bluuebunny/crossref_metadata_2025_split huggingface dataset:
Heavy chain variable region, light chain variable region, and heavy chain CDR3 influences on the mono- and polyreactivity and on the affinity of human monoclonal rheumatoid factors.
Authors: Crouzier R , Martin T , Pasquali J L 
Published: December 2022

Abstract
Monoreactive high affinity pathologic autoantibodies were supposed previously to derive through somatic mutation from polyreactive low affinity autoantibodies that are encoded by a small set of unmutated V region genes in fetal and neonatal B cells. However, recent data exploring the physiologically expressed Ab repertoire and the importance of the stochastically generated heavy chain CDR3 (H-CDR3) in autoreactivity suggest that this scheme is incomplete. Here we analyzed via gene-swapping experiments and site-directed mutagenesis the relative contributions of the mutations in the light chain variable region (VL) and the heavy chain variable region (VH) domains and of the H-CDR3 in the autoreactivity of two IgM rheumatoid factors (RF), one a polyreactive low affinity Ab, the other a monoreactive high affinity Ab. These two RFs derived from the same V kappa III (humkv325) and VH1 (51p1) genes, but differed from each other by a few mutations and by the structure of the H-CDR3. The analysis of the reactivity patterns of different combinations of wild-type and in vitro engineered hybrid gene products clearly demonstrates the main influence of the H-CDR3 in the autoAb activity profiles. The results directly demonstrate the previously proposed hypothesis, namely, that the H-CDR3 plays a critical role in distinguishing poly- from monospecific RF. However, the data also indicate that self polyreactivity is a very fragile property and is dependent upon the primary structure of the VH segment.
Peptides control the gain and loss of allele specificity by mutated MHC class I molecules.
Authors: Hengel H , Burke K , Kyburz D , Zinkernagel R M , Koszinowski U H 
Published: December 2022

Abstract
To analyze the molecular basis of MHC class I allele-restricted peptide recognition, a set of eight Ld/Lq mutants was constructed and tested for peptide recognition by allele-restricted and peptide-specific CTL. The MHC molecules H-2Ld and H-2Lq differ at six amino acid positions (95, 97, 107, 116, 155, 157) located within the alpha 2 domain of the molecule. Both molecules present the lymphocytic choriomeningitis virus (LCMV) nucleoprotein-derived peptide RPQASGVYM and the murine cytomegalovirus (MCMV) pp89-derived peptide YPHFMPTNL to the respective virus-specific CD8+ CTL is a strictly allele-restricted fashion. All mutated MHC class I molecules did still bind the LCMV peptide and seven of eight mutants retained MCMV peptide binding. The exchange Arg--&gt;Trp at position 97 of Lq in pocket C of the peptide binding groove prevented binding of the MCMV ligand and this loss was compensated by the additional exchange of Ile--&gt;Leu in position 95 (pocket F). Within the Lq molecule, single mutations at either position 97 on the floor of the groove or position 155 of the wall sufficed for a gain of LCMV peptide recognition by Ld-restricted CTL. Altogether, six of eight mutants resulted in a gain of recognition by CTL specific for the other allele. Thus, six of the eight mutants lost MHC-restricted recognition and were accepted by both Ld- as well as Lq-restricted CTL when presenting the LCMV peptide. Only one case of simultaneous recognition of the MCMV peptide by both Ld- as well as Lq-restricted CTL was noted. In other mutations, a gain of recognition by Ld-restricted CTL was associated with a loss of recognition of Lq-restricted CTL. Analysis of extracted MCMV peptide from mutant molecules excluded quantitative differences in presented MCMV peptide as a reason for the lack of CTL recognition. Altogether, the results show that, rather than aminoacids at certain residue positions, individual peptides govern MHC allele specificity of CTL recognition.
Rearrangement and expression of the human psi C lambda 6 gene segment results in a surface Ig receptor with a truncated light chain constant region.
Authors: Stiernholm N B , Verkoczy L K , Berinstein N L 
Published: December 2022

Abstract
The constant region of the human Ig lambda locus consists of seven tandemly organized J-C gene segments. Although it has been established that the J-C lambda 1, J-C lambda 2, J-C lambda 3, and J-C lambda 7 gene segments are functional, and code for the four distinct Ig lambda isotypes found in human serum, the J-C lambda 4, J-C lambda 5, and J-C lambda 6 gene segments are generally considered to be pseudogenes. Although one example of a functional J-C lambda 6 gene segment has been documented, in the majority of cases, J-C lambda 6 is rendered nonfunctional by virtue of a single duplication of four nucleotides, creating a premature translational arrest. We show here that rearrangements to the J-C lambda 6 gene segment do occur, and that such a rearrangement encodes an Ig lambda protein that lacks the terminal end of the constant region. We also show that this truncated protein is expressed on the surface with the IgH chain, creating an unusual surface Ig (sIg) receptor (sIg delta CL). Cells that express this receptor on the surface do so at significantly reduced levels compared with clonally related variants, which express sIg receptors with conventional Ig lambda L chains. However, the effects of sIg cross-linking on tyrosine phosphorylation and surface expression of the CD25 and CD71 Ags are similar in cells that express conventional sIg receptors and in those that express sIg delta CL receptors, suggesting that the latter could possibly function as an Ag receptor.
Active immunotherapy of cancer with a nonreplicating recombinant fowlpox virus encoding a model tumor-associated antigen.
Authors: Wang M , Bronte V , Chen P W , Gritz L , Panicali D , Rosenberg S A , Restifo N P 
Published: December 2022

Abstract
Some tumor cells express Ags that are potentially recognizable by T lymphocytes and yet do not elicit significant immune responses. To explore new immunotherapeutic strategies aimed at enhancing the recognition of these tumor-associated Ags (TAA), we developed an experimental mouse model consisting of a lethal clone of the BALB/c tumor line CT26 designated CT26.WT, which was transduced with the lacZ gene encoding beta-galactosidase, to create CT26.CL25. The growth rate and lethality of CT26.CL25 and CT26.WT were virtually identical despite the expression by CT26.CL25 of the model tumor Ag in vivo. A recombinant fowlpox virus (rFPV), which is replication incompetent in mammalian cells, was constructed that expressed the model TAA, beta-galactosidase, under the influence of the 40-kDa vaccinia virus early/late promoter. This recombinant, FPV.bg40k, functioned effectively in vivo as an immunogen, eliciting CD8+ T cells that could effectively lyse CT26.CL25 in vitro. FPV.bg40k protected mice from both subcutaneous and intravenous tumor challenge by CT26.CL25, and most surprisingly, mice bearing established 3-day pulmonary metastasis were found to have significant, Ag-specific decreases in tumor burden and prolonged survival after treatment with the rFPV. These observations constitute the first reported use of rFPV in the prevention and treatment of an experimental cancer and suggest that changing the context in which the immune system encounters a TAA can significantly and therapeutically alter the host immune response against cancer.
Chloromethyl ketones block induction of nitric oxide synthase in murine macrophages by preventing activation of nuclear factor-kappa B.
Authors: Kim H , Lee H S , Chang K T , Ko T H , Baek K J , Kwon N S 
Published: December 2022

Abstract
N-alpha-tosyl-L-phenylalanine chloromethyl ketone (TPCK) and N-alpha-tosyl-L-lysine chloromethyl ketone (TLCK), serine protease inhibitors, block many cytotoxic functions of immune cells including superoxide anion production, cytokine release, cell-mediated cytolysis, and nitric oxide (NO)-related macrophage functions. IFN-gamma/LPS-induced NO production from murine peritoneal macrophages was inhibited by TPCK and TLCK in a dose-dependent manner (EC50s: approximately 20 microM for TPCK and approximately 30 microM for TLCK). Viability exceeded 91% with 25 microM TPCK and with 80 microM TLCK. When TPCK treatment was delayed until 1 h of activation, the inhibitory effect was markedly reduced. After 2 h of the activation, TPCK was not effective anymore. Addition of either TNF-alpha or conditioned media from IFN-gamma/LPS-activated macrophage culture did not prevent the inhibitory effect of TPCK. Neither TPCK nor TLCK reduced enzymatic NO production from macrophage lysates. Lysates from TPCK-treated cells did not generate NO even after supplementing necessary cofactors for NO synthase. Immunoblotting analysis showed that simultaneous treatment of TPCK with IFN-gamma/LPS abolished the NO synthase expression, whereas delayed addition of TPCK was either partially effective or not effective at all. Furthermore, TPCK treatment reduced the concentration of mRNA for NO synthase without decreasing mRNA stability. Thus, the serine protease inhibitors directly blocked an early event in expression of NO synthase. Electrophoretic mobility shift assay indicated that TPCK blocked the activation of nuclear factor-kappa B, a transcription factor necessary for NO synthase induction. TPCK also blocked disappearance of I kappa B from cytosolic fraction, and nuclear translocation of NF-kappa B subunits p50 and p65. Delaying the addition of TPCK by 10 min partially prevented the inhibition of the NF-kappa B activation process and allowed partial resuming of NO production. Thus, TPCK inhibited NO synthase induction by blocking NF-kappa B activation.
Eosinophil major basic protein induces degranulation and IL-8 production by human eosinophils.
Authors: Kita H , Abu-Ghazaleh R I , Sur S , Gleich G J 
Published: December 2022

Abstract
Eosinophil granule proteins, such as major basic protein (MBP), eosinophil peroxidase (EPO), and eosinophil cationic protein (ECP), possess a wide range of biologic activities including the ability to activate other cells, such as basophils, neutrophils, and platelets. Here we have analyzed the effects of these proteins on eosinophils themselves. MBP and EPO, at concentrations as low as 0.1 micrograms/ml, induced eosinophil degranulation as measured by release of eosinophil-derived neurotoxin (EDN); in contrast, ECP, at 1 micrograms/ml, was inactive. MBP (10 micrograms/ml) and EPO (0.1 micrograms/ml) induced EDN release comparable with one of the strongest agonists for eosinophils, secretory IgA. Pretreatment of cells with dibutyryl cAMP or cytochalasin B completely abolished the EDN release induced by MBP and EPO, suggesting that the effects of MBP and EPO are not due to cytotoxic lysis of the cells. Degranulation induced by MBP was only partially dependent on calcium, and no elevation of intracellular Ca2+ concentration ([Ca2+]i) was observed in eosinophils stimulated with MBP. MBP stimulated the production, up to eightfold, of IL-8 by eosinophils in a dose-dependent manner. The MBP-stimulated expression of IL-8 mRNA by eosinophils was confirmed by reverse transcription-PCR. The MBP-stimulated production of IL-8 was inhibited by actinomycin D, but not by cyclosporin A. Furthermore, MBP and calcium ionophore ionomycin synergistically induced production of leukotriene C4 from eosinophils. Thus, MBP and EPO may act as autocrine mediators in the pathogenesis of eosinophil-associated diseases, such as bronchial asthma.
TNF-alpha induces tyrosine phosphorylation of mitogen-activated protein kinase in adherent human neutrophils.
Authors: Rafiee P , Lee J K , Leung C C , Raffin T A 
Published: December 2022

Abstract
Recombinant human TNF-alpha induces increased tyrosine phosphorylation of several proteins in human neutrophils (PMN) adhered to serum-coated plastic. When PMN are kept in suspension, TNF does not induce significant tyrosine phosphorylation. In adherent PMN, a 42-kDa protein (p42) displayed the most striking increase in tyrosine phosphorylation after TNF stimulation. Cell lysates of TNF-stimulated PMN were separated by two-dimensional gel electrophoresis and were immunoblotted with either anti-phosphotyrosine (alpha-PY) mAb or anti-mitogen-activated protein kinase (alpha-MAPK) mAb. Both Abs detected p42, and the spots were superimposable. Cell lysates were immunoprecipitated with agarose-conjugated alpha-PY mAb, electrophoresed, and then immunoblotted with alpha-MAPK Ab; alternatively, cell lysates were immunoprecipitated with agarose-conjugated alpha-MAPK Ab, electrophoresed, and then immunoblotted with alpha-PY mAb. In both cases, p42 was detected. These results demonstrate that p42 is a member of the MAPK family. TNF induces a time-and dose-dependent increase in tyrosine phosphorylation of p42 and MAPK activity. The degree of p42 tyrosine phosphorylation parallels the level of MAPK activity. MAPK activity was determined by measuring 32P phosphorylation of a synthetic peptide containing the recognition site on myelin basic protein for MAPK. PMN pretreatment with genistein, a tyrosine kinase inhibitor, inhibited the TNF-induced increase in tyrosine phosphorylation and MAPK activity. These results indicate that TNF signaling involves activation of MAPK.
Molecular basis of the complement C7 M/N polymorphism. A neutral amino acid substitution outside the epitope of the allospecific monoclonal antibody WU 4-15.
Authors: Würzner R , Fernie B A , Jones A M , Lachmann P J , Hobart M J 
Published: December 2022

Abstract
The allotypes of the C7 M/N polymorphism are determined by comparing the ELISA reaction pattern of the allospecific mAb WU 4-15 with polyclonal anti-C7 IgG. To characterize the molecular basis of this polymorphism, the WU 4-15 epitope was mapped by expression of cDNA fragments. It was found to be located within the boundary region of the two short consensus repeats of C7 (amino acid residues 595-612). Coincidentally an A or C substitution was found in the course of investigation of the gene structure at nucleotide 1759. This leads to an electrically neutral Pro/Thr substitution at residue 565 and alternative Sau961 or RsaI restriction sites, respectively. Enzymatic digestion or sequencing of PCR-amplified genomic DNA from C7 M/N-typed individuals showed a perfect correspondence of the genotype with the phenotype of the C7 M/N polymorphism. In contrast, no association was found with the genotypes of a second polymorphic site at residue 367. Thus, a threonine at amino acid residue 565 allows access of WU 4-15 to its epitope but its substitution with proline almost completely inhibits access in the native molecule, although the epitope is about 40 amino acid residues distant from the polymorphic site in the primary structure. Furthermore, the different conformational structure is probably responsible for the hypomorphic appearance of C7*N.
Cleavage of lymphocyte surface antigens CD2, CD4, and CD8 by polymorphonuclear leukocyte elastase and cathepsin G in patients with cystic fibrosis.
Authors: Döring G , Frank F , Boudier C , Herbert S , Fleischer B , Bellon G 
Published: December 2022

Abstract
Polymorphonuclear leukocytes (PMN) accumulating in airways of patients with cystic fibrosis (CF) as a response to chronic endobronchial bacterial lung infection, release lysosomal serine proteinases such as PMN-elastase at concentrations of approximately 0.5 microM to 5 microM into the airway lumen. Immunohistology of CF lung material and fluorescence activated cell sorter analysis of sequential CF bronchoalveolar lavages demonstrated loss of the CD4 and CD8 Ag on CD3+ T lymphocytes in sputum-filled airways. In 10 CF sputum samples 1.0%, 19.1%, and 15.7% of all CD3+ T lymphocytes expressed CD4, CD8, and CD2, respectively. Incubation of CF sputum supernatant fluids with peripheral blood T lymphocytes resulted in total reduction of CD4 and CD8 but not CD2. Addition of alpha 1-proteinase inhibitor abolished surface Ag cleavage completely. Purified PMN-elastase and cathepsin G cleaved CD2, CD4, and CD8 on peripheral blood T lymphocytes at proteinase concentrations of 0.83 to 8.3 microM in a dose-dependent manner. Cleaved CD4 and CD8 were reexpressed on the surface of T lymphocytes after 24 h in the absence of PMN-elastase. Incubation of a CD4+ T cell clone with PMN-elastase lead to a significant reduction of cytotoxicity toward target cells and significantly reduced IL-2 and IL-4 production. The results suggest a temporary functional impairment of T lymphocytes in foci of high inflammation characterized by stimulated PMN, which may lower tissue destruction.
Analysis of the heavy chain repertoire of human peripheral B cells using single-cell polymerase chain reaction.
Authors: Brezinschek H P , Brezinschek R I , Lipsky P E 
Published: December 2022

Abstract
The VH gene repertoire of human peripheral B cells was analyzed using PCR analysis of individual blood B cells. Because genomic DNA of single B cells was analyzed, data from both productive and nonproductive VDJ rearrangements were obtained. Nine out of 75 B cells contained both functional and nonfunctional rearrangement products, whereas 62/75 had a single productive VDJ rearrangement. The distribution of VH families was ordered in accordance with the germline complexity, although a bias toward VH3 and some of its members was found. This bias was noted in both the productively and nonproductively rearranged repertoires, indicating that it resulted from molecular and not selective processes. Evidence for negative selection of certain VH3 and VH4 family members was noted in that they were found less often as productive than nonproductive VDJ rearrangements. In addition, evidence for positive selection based on CDR3 was obtained, in that JH6 and DXP'1 were found at a higher frequency in the productively compared with the nonproductively rearranged repertoire. The CDR3 segments were markedly heterogenous, with a much greater degree of variability noted in the nonproductive VDJ rearrangements. Finally, a much greater frequency of mutations was noted in the nonproductively rearranged VH genes within individual B cells. These results have begun to delineate the human peripheral B cell repertoire and identify processes that shape it both positively and negatively.
Subverting lymph node trafficking by treatment with the Mel-14 monoclonal antibody to L-selectin does not prevent an effective host response to Sendai virus.
Authors: Hou S , Hyland L , Bradley L M , Watson S R , Doherty P C 
Published: December 2022

Abstract
A single 250-micrograms dose of the Mel-14 mAb to L-selectin greatly diminished the extent of L-selectin expression on lymphocytes and decreased (60 to 90%) the massive cellular recruitment to the cervical and mediastinal lymph nodes that follows intranasal infection of naive C57BL/6 mice with Sendai virus. The numbers of CD8+ CTL precursors in the mediastinal lymph nodes were considerably reduced on day 7, when compared with virus-infected mice given a control rat IgG2a, but potent CTL effectors were present in the lungs of both groups by day 10 after infection, and the overall magnitude of CTL precursor generation was not obviously compromised. The early dominance of Sendai virus-specific IgM Ab-forming cells was prolonged in the Mel-14-treated mice, whereas plasma cells producing virus-specific IgA were abnormally prominent in the lymph nodes but not in the spleen. The kinetics of virus-specific Ab-forming cells generation and the serum Ab response for the various IgG isotypes were also delayed. Thus, though L-selectin is clearly important for the localization of naive lymphocytes to regional lymph nodes, the Mel-14-treated mouse can still deal effectively with a virus that causes productive infection only in the respiratory tract. The spleen, where L-selectin does not determine lymphocyte trafficking, is a major site for the compensatory T cell and B cell responses.
Two waves of gamma delta T cells expressing different V delta genes are recruited into schistosome-induced liver granulomas.
Authors: Sandor M , Sperling A I , Cook G A , Weinstock J V , Lynch R G , Bluestone J A 
Published: December 2022

Abstract
Isolated granulomas provide a unique model to study T cells in the site of inflammation. Schistosoma mansoni-infected mice develop liver granulomas in response to schistosome egg deposition and the granulomas contain gamma delta T cells that appear to be activated (Pgp-1high and L-selectin low). Analysis of kinetics and TCR gene usage of granuloma gamma delta T cells revealed a limited TCR repertoire restricted to V gamma 1.1, V delta 4, and V delta 6 genes, suggesting that the occurrence of gamma delta T cells in the granuloma is influenced by TCR V gene usage. The V delta 4+, but not the V delta 6+, gamma delta T cells expressed CD69, a marker of recent activation. To determine if there was a preferred order of accumulation of the gamma delta T cells in granulomas, s.c. sponge grafts were implanted into schistosome-infected mice, schistosome eggs were injected into the grafts, and accumulated T cells were sequentially analyzed. The earliest gamma delta T cell immigrants expressed V delta 6 and later immigrants expressed V delta 4 V genes. Additional evidence for a role of TCR specificity in the accumulation of gamma delta T cells in granulomas is their absence from schistosome granulomas in TCR transgenic mice that express only a single MHC-specific gamma delta TCR. Finally, gamma delta T cell recruitment into the granulomas did not require beta 2-microglobulin, since gamma delta T cells were present in liver granulomas of beta 2-microglobulin gene-disrupted mice. The analysis of the influx of gamma delta T cells into schistosome-induced, liver granulomas and schistosome egg-containing sponges provides a model system to investigate the role, if any, of gamma delta T cells in schistosome infections.
Antigen-based heteropolymers facilitate, via primate erythrocyte complement receptor type 1, rapid erythrocyte binding of an autoantibody and its clearance from the circulation in rhesus monkeys.
Authors: Ferguson P J , Martin E N , Greene K L , Kuhn S , Cafiso D S , Addona G , Taylor R P 
Published: December 2022

Abstract
We investigated the feasibility of using the primate E complement receptor (CR1), in concert with Ag-based heteropolymers (AHP), as a potential therapy to remove autoantibodies from the circulation. AHP are prepared by cross-linking an anti-CR1 mAb with the acetylcholine receptor (AChR), the principal target Ag in myasthenia gravis. In vitro studies demonstrate that this methodology facilitates specific, rapid, and quantitative binding of an anti-AChR mAb to primate Es. In vivo experiments in rhesus monkeys indicate that AHP-mediated binding of an anti-AChR mAb to Es leads to the clearance of the mAb from the circulation. Once bound to the E via the AHP, the autoantibody is transported to the liver and spleen, where it is degraded without destruction of the E. It is therefore likely that the complexes of AHP and target mAb, when bound to Es, are recognized in vivo and processed by a mechanism quite similar to that which occurs when complement-opsonized immune complexes, bound to primate Es, are cleared from the circulation. It may be possible to extend and generalize this work to allow for the development of a simple, noninvasive therapy that can be made specific for the treatment of several different autoimmune diseases.