bluuebunny/crossref_metadata_2025_split huggingface dataset:
Repeated Intratracheal Instillations of Nonreplicating Adenovirus 2 Vector Attenuate CTL Responses and IFN-γ Production
Authors: Pascual David W. , Walters Nancy , Hillemeyer Peter 
Published: January 2023

AbstractThe proposed usage of replication-deficient adenovirus (Ad) vectors for corrective gene therapy or for mucosal immunization has been limited in part by the host reactivity to the Ad vector, thus limiting repeated Ad instillations. We have recently shown that the reactivity to the Ad vector is in large part due to increased CD4+ Th1 and Th2 responses as well as elevated IgG and mucosal IgA responses. It has been recently proposed that the diminution of transgene expression in respiratory epithelia was due to increased CTL reactivity to expressed Ad proteins. Herein, we report that repeated intratracheal delivery of a second generation Ad2 vector into mice results in no detectable CTL activity in freshly isolated lymphoid cells from lungs, lower respiratory lymph nodes, or spleens or after in vitro restimulation. In contrast, a single dose of Ad2 vector did elicit a robust CTL response. This attenuation of CTL activity was long lived and was not affected by macrophage depletion or due to a reduction in CD4+ or CD8+ T cells. Examination of cytokine production via MHC class I or class II restimulation by lymphoid cells from three intratracheally treated mice showed an attenuation in the production of IFN-γ by as much as 110-fold. This reduction in IFN-γ could not be attributed to increased IL-4 or IL-10 production. Thus, this study shows that the CTL response to Ad vectors is attenuated upon repeated administration.
Induction of In Vitro Human Macrophage Anti-Mycobacterium tuberculosis Activity: Requirement for IFN-γ and Primed Lymphocytes
Authors: Bonecini-Almeida M. Gloria , Chitale Sadhana , Boutsikakis Iosif , Geng Jiayuan , Doo Howard , He Suhui , Ho John L. 
Published: January 2023

Abstract
Mycobacterium tuberculosis (Mtb) is the world’s leading infectious cause of mortality. Despite the overwhelming data supporting the critical role of cellular immunity, little is known of the early microbial and immune cell interactions and whether human macrophages can be activated to express anti-Mtb activity. We report the reconstitution of an in vitro system whereby human macrophages express anti-Mtb activity only in coculture with PBL and with IFN-γ. Omission of IFN-γ in the cocultures or Mtb lysate/IFN-γ-primed lymphocytes was associated with high growth of Mtb, high IL-10 and IL-12 p40, nearly undetectable IL-12 p70 levels, and the highest percentages of CD4 and CD8 T cells. In contrast, IFN-γ treatment of cocultures containing Mtb lysate/IFN-γ-primed PBL reduced bacilli count by ∼2.5 log, decreased the production of IL-10 by 5.7-fold, increased IL-12 p70 by ∼50-fold, and reduced the percentages of CD4 and CD8 T cells. Activation of anti-Mtb activity was time and dose dependent. At 2000 U/ml of IFN-γ, bactericidal activity was achieved (10-fold reduction from initial inoculum). Anti-Mtb activity against several strains of M. tuberculosis (H37Ra and H37Rv, and C, a clinical isolate) was observed and was associated with expression of inducible nitric oxide synthase. These data suggest that induction of human macrophage anti-Mtb activity required dual signaling from PBL and IFN-γ. Thus, the development of an in vitro human system may greatly facilitate studies to delineate immune cells, cytokines, and effector functions/genes critical in controlling Mtb. Defining the mechanisms may also provide novel treatment strategies for tuberculosis.
Regulation of Human Mast Cell β-Tryptase: Conversion of Inactive Monomer to Active Tetramer at Acid pH
Authors: Ren Shunlin , Sakai Kentaro , Schwartz Lawrence B. 
Published: January 2023

Abstract
At neutral pH, human mast cell β-tryptase is stabilized in its enzymatically active, tetrameric form by heparin, and resists inhibition by biologic protease inhibitors. After dissociation of β-tryptase from heparin, active tetramers rapidly convert to inactive monomers in an isotonic, neutral pH environment. Although reversible transition states probably exist during this conversion, once inactive monomers form, addition of heparin fails to reconstitute active tetramer at neutral pH. The current study shows that complete reactivation of inactive monomers can occur at acidic pH in a heparin-independent manner. The respective rate-determining steps for formation of tetramer and active enzyme from inactive monomers exhibit second and first order kinetics based on an analysis of initial reaction rates. The optimal pH for tetramer formation and reactivation is about 6, suggesting His residues play a critical role. The optimal ionic strength equivalent is 160 mM NaCl; and the optimal temperature range is 22°C to 37°C. We propose a sequential three-step reactivation process at acidic pH, dimerization of monomers (rate-determining second order step), rapid formation of inactive tetramers, and slow formation of active tetramers (overall rate-determining first order step). Whether reactivation of human β-tryptase occurs at extracellular or intracellular sites, where the pH is acidic in vivo, should be considered.
An Immunodominant Epitope from Mycobacterial 65-kDa Heat Shock Protein Protects Against Pristane-Induced Arthritis
Authors: Thompson Stephen J. , Francis James N. , Siew L. Khai , Webb Ginette R. , Jenner Peter J. , Colston M. Joseph , Elson Christopher J. 
Published: January 2023

Abstract
Previous studies showed that mice with pristane-induced arthritis (PIA) and those protected from the disease by preimmunization with mycobacterial 65-kDa heat shock protein (hsp65) possess raised immune responses to hsp65. Additionally, T cells from hsp65-protected mice, but not from pristane-injected or normal mice, produced the Th2-associated cytokines IL-4, IL-5, and IL-10 in response to stimulation with hsp65. Here we demonstrate that the specificity of the immune response to hsp65 and related heat shock protein (hsps) differs between protected and PIA mice. T cells from hsp65-protected mice respond to the bacterial hsps tested but not to the mammalian homologue, hsp58. Similarly, they exhibit high serum titers of anti-hsp65 Abs, yet they have virtually undetectable levels of anti-hsp58 IgG. By contrast, both cellular and humoral immune responses are detectable to bacterial and mammalian hsps in mice with PIA. An immunodominant T cell epitope has been identified in hsp65-protected mice corresponding to amino acids 261–271 from hsp65. Immunization of mice, either before or after the induction of arthritis, with this bacterial peptide, but not its mammalian homologue, protects mice from the development of PIA, and protection is associated with the production of Th2-type cytokines. Other experiments revealed that T cells primed with bacterial 261–271 or the mammalian homologue do not cross-react at the proliferative or cytokine level. These results demonstrate that an hsp65 peptide-specific Th2 response confers protection from PIA but do not support the idea that protection is mediated by a cross-reaction with self hsp58 in the joints.
A Comparison of Signaling Requirements for Apoptosis of Human B Lymphocytes Induced by the B Cell Receptor and CD95/Fas
Authors: Graves Jonathan D. , Draves Kevin E. , Craxton Andrew , Krebs Edwin G. , Clark Edward A. 
Published: January 2023

AbstractTo define how the signaling pathways that mediate the B cell receptor (BCR) death pathway differ from those responsible for CD95/Fas-mediated death, we compared the BCR and Fas death pathways in two human B cell lines, B104 and BJAB. Both BCR- and Fas-induced apoptosis are blocked by the peptide cysteine protease inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (ZVAD (mlz)), demonstrating a common requirement caspase activity. Despite this common characteristic, the ability of actinomycin D and cycloheximide to block BCR-induced apoptosis, but not apoptosis induced by Fas cross-linking, suggests that a major difference between these two pathways is their differential requirements for new gene and protein synthesis. BCR- and Fas-mediated apoptosis are both accompanied by activation of stress-activated protein kinase and p38 mitogen-activated protein kinase (MAPK). Activation of both stress-activated protein kinase and p38 MAPK was inhibited by ZVAD (mlz), suggesting the involvement of caspases. To determine the role of p38 MAPK activation in BCR- and Fas-induced apoptosis, we employed SB203580, a specific inhibitor of p38 MAPK. SB203580 inhibited BCR-induced apoptosis, but not apoptosis induced by cross-linking Fas. Furthermore, both actinomycin D and SB203580 inhibited BCR-induced, but not Fas-induced, activation of caspase. Collectively, these findings establish a role for p38 MAPK in BCR-induced apoptosis both upstream and downstream of caspase activity. The p38 MAPK pathway may function to regulate transcriptional or translational events that are critical for BCR-induced apoptosis.
Systemic Administration of Endotoxin Induces Bronchopulmonary Hyperreactivity Dissociated from TNF-α Formation and Neutrophil Sequestration into the Murine Lungs
Authors: Lefort Jean , Singer Monique , Leduc Dominique , Renesto Patricia , Nahori Marie Anne , Huerre Michel , Créminon Christophe , Chignard Michel , Vargaftig B. Boris 
Published: January 2023

Abstract
Bronchopulmonary hyperreactivity (BHR), an increased responsiveness to nonspecific bronchoconstrictor agents, is a well-known characteristic of bronchial asthma. It has been recently suggested that the severity of this disease is related to the endotoxin content of house dust. In the present report, it is shown that the i.p. administration of bacterial LPS to mice is followed by a marked early dose-dependent BHR in response to methacholine. The microscopic examination showed no ultrastructural lesions of the lungs or of the airways, but a marked neutrophil accumulation in the capillaries, as confirmed by an increase of the lung content in the neutrophil enzyme marker myeloperoxidase. In parallel, high levels of TNF-α were found in plasma as well as its transcripts in the lung tissues. Using immunologic (anti-TNF-α and anti-granulocyte Abs), and pharmacologic (dexamethasone and vinblastine) tools, it is demonstrated that BHR is apparently neither related to the presence of neutrophils in the pulmonary microvasculature nor to the synthesis of TNF-α.
Allelic Differences in the Relationship Between Proteasome Activity and MHC Class I Peptide Loading
Authors: Benham Adam M. , Grommé Monique , Neefjes Jacques 
Published: January 2023

AbstractMHC class I molecules are cell surface glycoproteins that play a pivotal role in the response to intracellular pathogens. The loading of MHC class I molecules with antigenic substrates takes place in the endoplasmic reticulum. This requires a functional TAP transporter, which translocates peptides into the endoplasmic reticulum from the cytosol. The generation of antigenic peptides from polypeptide precursors is thought to be mediated in the cytosol by the proteasome. Previously, we have demonstrated that inhibiting the proteasome with the specific covalent inhibitor lactacystin results in a direct reduction of peptide-loaded MHC class I molecules. This indicates that the proteasome is the limiting step in the MHC class I pathway. In this study we use isoelectric focusing to demonstrate that two related MHC class I alleles, HLA-A3 and HLA-A11, as well as HLA-B35 do not follow this behavior. In contrast to other class I alleles expressed by the same cells, these alleles are loaded with peptides and mature normally when proteasome activity is severely inhibited. Our observations highlight a new level of diversity in the MHC class I system and indicate that there are allele-specific differences in the linkage between proteasome activity and MHC class I peptide loading.
Cutting Edge: BASH, A Novel Signaling Molecule Preferentially Expressed in B Cells of the Bursa of Fabricius
Authors: Goitsuka Ryo , Fujimura Yu-ichi , Mamada Hiroshi , Umeda Akiko , Morimura Toshifumi , Uetsuka Koji , Doi Kunio , Tsuji Sachiyo , Kitamura Daisuke 
Published: January 2023

Abstract
The bursa of Fabricius is a gut-associated lymphoid organ that is essential for the generation of a diversified B cell repertoire in the chicken. We describe here a novel gene preferentially expressed in bursal B cells. The gene encodes an 85-kDa protein, designated BASH (B cell adaptor containing SH2 domain), that contains N-terminal acidic domains with SH2 domain-binding phosphotyrosine-based motifs, a proline-rich domain, and a C-terminal SH2 domain. BASH shows a substantial sequence similarity to SLP-76, an adaptor protein functioning in TCR-signal transduction. BASH becomes tyrosine-phosphorylated with the B cell Ag receptor (BCR) cross-link or by coexpression with Syk and Lyn and associates with signaling molecules including Syk and a putative chicken Shc homologue. Overexpression of BASH results in suppression of the NF-AT activation induced by BCR-cross-linking. These findings suggest that BASH is involved in BCR-mediated signal transduction and could play a critical role in B cell development in the bursa.
Cutting Edge: Identification of the 2B4 Molecule as a Counter-Receptor for CD48
Authors: Latchman Yvette , McKay Paul F. , Reiser Hans 
Published: January 2023

AbstractThe CD48 molecule belongs to a subfamily of the Ig superfamily that also includes the CD2, CD58, 2B4, Signaling lymphocyte activation molecule (SLAM), and Ly-9 molecules. Receptor-ligand interactions are known to occur between several members of this family, and these interactions can strengthen cell to cell adhesion. In mice, the CD48 molecule can bind to CD2. To search for additional ligands of murine CD48, we have generated a chimeric fusion protein consisting of the extracellular domain of murine CD48 and the C region of human IgG1. The results of immunofluorescence and immunoprecipitation experiments in which this reagent was used identify the 2B4 molecule as a novel counter-receptor of CD48.
Cutting Edge: Differentiation of Human NK Cells into NK1 and NK2 Subsets
Authors: Peritt David , Robertson Susan , Gri Giorgia , Showe Louise , Aste-Amezaga Miguel , Trinchieri Giorgio 
Published: January 2023

Abstract
Human NK cells cultured in the presence of IL-12 or IL-4 differentiate into cell populations with distinct patterns of cytokine secretion similar to Th1 and Th2 cells. NK cells grown in IL-12 (NK1) produce IL-10 and IFN-γ, whereas NK cells grown in IL-4 (NK2) produce IL-5 and IL-13. Although these NK cell subsets do not differ in cytotoxic activity, NK1 cells express higher levels of cell surface CD95 (Fas) Ag than NK2 cells and are more sensitive to Ab or chemically induced apoptosis. Like Th1 cells, NK1 cells accumulate much higher levels of the IL-12Rβ2-chain mRNA and are significantly more responsive to IL-12 than NK2 cells at the level of activation of STAT4 transcription factor. The identification of NK cell subsets that are analogous to T cell subsets suggests a new role for NK cells in innate inflammatory responses and in their effect on adaptive immunity.
B Cell Responses to a Peptide Epitope. VII. Antigen-Dependent Modulation of the Germinal Center Reaction
Authors: Agarwal Anshu , Nayak Bishnu P. , Rao Kanury V. S. 
Published: January 2023

AbstractGerminal center responses to two analogous peptides, PS1CT3 and G32CT3, that differ in sequence only at one position within the B cell epitopic region were examined. In comparison with peptide PS1CT3, peptide G32CT3 elicited a poor germinal center response. By demonstrating equal facility of immune complexes with IgM and IgG Ab isotypes to seed germinal centers, we excluded differences in isotype profiles of early primary anti-PS1CT3 and anti-G32CT3 Ig as the probable cause. Quantitative differences in germinal center responses to the two peptides were also not due to either qualitative/quantitative differences in T cell priming or variation in the frequency of the early Ag-activated B cells induced. Rather, they resulted from qualitative differences in the nature of B cells primed. Analysis of early primary anti-PS1CT3 and anti-G32CT3 IgMs revealed that the latter population was of a distinctly lower affinity, implying the existence of an Ag affinity threshold that restricts germinal center recruitment of G32CT3-specific B cells. The impediment in anti-G32CT3 germinal center initiation could be overcome by making available an excess of Ag-activated Th cells at the time of immunization. This resulted in the appearance of a higher affinity population of G32CT3-specific B cells that, presumably, are now capable of seeding germinal centers. These data suggest that the strength of a germinal center reaction generated is Ag dependent. At least one regulatory parameter represents the quality of B cells that are initially primed.
In Vivo Anergized CD4+ T Cells Have Defective Expression and Function of the Activating Protein-1 Transcription Factor
Authors: Sundstedt Anette , Dohlsten Mikael 
Published: January 2023

Abstract
The transcription factor activating protein-1 (AP-1) contributes significantly to the regulation of IL-2 gene expression during T cell activation and has been suggested to play a unique role in T cell anergy in vitro. In this study we have used the superantigen staphylococcal enterotoxin A to investigate the regulation of AP-1 in T cell anergy in vivo. Repeated injections of staphylococcal enterotoxin A induce a state of anergy in CD4+ T cells, characterized by reduced expression of IL-2 at mRNA and protein levels. The perturbed IL-2 response in anergic T cells correlated with reduced DNA binding activity of the transcription factors AP-1 and Fos/Jun-containing NF-AT. Using AP-1-luciferase reporter transgenic mice, we now demonstrate the lack of AP-1-dependent transcription. AP-1 activity is controlled by synthesis of its subunits Fos and Jun and by posttranslational phosphorylations. Analysis of Fos and Jun protein levels revealed no major differences in the expression of Jun proteins, but a marked decrease in c-Fos in anergic T cells. Experiments in transgenic mice overexpressing c-Fos (H2-c-fos) showed reconstituted AP-1 DNA binding. In contrast, the AP-1-driven transcription and IL-2 production remained suppressed. The Jun N-terminal kinase is known to play a critical role in regulating AP-1 trans-activation. Analyses of Jun N-terminal kinase demonstrated normal protein amounts, but reduced enzymatic activity, in anergic compared with activated CD4+ T cells. This suggests that in vivo anergized T cells have defects in the AP-1 pathway due to both reduced protein expression and perturbed posttranslational modifications.
Somatic Mutation in the Neonatal Mouse
Authors: Giorgetti Carol A. , Press Joan L. 
Published: January 2023

AbstractSeveral mechanisms that diversify the adult immune repertoire, such as terminal deoxynucleotidyl transferase-dependent N region addition, are not available to the neonatal mouse. One important process that contributes to protective immunity in the adult is somatic mutation, which plays a major role in the generation of high affinity memory B cells. It is not clear whether B cells in the neonatal mouse can activate the somatic mutation machinery. To investigate this, we immunized neonates with poly(l-Tyr,l-Glu)-poly-d, l-Ala–poly-l-Lys complexed with methylated BSA, or (4-hydroxy-3-nitrophenyl)acetyl coupled to chicken γ-globulin. Eight to fourteen days after priming, V(D)J rearrangements of known VH genes (VHSM7 family) were screened for mutations using a temperature-melt hybridization assay and oligonucleotide probes specific for complementarity-determining regions I and II; possible mutations were confirmed by sequence analysis. More mutations per sequence were found in heavy chains from neonates immunized with (4-hydroxy-3-nitrophenyl)acetyl coupled to chicken γ-globulin than in those from neonates immunized with poly(l-Tyr, l-Glu)-poly-d,l-Ala-poly-l-Lys complexed with methylated BSA. Mutations were found in heavy chains lacking N regions, suggesting that B cells of the putative fetal lineage can somatically mutate and diversify an initially limited repertoire. Since neonates immunized as early as 1 or 2 days after birth had mutations, the somatic mutation machinery can be activated soon after birth, suggesting that early vaccination should result in affinity maturation and protective immunity in the neonate.
OPG/FDCR-1, a TNF Receptor Family Member, Is Expressed in Lymphoid Cells and Is Up-Regulated by Ligating CD40
Authors: Yun Theodore J. , Chaudhary Preet M. , Shu Geraldine L. , Frazer J. Kimble , Ewings Maria K. , Schwartz Stephen M. , Pascual Virginia , Hood Leroy E. , Clark3 Edward A. 
Published: January 2023

Abstract
We have cloned a TNFR family member from a follicular dendritic cell (FDC)-like cell line, FDC-1. This molecule, FDC-derived receptor-1 (FDCR-1), is identical to osteoprotegerin (OPG), a soluble cytokine that regulates osteoclast differentiation. Recently, OPG/FDCR-1 has been characterized as a second receptor for receptor activator of NF-κB ligand (RANKL)/TNF-related activation-induced cytokine (TRANCE), a primarily T-cell restricted TNF family member that augments dendritic cell (DC) function. In this report, we demonstrate that OPG/FDCR-1 is membrane bound on the surface of transfected baby hamster kidney (BHK) and untransfected FDC-1 cells. We also found a restricted OPG/FDCR-1 expression pattern in lymphoid cells, specifically in B cells, DCs and FDC-enriched fractions, which in B cells and DCs is up-regulated by CD40 stimulation. Because OPG/FDCR-1 shares some properties with RANK, the first RANKL/TRANCE receptor, we discuss how the balance between RANK and OPG/FDCR-1 expression could influence immune responses and, ultimately, germinal center formation.
Isolated Human Germinal Center Centroblasts Have an Intact Mismatch Repair System
Authors: Park Kyoungsook , Kim Jaehoon , Kim Han-Soo , Shin Hyun S. 
Published: January 2023

Abstract
Ig somatic hypermutation contributes to the generation of high-affinity Abs that are essential for efficient humoral defense. The presence of multiple point mutations in rearranged Ig V genes and their immediate flanking sequences suggests that the DNA repair system may not be working properly in correcting point mutations introduced to the restricted region of Ig genes. We examined the DNA repair functions of germinal center (GC) centroblasts, which are the cells in which ongoing Ig hypermutation takes place. We found that GC centroblasts express all known components of the human DNA mismatch repair system, and that the system corrects DNA mismatches in a strand-specific manner in vitro. We conclude that general suppression of mismatch repair at the cellular level does not occur during somatic hypermutation.