosoma mansoni induce the production of TGF-beta, which down-regulates macrophage antiparasitic activity.
Authors: Williams M E , Caspar P , Oswald I , Sharma H K , Pankewycz O , Sher A , James S L 
Published: December 2022

Abstract
C57BL/6 mice immunized intradermally (i.d.) with bacillus Calmette Guerin (BCG) plus killed skin-stage schistosomula are protected against subsequent infection with Schistosoma mansoni, whereas immunization by i.v. or i.m. routes is not protective. Moreover, previous immunization via the nonprotective i.v. route interfered with the ability to subsequently induce protection by i.d. vaccination, suggesting that inhibitory responses are invoked. Given the evidence that activated macrophages (M phi) play a role as effector cells in protection against schistosomiasis, we investigated the ability of spleen cells from protected and nonprotected immunized mice to produce M phi activating and deactivating cytokines. Exposure to supernatant fluids (SNs) from Ag stimulated spleen cells of i.d., but not i.v. or i.m., immunized mice activated inflammatory M phi for in vitro killing of schistosome larvae, through a mechanism dependent on both IFN gamma and TNF-alpha. No evidence was observed for the preferential induction of the M phi activating Th1 cytokines IFN-gamma and IL-2 in i.d. immunized mice, nor did spleen cells from nonprotected animals produce higher levels of the Th2 associated cytokines IL-4 and IL-10, which are known to prevent M phi activation. TGF-beta was, however, detected in SNs from unprotected mice. Moreover, the M phi inhibitory activity detected in these SNs was heat stable and neutralized by anti-TGF-beta Abs, suggesting that production of TGF-beta is at least partially responsible for the failure of i.m. and i.v. immunized mice to develop immunity to S. mansoni. Thus, the induction of down-regulatory cytokines may be an important factor limiting the efficacy of certain vaccination protocols.