bluuebunny/crossref_metadata_2025_split huggingface dataset:
Bcl-2 Controls Dendritic Cell Longevity In Vivo
Authors: Nopora Adam , Brocker Thomas 
Published: April 2014

Abstract
Dendritic cells (DC) were found to down-regulate Bcl-2 protein upon maturation in vivo. Because Bcl-2 has been shown to exert anti-apoptotic functions, down-regulation of Bcl-2 could be a mechanism by which DC longevity is controlled. To dysregulate this potential control system and to study the role of Bcl-2 in DC, we expressed human Bcl-2 under control of the murine CD11c-promoter as a transgene specifically in DC and show that DC frequencies and numbers increase in transgenic mice. In vivo bromodeoxyuridin, as well as adoptive, DC transfer studies show that the relative turnover/survival of mature Bcl-2 transgenic DC is increased. This had a direct impact on CD4+ T cell, as well as humoral immune, responses, which were elevated in transgenic animals. When Bcl-2 transgenic DC were used as DC vaccines, they induced 2- to 3-fold greater expansion of Ag-specific CTL, and stronger in vivo cytotoxicity. Overall, these data indicate that down-regulation of Bcl-2 controls DC longevity, which in turn directly regulates immune responses and the efficacy of DC when used as vaccines.
Active Form of Notch Imposes T Cell Fate in Human Progenitor Cells
Authors: De Smedt Magda , Reynvoet Katia , Kerre Tessa , Taghon Tom , Verhasselt Bruno , Vandekerckhove Bart , Leclercq Georges , Plum Jean 
Published: April 2014

AbstractThe crucial role of Notch signaling in cell fate decisions in hematopoietic lineage and T lymphocyte development has been well established in mice. Overexpression of the intracellular domain of Notch mediates signal transduction of the protein. By retroviral transduction of this constitutively active truncated intracellular domain in human CD34+ umbilical cord blood progenitor cells, we were able to show that, in coculture with the stromal MS-5 cell line, depending on the cytokines added, the differentiation toward CD19+ B lymphocytes was blocked, the differentiation toward CD14+ monocytes was inhibited, and the differentiation toward CD56+ NK cells was favored. The number of CD7+cyCD3+ cells, a phenotype similar to T/NK progenitor cells, was also markedly increased. In fetal thymus organ culture, transduced CD34+ progenitor cells from umbilical cord blood cells or from thymus consistently generated more TCR-γδ T cells, whereas the other T cell subpopulations were largely unaffected. Interestingly, when injected in vivo in SCID-nonobese diabetic mice, the transduced cells generated ectopically human CD4+CD8+ TCR-αβ cells in the bone marrow, cells that are normally only present in the thymus, and lacked B cell differentiation potential. Our results show unequivocally that, in human, Notch signaling inhibits the monocyte and B cell fate, promotes the T cell fate, and alters the normal T cell differentiation pathway compatible with a pretumoral state.
Regulation of Immunoproteasome Subunit Expression In Vivo Following Pathogenic Fungal Infection
Authors: Barton Lance F. , Cruz Miguel , Rangwala Reshma , Deepe George S. , Monaco John J. 
Published: April 2014

AbstractThe proteasome catalytic β subunits LMP2, LMP7, and MECL-1 and two proteasome activator proteins, PA28 α and β, are induced following exposure to IFN-γ in vitro. Induction of these immunosubunits and the PA28 α/β hetero-oligomer alters proteasome catalytic functions and specificity and enhances production of certain MHC class I epitopes. We sought to determine whether and to what extent proteasome subunit composition is regulated in vivo and to elucidate the mechanisms of such regulation. We analyzed basal expression levels of these inducible genes in normal, IFN-γ-deficient, and Stat-1-deficient mice. Mice of all three genotypes display constitutive expression of the immunosubunits and PA28, demonstrating that basal expression in vivo is independent of endogenous IFN-γ production. However, basal expression levels are reduced in Stat-1−/− mice, demonstrating a role for Stat-1 independent of IFN-γ signaling. To demonstrate that IFN-γ can induce these genes in vivo, mice were infected with Histoplasma capsulatum. Elevated expression of these genes followed the same time course as IFN-γ expression in infected mice. IFN-γ-deficient mice did not display elevated protein expression following infection, suggesting that other inflammatory cytokines produced in infected mice are unable to influence proteasome expression. Cytokines other than IFN-γ also failed to influence proteasome gene expression in vitro in cell lines that had no basal expression of LMP2, LMP7, or MECL-1. Thus, both in vitro and in vivo data demonstrate that IFN-γ is essential for up-regulation, but not constitutive expression, of immunoproteasome subunits in mice.
Gene Conversion-Like Sequence Transfers Between Transgenic Antibody V Genes Are Independent of RAD54
Authors: D’Avirro Nicole , Truong David , Luong Michael , Kanaar Roland , Selsing Erik 
Published: April 2014

Abstract
Homology-based Ig gene conversion is a major mechanism for Ab diversification in chickens and the Rad54 DNA repair protein plays an important role in this process. In mice, although gene conversion appears to be rare among endogenous Ig genes, Ab H chain transgenes undergo isotype switching and gene conversion-like sequence transfer processes that also appear to involve homologous recombination or gene conversion. Furthermore, homology-based DNA repair has been suggested to be important for somatic mutation of endogenous mouse Ig genes. To assess the role of Rad54 in these mouse B cell processes, we have analyzed H chain transgene isotype switching, sequence transfer, and somatic hypermutation in mice that lack RAD54. We find that Rad54 is not required for either transgene switching or transgene hypermutation. Furthermore, even transgene sequence transfers that are known to require homology-based recombinations are Rad54 independent. These results indicate that mouse B cells must use factors for promoting homologous recombination that are distinct from the Rad54 proteins important in homology-based chicken Ab gene recombinations. Our findings also suggest that mouse H chain transgene sequence transfers might be more closely related to an error-prone homology-based somatic hypermutational mechanism than to the hyperconversion mechanism that operates in chicken B cells.
Polymorphism at Position 97 in MHC Class I Molecules Affects Peptide Specificity, Cell Surface Stability, and Affinity for β2-Microglobulin
Authors: Smith Ruth A. , Myers Nancy B. , Robinson Melanie , Hansen Ted H. , Lee David R. 
Published: April 2014

AbstractThe two mouse MHC class I alleles, Ld and Lq, share complete amino acid sequence identity except in the α2 domain, where they differ at six positions. Despite their similarity, Lq has a stronger association with β2-microglobulin (β2m), is expressed at higher levels on the cell surface, demonstrates an increased cell surface half-life, and has fewer open forms on the cell surface than Ld. To determine the basis for their phenotypic differences, Ld molecules containing chimeric Ld-Lq α2 domains were characterized, and these analyses implicated residue 97 (LdTrp and LqArg) as the polymorphic site responsible for the disparity in β2m association between the two alleles. Single substitution analysis at this site (LdW97R and LqR97W) confirmed this. Furthermore, the LdW97R mutant molecule has a longer cell surface half-life than either Lq or Ld, and fewer open forms of LdW97R are observed on the cell surface. In addition, both LdW97R and Lq possess decreased binding affinity for the Ld-restricted tum− P91A14–22 peptide compared with Ld. Collectively, these results and the known location of Trp97 in the peptide binding cleft of Ld strongly suggest that the substitution of Arg for Trp97 in Ld alters the peptide binding cleft, increasing its affinity for endogenous peptides, which results in greater cell surface stability and better retention of β2m. Furthermore, these results imply that Trp97 plays an important role in the ability of Ld to efficiently participate in alternative MHC class I Ag presentation pathways.
Identification of a Novel HLA-B60-Restricted T Cell Epitope of the Minor Histocompatibility Antigen HA-1 Locus
Authors: Mommaas Bregje , Kamp Janine , Drijfhout Jan-Wouter , Beekman Nico , Ossendorp Ferry , van Veelen Peter , den Haan Joke , Goulmy Els , Mutis Tuna 
Published: April 2014

AbstractThe polymorphic minor histocompatibility Ag HA-1 locus encodes two peptides, HA-1H and HA-1R, with a single amino acid difference. Whereas the immunogenicity of the HA-1R allele has not yet been shown, the nonameric HA-1H peptide induces HLA-A2-restricted cytotoxic T cells in vivo and in vitro. It is not known whether the mHag HA-1H or HA-1R associates with other HLA class I molecules. Therefore, the polymorphic regions of both HA-1 alleles were analyzed to identify HLA class I binding peptides that are properly processed by proteasomal degradation. Peptide binding analyses were performed for all nonameric HA-1H/R peptides for binding to nine HLA class I molecules with &gt;10% prevalence in the Caucasian population and for seven nonameric/decameric HA-1H/R peptides predicted to bind to HLA-A3, -B14, and -B60. Only the nonameric KECVLH/RDDL and decameric KECVLH/RDDLL peptides showed strong and stable binding to HLA-B60. In vitro digestion of 29-aa-long HA-1 peptides by purified 20S proteasomes revealed proper cleavage at the COOH termini of both HLA-B60 binding HA-1H and HA-1R peptides. In subsequent analyses, dendritic cells pulsed with the nonameric HA-1R peptide did not induce CTLs that recognize the natural HLA-B60/HA-1R ligand. In contrast, dendritic cells pulsed with the nonameric HA-1H peptide induced IFN-γ-secreting T cells specific for the natural HLA-B60/HA-1H ligand in three HLA-B60+ HA-1RR individuals, demonstrating the immunogenicity of the HLA-B60/HA-1H ligand. In conclusion, this study shows a novel HLA-B60-restricted T cell epitope of the minor histocompatibility Ag HA-1 locus.
Peptidic Termini Play a Significant Role in TCR Recognition
Authors: Wang Bo , Sharma Ashawni , Maile Robert , Saad Mohamed , Collins Edward J. , Frelinger Jeffrey A. 
Published: April 2014

AbstractTCR recognition of class I MHC is dependent on the composition of the antigenic peptide and the MHC. Single amino acid substitutions in either the MHC or the peptide may dramatically alter recognition. While the major interactions between TCR and the peptide/MHC complex appear to be focused on the complementarity-determining region (CDR)3, it is also clear from the cocrystal structure of class I MHC and TCR that the amino and carboxyl ends of the peptide may play a role through interactions with the CDR1. In this work we show that gp33 variants substituted at the peptidic termini at the putative CDR1 contact regions show improved recognition in B6 mice. The rank order of recognition is different using the P14 transgenic T cells, suggesting that one reason for improved recognition is a change in the TCR repertoire that recognizes the peptide. However, the affinity of the TCR by some of the peptide/MHC complex with increased recognition is improved, as shown by increased tetramer binding to P14 T cells. These substitutions at the termini of the peptide-binding cleft cause localized conformational changes as seen by changes in mAb binding and crystallographic structures. The different peptide structures also show different conformations in the center of the peptide, but these are shown to be energetically similar and thus most likely have no significance with respect to TCR recognition. Therefore, small conformational changes, localized to the CDR1 contact regions, may play a significant role in TCR recognition.
MHC Allele-Specific Molecular Features Determine Peptide/HLA-A2 Conformations That Are Recognized by HLA-A2-Restricted T Cell Receptors
Authors: Wang Zichun , Turner Richard , Baker Brian M. , Biddison William E. 
Published: April 2014

AbstractThe structures of αβ TCRs bound to complexes of class I MHC molecules and peptide show that the TCRs make multiple contacts with the α1 and α2 helixes of the MHC. Previously we have shown that the A6 TCR in complex with the HLA-A2/Tax peptide has 15 contact sites on HLA-A2. Single amino acid mutagenesis of these contact sites demonstrated that mutation of only three amino acids clustered on the α1 helix (R65, K66, A69) disrupted recognition by the A6 TCR. In the present study we have asked whether TCRs that recognize four other peptides presented by HLA-A2 interact with the MHC in identical, similar, or different patterns as the A6 TCR. Mutants K66A and Q155A had the highest frequency of negative effects on lysis. A subset of peptide-specific CTL also selectively recognized mutants K66A or Q155A in the absence of exogenous cognate peptides, indicating that these mutations affected the presentation of endogenous peptide/HLA-A2 complexes. These findings suggest that most HLA-A2-restricted TCRs recognize surfaces on the HLA-A2/peptide complex that are dependent upon the side chains of K66 and Q155 in the central portion of the peptide binding groove. Crystallographic structures of several peptide/HLA-A2 structures have shown that the side chains of these critical amino acids that make contact with the A6 TCR also contact the bound peptide. Collectively, our results indicate that the generalized effects of changes at these critical amino acids are probably due to the fact that they can be directly contacted by TCRs as well as influence the binding and presentation of the bound peptides.
γ/δ T Cell-Deficient Mice Exhibit Reduced Disease Severity and Decreased Inflammatory Response in the Brain in Murine Neurocysticercosis
Authors: Cardona Astrid E. , Teale Judy M. 
Published: April 2014

AbstractIn a recently developed mouse model for neurocysticercosis, the immune response was characterized by a massive influx of γδ T cells and a type 1 pathway of cytokine expression. To understand the role of γδ T cells during this infection, the cellular and cytokine response was analyzed in mice that lack γδ T cells (TCRδ−/−). In TCRδ−/− mice, Mesocestoides corti metacestodes preferentially invaded the extraparenchymal areas of the brain. Furthermore, parasites were able to escape from the brain and establish a systemic infection with liver and peritoneal involvement. Immunopathological studies indicated that TCRδ−/− mice develop little inflammatory response and less neurological symptomatology. Significantly reduced numbers of T cells, macrophages, dendritic cells, and mast cells were present in the brain. The cytokine response in the brain of TCRδ−/− mice appears to be a mixed type1/type 2 response with low levels of IL-2, IL-4, IL-10, IL-12, IL-13, IL-15, and IFN-γ. To further investigate the immunological significance of this cell population, γδ T cells were adoptively transferred into intracranially infected TCRδ−/− mice. γδ T cells were specifically recruited into the CNS in response to this parasitic infection, and they were able to target the infected brain within 12 h after transfer. These results suggest that γδ T cells are key players in the immune response elicited during this CNS infection and direct a type 1 response in wild-type mice upon infection.
A C-Reactive Protein Mutant That Does Not Bind to Phosphocholine and Pneumococcal C-Polysaccharide
Authors: Agrawal Alok , Simpson Melanie J. , Black Steven , Carey Marianne P. , Samols David 
Published: April 2014

AbstractC-reactive protein (CRP), the major human acute-phase plasma protein, binds to phosphocholine (PCh) residues present in pneumococcal C-polysaccharide (PnC) of Streptococcus pneumoniae and to PCh exposed on damaged and apoptotic cells. CRP also binds, in a PCh-inhibitable manner, to ligands that do not contain PCh, such as fibronectin (Fn). Crystallographic data on CRP-PCh complexes indicate that Phe66 and Glu81 contribute to the formation of the PCh binding site of CRP. We used site-directed mutagenesis to analyze the contribution of Phe66 and Glu81 to the binding of CRP to PCh, and to generate a CRP mutant that does not bind to PCh-containing ligands. Five CRP mutants, F66A, F66Y, E81A, E81K, and F66A/E81A, were constructed, expressed in COS cells, purified, and characterized for their binding to PnC, PCh-BSA, and Fn. Wild-type and F66Y CRP bound to PnC with similar avidities, while binding of E81A and E81K mutants to PnC was substantially reduced. The F66A and F66A/E81A mutants did not bind to PnC. Identical results were obtained with PCh-BSA. In contrast, all five CRP mutants bound to Fn as well as did wild-type CRP. We conclude that Phe66 is the major determinant of CRP-PCh interaction and is critical for binding of CRP to PnC. The data also suggest that the binding sites for PCh and Fn on CRP are distinct. A CRP mutant incapable of binding to PCh provides a tool to assess PCh-inhibitable interactions of CRP with its other biologically significant ligands, and to further investigate the functions of CRP in host defense and inflammation.
Human Papillomavirus Virus-Like Particles Do Not Activate Langerhans Cells: A Possible Immune Escape Mechanism Used by Human Papillomaviruses
Authors: Fausch Steven C. , Da Silva Diane M. , Rudolf Michael P. , Kast W. Martin 
Published: April 2014

Abstract
High-risk human papillomaviruses are linked to several malignancies including cervical cancer. Because human papillomavirus-infected women do not always mount protective antiviral immunity, we explored the interaction of human papillomavirus with Langerhans cells, which would be the first APCs the virus comes into contact with during infection. We determined that dendritic cells, normally targeted by vaccination procedures and Langerhans cells, normally targeted by the natural virus equally internalize human papillomavirus virus-like particles. However, in contrast to dendritic cells, Langerhans cells are not activated by human papillomavirus virus-like particles, illustrated by the lack of: up-regulating activation markers, secreting IL-12, stimulating T cells in an MLR, inducing human papillomavirus-specific immunity, and migrating from epidermal tissue. Langerhans cells, like dendritic cells, can display all of these characteristics when stimulated by proinflammatory agents. These data may define an intriguing immune escape mechanism used by human papillomavirus and form the basis for designing optimal vaccination strategies.
Opsonization of HIV-1 by Semen Complement Enhances Infection of Human Epithelial Cells
Authors: Bouhlal Hicham , Chomont Nicolas , Haeffner-Cavaillon Nicole , Kazatchkine Michel D. , Belec Laurent , Hocini Hakim 
Published: April 2014

AbstractIn the present study we demonstrate that both X4- and R5-tropic HIV-1 strains are able to infect the human epithelial cell line HT-29. Infection was enhanced 2-fold when HIV was added to semen before contact with the cell cultures. The enhancing effect of semen was complement dependent, as evidenced by blockage of generation of C3a/C3adesArg in semen by heat or EDTA treatment of semen and suppression of semen-dependent enhancement with mAbs directed to complement receptor type 3 (CD11b/CD18) and soluble CD16. Infection of HT-29 cells was assessed by the release of p24 Ag in cultures and semiquantitative PCR of the HIV-1 pol gene. Inhibition of infection of HT-29 by stromal cell-derived factor 1 was decreased in the case of semen-opsonized X4- and R5-tropic virus compared with unopsonized virus. In contrast, inhibition of infection by RANTES was increased for opsonized X4-tropic HIV-1 compared with unopsonized virus. Taken together these observations indicate that activation of complement in semen may play an enhancing role in mucosal transmission of HIV-1 by facilitating infection of epithelial cells and/or enhancing infection of complement receptor-expressing target cells in the mucosa.
HIV-1 Transcription and Virus Production Are Both Accentuated by the Proinflammatory Myeloid-Related Proteins in Human CD4+ T Lymphocytes
Authors: Ryckman Carle , Robichaud Gilles A. , Roy Jocelyn , Cantin Réjean , Tremblay Michel J. , Tessier Philippe A. 
Published: April 2014

Abstract
S100A8, S100A9, and S100A12, collectively known as myeloid-related proteins (MRPs), are highly expressed by the myeloid cell lineage and are found in the extracellular milieu during infections and inflammatory conditions. Recent data showed high levels of MRPs in the serum of HIV type 1 (HIV-1)-infected patients which correlated with disease progression and low CD4+ counts. Therefore, we set out to investigate the effect of MRPs on HIV-1 replication. We observed a 4- to 5-fold induction of virus production in J1.1, a human T lymphoid cell line latently infected with HIV-1, following treatment with MRPs. Using luciferase-based reporter gene assays, we demonstrated that MRPs induce a dose- and time-dependent activation of the HIV-1 long terminal repeat promoter region that could be blocked by specific anti-MRP polyclonal Abs and by physical denaturation of these proteins. The MRP-mediated induction was acting through the HIV-1 enhancer sequence and was dependent upon NF-κB activity. These latter results were also confirmed by EMSA experiments conducted in Jurkat cells and freshly isolated PBMCs. In conclusion, we demonstrate that MRPs induce HIV-1 transcriptional activity and viral replication in infected CD4+ T-lymphocytes at concentrations similar to those found in the serum of HIV-1-infected patients.
Adoptive Immunotherapy of Advanced Tumors with CD62 L-Selectinlow Tumor-Sensitized T Lymphocytes Following Ex Vivo Hyperexpansion
Authors: Wang Li-Xin , Chen Bing-Guan , Plautz Gregory E. 
Published: April 2014

AbstractTumor-draining lymph nodes (TDLN) contain sensitized T cells with the phenotype CD62 L-selectinlow (CD62Llow) that can be activated ex vivo with anti-CD3 mAb and IL-2 to acquire potent dose-dependent effector function manifested upon adoptive transfer to secondary tumor-bearing hosts. In this study advanced tumor models were used as a stringent comparison of efficacy for the CD62Llow subset, comprising 5–7% of the TDLN cells, vs the total population of TDLN cells following culture in high dose IL-2 (100 U/ml). During the 9-day activation period the total number of CD8+ T cells increased 1500-fold, with equivalent proliferation in the CD62Llow vs the total TDLN cell cultures. Adoptive transfer of activated CD62Llow cells eliminated 14-day pulmonary metastases and cured 10-day s.c. tumors, whereas transfer of maximally tolerated numbers of total TDLN cells was not therapeutic. Despite their propagation in a high concentration of IL-2, the hyperexpanded CD62Llow subset of TDLN cells functioned in vivo without exogenous IL-2, and CD8+ T cells demonstrated relative helper independence. Moreover, the anti-tumor response was specific for the sensitizing tumor, and long term memory was established. The facile enrichment of tumor-reactive TDLN T cells, based on the CD62Llow phenotype, circumvents the need for prior knowledge of the relevant tumor Ags. Coupling the isolation of pre-effector T cells with rapid ex vivo expansion to &gt;3 logs could overcome some of the shortcomings of active immunotherapy or in vivo cytokine treatment, where selective robust expansion of effector cells has been difficult to achieve.