bluuebunny/crossref_metadata_2025_split huggingface dataset:
Heavy chain joining region segments of the channel catfish. Genomic organization and phylogenetic implications.
Authors: Hayman J R , Ghaffari S H , Lobb C J 
Published: December 2022

Abstract
The JH locus of the channel catfish has been characterized to determine the organization and structural diversity of JH segments. These analyses indicate that there are a total of nine JH segments tightly clustered within a region spanning about 2.2 kb. The JH locus is closely linked to the CH 1 domain of the expressed catfish H chain; the distance between the CH proximal JH segment (JH9) and the CH 1 domain is about 1.8 kb. Each JH segment has an upstream recombination sequence, which includes a T-rich nonamer, a 22- to 24-bp spacer, and a phylogenetically conserved heptamer. Each JH segment also has an open reading frame that encodes the conserved framework region 4 tryptophan (Trp-103) and terminates with a RNA donor splice site. The catfish JH locus contains an internal repetitive sequence region characterized by a short (183-188 bp) repeat that occurs sequentially five times. Strong sequence homology as well as the unified length of the repeated sequences indicate that JH segments JH3-JH7 probably arose as the result of a series of homologous but unequal crossover events. Sequence alignments of the duplicated JH segments indicates that there is diversity within the 5-11 nucleotides located immediately downstream from the heptamer, an observation which indicates that closely related JH segments can serve to enhance CDR3 diversity in the expressed H chain. Comparisons of the genomic JH sequences with different cDNA clones indicate that each JH segment is probably functional and that junctional diversity serves an important role in the generation of CDR3 diversity. In addition, single base differences observed in comparisons of JH-encoded regions indicate that there is probably somatic mutation or allelic variation of genomic JH segments. These studies suggest that the characteristic structure and organizational pattern of JH segments in higher vertebrates may have evolved early in vertebrate phylogeny at the level of the bony fish.
Structural analysis of the VH-D-JH segments of human polyreactive IgG mAb. Evidence for somatic selection.
Authors: Ikematsu H , Kasaian M T , Schettino E W , Casali P 
Published: December 2022

Abstract
Polyreactive (natural) antibodies are primarily IgM and account for a major proportion of circulating Ig in humans. They use various V gene segments, in general, in germ line (unmutated) configuration. To analyze the VH regions of polyreactive antibodies, with particular attention at their somatically mutated status, we generated five IgG (three IgG1 and two IgG3) mAb (using B cells from a healthy subject, a patient with insulin-dependent diabetes mellitus and a patient with SLE), which bound with various efficiencies a number of different self and foreign Ag. Gene cloning experiments showed that the VH region sequences were unique to each IgG mAb. The H chain complementary determining region (CDR3) of two IgG (mAb10 and mAb426.4.2F20) displayed an identical stretch of five amino acids (RFLEW), but the other three IgG mAb CDR3 were divergent in both length and composition. The VH gene sequences of two IgG, mAb426.4.2F20 and mAb410.7.F91, were 99% identical to those of the germ line VH4.11 and VH4.21 genes, respectively. Those of the remaining three IgG mAb displayed a number of differences (93.6 to 95.9% identity) when compared with the germ line VH4.18, VH4.11, and hv1263 gene sequences. These and the VH4.21 gene have been found to encode polyreactive IgM and IgA and, in mutated configuration, monoreactive high affinity autoantibodies and antibodies induced by foreign Ag. When compared with the respective framework region, the CDR of three IgG mAb VH segment sequences displayed a significantly higher: 1) frequency of total nucleotide differences (6.1 x 10(-2) vs 4.5 x 10(-2) difference/base); 2) frequency of putative nucleotide changes yielding amino acid replacements (5.6 x 10(-2) vs 1.4 x 10(-2) replacement change/base); and 3) ratio of overall putative replacement to silent (R:S) mutations (11.0 vs 0.4). Thus, the distribution and nature of the nucleotide differences were consistent with a process of somatic mutation and Ag-dependent clonal selection. This was formally proved in IgG mAb426.12.3F1.4 and IgG mAb10 by differentially targeted polymerase chain reaction amplification and cloning and sequencing of the germ line genes that gave rise to the expressed VH segments, using DNA from polymorphonuclear cells of the same subjects whose B cells were used for the generation of these IgG mAb. Somatic mutations might have been responsible for bringing about polyreactivity in originally monoreactive antibodies or, more likely, they accumulated in originally polyreactive antibodies, which after undergoing a process of Ag selection, retained polyreactivity and may have or may have not acquired a higher affinity for the selecting Ag.
Cellular distribution of HLA-G mRNA in transgenic mouse placentas.
Authors: Yelavarthi K K , Schmidt C M , Ehlenfeldt R G , Orr H T , Hunt J S 
Published: December 2022

Abstract
The human MHC class I gene, HLA-G, is unique among members of the class I gene family in that it is nonpolymorphic, and expression is primarily restricted to extraembryonic tissues. To examine regulatory elements that direct tissue- and cell lineage-specific expression of this gene, transgenic mice expressing HLA-G have been established. In this study, in situ hybridization was used to evaluate the cellular distribution of HLA-G mRNA in transgenic placentas. Extraembryonic tissues were obtained at gestation day 12.5 from embryos that had been microinjected with either 6.0 or 5.7 kb of HLA-G genomic DNA and had been transferred into pseudopregnant HLA-G transgenic mice or Swiss mice. The 6.0 kb transgene contained an additional 250 bp at the extreme 5'-end of the upstream region. Genotype of the recipient had no discernable effect on the cellular distribution of HLA-G mRNA. HLA-G mRNA was present in both trophoblast and mesenchymal cells in transgenic placentas carrying 6.0 kb of genomic HLA-G, a pattern strikingly similar to that of HLA-G message distribution in early gestation human placentas. By contrast, in placentas from embryos carrying 5.7 kb HLA-G DNA, specific mRNA was found primarily in mesenchymal cells at the base of the placenta. Thus, the 6.0 genomic fragment contains elements capable of directing HLA-G expression in placentas, and is particularly influential in the trophoblastic cell lineage.
The hemopoietic progenitor 32DCl3(G) cells synthesize C3 molecules and acquire C3b acceptor sites upon differentiation or neoplastic transformation.
Authors: Di Renzo L , Zicari A , Longo A , Realacci M , Naso G , Pontieri G , Lipari M 
Published: December 2022

Abstract
32DCl3(G) cells are a diploid, nontumorigenic, IL-3-dependent hemopoietic progenitor cell line, which undergoes terminal differentiation into neutrophilic granulocytes when cultured in presence of G-CSF. The infection with BALB-Moloney murine sarcoma virus, containing a v-HA-ras oncogene, renders this cell line IL-3 independent and continuously growing in the presence of G-CSF, nontumorigenic and with an apparent block at the level of promyelocyte/myelocyte (32D-Ha-ras). After infection with Abelson murine leukemia virus containing a v-abl oncogene, the cell line originates (32D-abl) that is also IL-3 independent but is tumorigenic and unable to differentiate in the presence of G-CSF. This cellular model allowed us to study the relationship between distinct steps of cell differentiation, neoplastic transformation, and C3 synthesis, activation, and characteristics of binding. We demonstrated that C3 synthesis, release, and cleavage are properties already present in the progenitor 32DCl3(G) cells. The more differentiated 32D-Ha-ras cells acquired C3 acceptor sites, apparently completely saturated by the constitutively released molecules. The transformation with Abelson murine leukemia virus, although it did not affect all these properties, let the cells bind considerable amounts of C3-related fragments activated in normal murine serum through the alternative pathway. This last event was a result of the acquisition of PMSF-sensitive serine proteases associated with the plasma membrane.
Enhanced production of LPS-induced cytokines during differentiation of human monocytes to macrophages. Role of LPS receptors.
Authors: Gessani S , Testa U , Varano B , Di Marzio P , Borghi P , Conti L , Barberi T , Tritarelli E , Martucci R , Seripa D 
Published: December 2022

Abstract
In vitro cultivated human monocytes isolated from normal peripheral blood show a time-dependent differentiation into macrophages characterized by an increased expression of transferrin receptors, CD11/CD18, and CD14 Ag. We measured the secretion of TNF-alpha and IL-6 in freshly isolated monocytes and in differentiated macrophages after LPS treatment. Differentiated macrophages produced significantly higher amounts of TNF-alpha and IL-6 than freshly isolated monocytes. This increased secretion was not a result of an enhanced accumulation of TNF-alpha and IL-6 mRNA, as comparative levels of these transcripts were found in both cell types after LPS treatment. Furthermore, LPS did not induce an antiviral state to VSV3 in monocytes, but it reduced by 3 to 5 log10 the virus yield in differentiated macrophages. The addition of antibodies to IFN-beta completely inhibited the LPS-induced antiviral state to VSV, but antibodies to IFN-alpha, TNF-alpha, or IL-6 were ineffective. A marked accumulation of IFN-beta mRNA was found in both cell types after LPS treatment. Binding experiments with FITC-LPS revealed a slightly higher overall binding affinity for LPS in freshly explanted monocytes as compared with differentiated macrophages, even though the maximal binding was higher in macrophages. In both cell types, the LPS binding was partially inhibited by antibodies to CD14. These results demonstrate that: 1) in vitro differentiation of human monocytes to macrophages leads to an enhanced LPS response in terms of (a) progressive increase of IL-6/TNF-alpha production and (b) acquisition of an IFN-beta mediated antiviral state; 2) this enhanced response to LPS, largely CD14-independent, is not linked to any increased accumulation of cytokine mRNA, but is probably a result of an increased synthesis and/or secretion of these cytokines.
IL-3-dependent murine mast cells undergo apoptosis on removal of IL-3. Prevention of apoptosis by c-kit ligand.
Authors: Mekori Y A , Oh C K , Metcalfe D D 
Published: December 2022

Abstract
It is well established that mast cell proliferation and maturation are regulated by two principle cytokines, IL-3 and the c-kit ligand stem cell factor (SCF). Little is known, however, how these two processes are negatively regulated and thus, how mast cell number is controlled in normal or pathologic processes. In this study we hypothesized that IL-3-dependent mast cells would undergo programmed cell death (apoptosis) on removal of IL-3 as was shown with other growth factor-dependent hemopoietic cells. Apoptotic changes were analyzed using light microscopy, fluorescent staining with acridine orange, flow cytometric analysis, and DNA electrophoresis. We could demonstrate that elimination of IL-3 from either primary bone marrow-derived cultured mast cell cultures (BMCMC) or from the growth factor-dependent mast cell line MCP5 resulted in the characteristic changes of apoptosis including condensed chromatin, fragmented nuclei, cellular vacuolization, typical pattern of propidium iodide or Hoechst 33342 uptake by flow cytometry, and the characteristic 200 bp "ladder" pattern of DNA cleavage. These events were prevented by SCF, an action that was in part mediated by tyrosine kinases, in that the tyrosine kinase inhibitor herbimycin inhibited the action of SCF in preventing apoptosis in IL-3-deprived cells. By using anti c-kit mAb and IL-3-dependent BMCMC obtained from W/Wv mice homozygous for mutation at the w locus that encodes the c-kit receptor, we could also show that SCF exerted its effect via c-kit. Neither dexamethasone nor cyclosporin A inhibited the "rescue" effect of SCF, suggesting that "rescue" was mediated by SCF and not through the induction of other cytokines. Thus, IL-3-dependent mast cells undergo apoptosis on removal of IL-3, an event that is prevented by the addition of SCF through its ligand c-kit, thus demonstrating how these principle mast cell growth factors may act in concert to regulate mast cell number under physiologic conditions.
Immunolocalization of cytokines in the nasal mucosa of normal and perennial rhinitic subjects. The mast cell as a source of IL-4, IL-5, and IL-6 in human allergic mucosal inflammation.
Authors: Bradding P , Feather I H , Wilson S , Bardin P G , Heusser C H , Holgate S T , Howarth P H 
Published: December 2022

Abstract
Allergic mucosal inflammation is characterized by the presence of cell infiltration, predominantly with IgE-sensitized mast cells and activated eosinophils, and appears to be regulated by the local production and release of several cytokines, particularly IL-4 and IL-5. Although attention has focused on the Th2 subpopulation of CD4+ T lymphocytes as an important source of these cytokines, human mast cells have been shown to both store and secrete IL-4 and TNF-alpha. To investigate the expression of cytokines relevant to allergic inflammation and to identify their cellular localization within the nasal mucosa, we have undertaken specific immunohistochemical staining of thin sections of inferior turbinate biopsies from patients with perennial allergic rhinitis and, for comparison, from nonatopic healthy volunteers. The cytokines investigated were IL-4, IL-5, IL-6, and IL-8. In both the normal and rhinitic biopsies numerous cells immunoreactive for IL-4, IL-5, and IL-6 were seen. Staining of adjacent 2-microns sections for CD3, mast cell tryptase, and eosinophil cationic protein revealed that 90% of the IL-4 immunoreactive cells were mast cells, with biopsies from rhinitic subjects containing significantly more IL-4+ cells than biopsies from normal controls (p = 0.02), especially when assessed with the anti-IL-4 mAb 3H4. Mast cells also accounted for &gt; 90% of IL-6 and &gt; 50% of IL-5 immunoreactive cells. IL-5 immunoreactivity was also localized to eosinophils, whereas IL-8 localized predominantly to the nasal epithelium in both groups. No cytokines were found in association with T lymphocytes. These findings indicate that the mast cell is an important source of preformed cytokines and as such may contribute to the chronicity of the mucosal inflammation that characterizes allergic rhinitis.
IL-2 receptor-positive intrathyroidal lymphocytes in Graves' disease. Analysis of V alpha transcript microheterogeneity.
Authors: McIntosh R S , Tandon N , Pickerill A P , Davies R , Barnett D , Weetman A P 
Published: December 2022

Abstract
There has been considerable interest recently in the possible restriction of the TCR repertoire in autoimmune disorders, because such restriction would have important therapeutic implications. Reports of restriction of the TCR V alpha but not V beta repertoire in the thyroid in Graves' disease could not be repeated in an earlier study. Using RNA derived from matched peripheral blood, thyroid tissue, intrathyroidal lymphocytes (ITL), and IL-2R+ and IL-2R- subpopulations of ITL from Graves' patients, we conducted reverse transcription polymerase chain reaction/Southern blot analysis of TCR V alpha family usage. No evidence was found for V alpha restriction in the IL-2R+ subpopulation of ITL from eight patients, one of whom was operated on within 1 mo of diagnosis. We have further analyzed samples from seven of these patients by resolution on denaturing polyacrylamide gels. Typically, a single dominant band was amplified with surrounding minor bands in a normal distribution. Dominant and minor species were both the result of amplification from in frame V alpha transcripts, and the minor bands were separated in size by increments of 3 bp. We found no evidence for reduced heterogeneity of the V alpha transcripts in ITL or IL-2R+ ITL populations relative to peripheral blood in the vast majority of samples. This therefore suggests that there is little difference between the blood lymphocyte population and the activated T cell population in the thyroid in patients with Graves' disease, and indicates that in autoimmune thyroid disease, this method of analysis is not sufficient to distinguish between autoreactive and bystander T cell populations.
T cell-antigen-presenting cell interactions in human systemic lupus erythematosus. Evidence for heterogeneous expression of multiple defects.
Authors: Via C S , Tsokos G C , Bermas B , Clerici M , Shearer G M 
Published: December 2022

Abstract
To assess interactions between T cells and APC in patients with SLE, PBL were stimulated in vitro and IL-2 production measured after stimulation with either a MHC-self-restricted Ag (influenza A virus) or an Ag which can use both MHC-self-restricted or unrestricted T cell activation pathways (HLA-alloantigen). SLE patients (n = 26) and controls (n = 8) were categorized as responder (+) or nonresponder (-) for each stimulus and grouped according to their paired response pattern. All controls responded to both influenza virus (Flu) or alloantigen (Allo) and were categorized as +/+. In contrast, SLE patient response patterns were heterogeneous with no evidence for a single SLE-associated defect. Instead, SLE patient responses fell into one of three different patterns: a) normal responses to both Flu and Allo (+/+), observed in nine (35%) patients; b) defective responses to Flu but intact Allo responses (-/+) observed in 12 (46%) SLE patients; and c) defective responses to both Flu and Allo (-/-), observed in five (19%) SLE patients. There was no statistically significant correlation between immune response pattern and the use of immunosuppressants. Further analysis of -/- SLE patients indicated defects in APC function and in both CD4+ and CD8+ T cell function. In contrast, cell depletion and add-back studies in -/+ SLE patients demonstrated defects in APC function only. Thus, similar to the well recognized clinical heterogeneity among SLE patients, our data support the concept that SLE patients are heterogeneous with respect to in vitro T cell-APC function, exhibiting responses ranging from normal function to defects in APC and in both T cell subsets. Prospective studies are in progress to determine the clinical relevance of these observations.
A conservative mutation in a class I MHC molecule outside the peptide binding groove stimulates responses to self peptides.
Authors: Grandea A G , Bevan M J 
Published: December 2022

Abstract
A transgenic mouse has been made that expresses a mutant MHC class I H-2Kb molecule with glutamic acid at position 65 (E65) in place of glutamine. The side chain at position 65, on the outward face of the alpha-helix of the alpha 1 domain of the class I molecule, interacts with the TCR, and not with the peptide binding groove. The transgenic mouse, on a DBA/2 background, mounts Kb,E65-restricted Ag-specific responses to conventional Kb-restricted Ag such as OVA and vesicular stomatitis virus, and shows strong alloreactivity to wild-type Kb. The transgenic mouse also mounts a primary in vitro alloreactive response directed to a mutant molecule with aspartic acid at position 65 (D65). This response is relatively weak, probably because of the structural similarities between aspartic and glutamic acid side chains; both have carboxylic termini, and the aspartic acid side chain is shorter by a single secondary carbon. The alloreactive CTL lines elicited by this conservative change are cross-reactive among several position-65 variants of H-2Kb. Individual CTL clones are specific for self peptides that can be extracted from cells expressing Kb,E65, and from purified wild-type Kb molecules, and that are recognized in the context of the D65 residue. Thus, the smallest variance from self in a class I molecule, even outside the peptide binding groove, can be antigenic.
Immunogenic peptides bind to class II MHC molecules in an early lysosomal compartment.
Authors: Harding C V , Geuze H J 
Published: December 2022

Abstract
Exogenous protein Ag are processed within endocytic compartments to produce peptides that bind to class II MHC (MHC-II) molecules for presentation to T cells. We have now identified a subcellular compartment in which immunogenic peptides bind to MHC-II as a subset of high density lysosomes. Immunoelectron microscopy of whole cells and dense Percoll gradient subcellular fractions showed early tubulovesicular lysosomes with high levels of MHC-II. Typical mature lysosomes contained less MHC-II. Pulse-chase biosynthetic labeling of macrophages followed by immunoprecipitation of MHC-II from dense lysosomal fractions showed that MHC-II molecules targeted efficiently to lysosomes after biosynthesis. Moreover, lysosomal MHC-II molecules were rapidly loaded with immunogenic peptide (as detected by T cells) soon after exposure of macrophages to Ag and before similar expression of peptide-MHC-II complexes on the plasma membrane; this loading was blocked at 18 degrees C. We propose that nascent MHC-II molecules target to early tubulovesicular lysosomes and bind immunogenic peptides therein; the resulting peptide-MHC-II complexes are then transported to the plasma membrane.
Novel mechanism for inhibition of human T cells by glucocorticoids. Glucocorticoids inhibit signal transduction through IL-2 receptor.
Authors: Paliogianni F , Ahuja S S , Balow J P , Balow J E , Boumpas D T 
Published: December 2022

Abstract
Interaction of IL-2 with its high affinity membrane receptor complex (IL-2R) present on activated T lymphocytes induces cell proliferation and mediates effector functions. Glucocorticoids inhibit IL-2 production by inhibiting TCR-mediated signal transduction. We asked whether they also inhibit the action of IL-2 by inhibiting signal transduction through IL-2R. Human peripheral blood T cells, stimulated with PMA for 48 h (PMA blasts), were incubated with IL-2 in the presence of incremental dosages of dexamethasone (Dex; 10(-5)-10(-9) M). Dex inhibited the IL-2-dependent proliferation of PMA blasts in a dose-dependent fashion (IC50, 5 x 10(-8) M). Cell surface expression of IL-2R alpha- and beta-chains as determined by immunofluorescence analysis was not affected by Dex. In addition, Scatchard plot analysis of 125I-labeled IL-2 showed that Dex did not affect the binding of IL-2, thus suggesting that inhibition is due to a postreceptor effect. Inhibition of T cell proliferation by Dex was associated with decreased IL-2-dependent tyrosine phosphorylation of several intracellular proteins and decreased phosphorylation of the retinoblastoma gene product Rb, a protein essential for controlling the progression of cells through the cell cycle. IL-2-dependent IL-2R alpha expression in PMA blasts and NF-kB induction in resting human T cells were also inhibited by Dex. These results demonstrate that glucocorticoids inhibit preactivated T cells by down-regulating signal transduction through IL-2R.