bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effect of an ectokinase inhibitor, K252b, on degranulation and Ca2+ signals of RBL-2H3 cells and human basophils.
Authors: Teshima R , Saito Y , Ikebuchi H , Rajiva De Silva N , Morita Y , Nakanishi M , Sawada J , Kitani S 
Published: January 2023

Abstract
We examined the effects of K252b, an ectoprotein kinase inhibitor of microbial origin, on the activation process of RBL-2H3 cells by cross-linking of IgE receptors by the endoplasmic reticulum Ca2+-ATPase inhibitor 2,5-di(tert-butyl)-1,4-hydroquinone or by the Ca2+ ionophore A23187. Analysis of phosphorylation of ectoproteins following IgE receptor cross-linking revealed that K252b mainly inhibited the phosphorylation of a 130-kDa protein. The inhibitor simultaneously inhibited degranulation and the sustained increase in the cytosolic calcium ion concentration even after addition of Ag. In contrast, K252b did not inhibit the increase in degranulation and cytosolic calcium ion concentration caused by stimulation with 2,5-di(tert-butyl)-1,4-hydroquinone and A23187. Permeation of K252b into RBL-2H3 cells, assessed by fluorescence intensity, was very low. K252b also inhibited degranulation caused by IgE receptor cross-linking in human basophils, but did not inhibit the degranulation caused by A23187. Thus, our findings suggest that the effects of K252b may be mediated by outer surface-bound or -anchored K252b-sensitive molecules on RBL-2H3 cells and human basophils, and that the phosphorylation of ectoprotein may involve a transmembrane influx of Ca2+ by IgE receptor cross-linking.
Fas-mediated liver damage in MRL hemopoietic chimeras undergoing lpr-mediated graft-versus-host disease.
Authors: Bobé P , Bonardelle D , Reynès M , Godeau F , Mahiou J , Joulin V , Kiger N 
Published: January 2023

Abstract
Fas is an apoptosis-signaling receptor that triggers cell death upon binding to its ligand (FasL). Autoimmune-prone MRL/lpr mice, characterized by a spontaneous mutation of Fas, exhibit a defect in the activation-induced cell death of mature T cells through a Fas-mediated pathway. As a consequence of this defect, activated T cells accumulating in this strain overexpress the FasL and can therefore mediate in vitro Fas-dependent cytotoxicity. To determine whether hepatic injury could be the result of an interaction between T lymphocytes bearing FasL and Fas-expressing liver cells, the livers of lethally irradiated MRL+/+ recipients reconstituted with MRL/lpr lymphoid cells were studied. After transfer of MRL/lpr spleen cells, livers were infiltrated by polyclonal CD8+ T lymphocytes of lpr origin with a peak on day 21 postgrafting. These donor-derived intrahepatic lymphocytes overexpressed the FasL and exerted in vitro Fas-mediated cytotoxicity against Fas+ thymocytes, which was specifically inhibited by soluble recombinant Fas in a dose-dependent manner. These intrahepatic lymphocytes induced apoptosis in vitro, irrespective of MHC restriction, in Fas-expressing primary cultured hepatocytes. Histologic examination of the liver revealed severe endothelialitis as well as periportal and intralobular infiltrations of activated lymphocytes with apoptotic hepatocytes in their vicinity. Simultaneously, liver damage was ascertained by elevated serum transaminase levels. These observations support the notion that an Ag-independent mechanism involving FasL may play a role in certain liver pathologies.
B cell antigen receptor-evoked calcium influx is enhanced in CD22-deficient B cell lines.
Authors: Nadler M J , McLean P A , Neel B G , Wortis H H 
Published: January 2023

Abstract
CD22 is a B cell membrane glycoprotein that, upon Ag receptor engagement, becomes rapidly tyrosyl phosphorylated and associates with several signaling molecules including Lyn, Syk, PLCgamma1, and the protein-tyrosine phosphatase, SHP-1. Two allelic forms of murine CD22 exist: CD22.1 is expressed in strains such as NZB and DBA/2, whereas CD22.2 is expressed in BALB/c and most other strains. WEHI-231 cells, which derive from a (BALB/c x NZB)F1 mouse, express one copy of each allele. Previous studies have proposed both positive and negative functions for CD22. We explored the role of CD22 in surface IgM Ag receptor signal transduction by examining signaling in three clonally independent WEHI-231 variants that have lost expression of the CD22.2 allele. This experimental design allowed us to assess the signaling functions of CD22 independent of its developmental role. These variants, which exhibit a 50% reduction of total surface CD22, are hyper-responsive to Ag receptor stimulation: several cellular proteins are hyperphosphorylated on tyrosyl residues and surface IgM-mediated calcium flux is markedly increased. Interestingly, the increased calcium response observed in CD22-deficient cells is due largely to enhanced calcium influx. Reconstitution of CD22 expression reduces these changes. The SHP-1/CD22 association is reduced in CD22-deficient cell lines and is restored by re-expression of CD22. Our results demonstrate that CD22 is a cell autonomous negative regulator of B cell Ag receptor signaling, and suggest that it regulates calcium entry via a mechanism downstream from or independent of calcium release from intracellular stores.
Regulation of B cell functions by C3a and C3a(desArg): suppression of TNF-alpha, IL-6, and the polyclonal immune response.
Authors: Fischer W H , Hugli T E 
Published: January 2023

Abstract
Regulation of the humoral immune response by fragments of complement component C3 has been shown to play an important role in host defense. In this study, we investigated expression of the C3a receptor (C3aR) by human B cells and the effects of C3a and C3a(desArg) on IgG, TNF-alpha, and IL-6 production in Staphylococcus aureus Cowan strain I (SAC)/IL-2-activated B cells. Here we report that tonsil-derived human B cells express a C3aR that is indistinguishable at both the protein (by flow cytometry) and mRNA (by reverse transcription-PCR) levels from that on human neutrophils or on C3aR-transfected mouse L cells. Incubation of SAC/IL-2-activated B cells with C3a or C3a(desArg) resulted in a dose-dependent suppression of the polyclonal immune response. Under these same experimental conditions, IL-6 and TNF-alpha release was suppressed in a dose-dependent manner. Kinetic studies with SAC/IL-2-activated B cells revealed that C3a must be present at initiation of the culture to exert its suppressive effect on IgG and IL-6 production. These results demonstrate that tonsil-derived B cells express the C3aR and that C3a as well as C3a(desArg) have a direct immunomodulatory effect on these cells.
Rigidification of the alpha2 helix of an MHC class I molecule by a valine to proline mutation in position 165 does not prevent peptide-specific antigen presentation.
Authors: Plaksin D , Polakova K , Mage M G , Margulies D H 
Published: January 2023

Abstract
Although classical MHC class I glycoproteins bind peptide Ags for display at the cell surface, some MHC class I-related molecules such as the neonatal Fc receptor (FcRn) execute their function without binding peptide ligands. The three-dimensional structure of the FcRn suggested that a substitution of the conserved valine at position 165 of the alpha2 helix by proline contributed to a kink in the position of this helix relative to the alpha1 helix, and resulted in closing of the potential peptide-binding cleft. To test the contribution of proline 165 to the occlusion of the cleft and the binding of potential antigenic peptides, we introduced this mutation into the classical murine MHC class I molecule, H-2Dd, and characterized the ability of such a mutant to present peptide Ags to either a peptide-specific, H-2Dd-restricted T cell hybridoma (B4.2.3), or an allospecific, peptide-dependent, T cell hybridoma (3DT52.5.8). We show that the V165P mutation, expressed at the cell surface either in H-2Dd or in a single chain membrane version of H-2Dd, fails to eliminate recognition of the peptide/MHC complexes by two different T cells. Evaluation of a panel of synthetic substituted peptides suggests that subtle differences in the fine specificity of presentation can be discerned. Thus, the proline substitution at position 165 of FcRn and some other class I-like molecules is not the sole cause of the lack of peptide presentation.
T cell receptor-mediated signaling is defective in T cells obtained from patients with primary intracranial tumors.
Authors: Morford L A , Elliott L H , Carlson S L , Brooks W H , Roszman T L 
Published: January 2023

Abstract
It has been well established that patients with malignant glioblastomas exhibit T cell anergy. In this report, we further investigate the nature of this T cell anergy. The results demonstrate that tumor size but not location correlates with decreased mitogen or anti-CD3 mAb responsiveness of T cells obtained from patients. Stimulation of the TCR/CD3 complex on these patients' T cells revealed defects in early transmembrane signaling. Both PHA and anti-CD3 mAb activated PBL and T cells obtained from patients exhibited a marked decrease in the tyrosine phosphorylation of a number of proteins. In particular, decreased phosphorylation of pp100 and phospholipase Cgamma1 (PLCgamma1) was observed. In addition, PLCgamma1 and p56(lck) protein levels were dramatically reduced in T cells obtained from patients harboring a glioma. In contrast, the protein levels of p59(fyn) were normal or only slightly reduced in T cells obtained from patients with gliomas. Quantitation of free intracellular calcium concentrations ([Ca2+]i) after mitogen (PHA) stimulation or ionomycin treatment of T cells obtained from patients revealed that they mobilize less calcium than do T cells obtained from normal subjects. Stimulation of T cells obtained from patients with PMA and ionomycin, which should bypass the requirement for PLCgamma1 activation as well as directly activate the p21(ras) signaling pathway, did not restore the proliferative capacity of these T cells to normal levels. These results indicate that the anergy observed in T cells obtained from these patients is a consequence of one or more defects in the early transmembrane signaling events associated with TCR/CD3 stimulation.
TNF Receptor-Deficient Mice Reveal Striking Differences Between Several Models of Thymocyte Negative Selection
Authors: Page Dawne M. , Roberts Edda M. , Peschon Jacques J. , Hedrick Stephen M. 
Published: January 2023

Abstract
Central tolerance depends upon Ag-mediated cell death in developing thymocytes. However, the mechanism of induced death is poorly understood. Among the known death-inducing proteins, TNF was previously found to be constitutively expressed in the thymus. The role of TNF in thymocyte negative selection was therefore investigated using TNF receptor (TNFR)-deficient mice containing a TCR transgene. TNFR-deficient mice displayed aberrant negative selection in two models: an in vitro system in which APC are cultured with thymocytes, and a popular in vivo system in which mice are treated with anti-CD3 Abs. In contrast, TNFR-deficient mice displayed normal thymocyte deletion in two Ag-induced in vivo models of negative selection. Current models of negative selection and the role of TNFR family members in this process are discussed in light of these results.
Detection and Delineation of CXCR-4 (Fusin) as an Entry and Fusion Cofactor for T Cell-Tropic HIV-1 by Three Different Monoclonal Antibodies
Authors: Hori Toshiyuki , Sakaida Hitoshi , Sato Akihiko , Nakajima Toshihiro , Shida Hisatoshi , Yoshie Osamu , Uchiyama Takashi 
Published: January 2023

Abstract
A chemokine receptor, CXCR-4, has been identified as an entry cofactor for T cell line-tropic (T-tropic) HIV-1. To detect expression of CXCR-4 at the single cell level and dissect postbinding events of HIV-1 infection, we generated three mAbs against human CXCR-4. These mAbs inhibited SDF-1-induced intracellular Ca2+ mobilization, and one of the mAbs immunoprecipitated a specific 47-kDa component from CXCR-4+ cells. Flow cytometric analysis showed that most human cell lines examined expressed CXCR-4. A fraction of normal PBMC expressed CXCR-4, but neutrophils were negative. Two-color analysis revealed that the majority of T cells, virtually all B cells, and all monocytes expressed CXCR-4, while it was only weakly present on NK cells. Thus, expression of CXCR-4 is not ubiquitous but cell type specific in hemopoietic cells. The three mAbs were shown to suppress cell fusion mediated by envelope proteins of a T-tropic NL432 virus but not by those of an M-tropic JRCSF virus. Likewise, they suppressed infection of NL432 but not that of an M-tropic NL162 virus. In both cases it was noted that the suppressive activity varied considerably among the mAbs. These data confirmed that CXCR-4 is directly involved in env-mediated entry and fusion of T-tropic HIV-1 and suggest that the epitopes on CXCR-4 recognized by the three mAbs may have different roles in interaction with the envelope proteins of T-tropic HIV-1.
Staphylococcal Enterotoxin D Is a Promiscuous Superantigen Offering Multiple Modes of Interactions With the MHC Class II Receptors
Authors: Al-Daccak Reem , Mehindate Khalil , Damdoumi Farida , Etongué-Mayer Pierre , Nilsson Helen , Antonsson Per , Sundström Michael , Dohlsten Mikael , Sékaly Rafick-Pierre , Mourad Walid 
Published: January 2023

Abstract
Dimerization of MHC class II molecules on the cell surface of human THP-1 monocytic cell line is a requirement for staphylococcal superantigen (SAG)-induced cytokine gene expression. The capacities of various SAG to induce this response are governed by their modes of interaction with MHC class II molecules. Staphylococcal enterotoxin A (SEA), with its two binding sites, dimerizes MHC class II molecules and subsequently induces cytokine gene expression in THP-1 cells. Here, we demonstrate that staphylococcal enterotoxin D (SED) and staphylococcal enterotoxin E (SEE) induce, similarly, IL-1β and TNF-α gene expression in these cells. Using mutated toxins that lost their binding site with the MHC class II α- or β-chain, we demonstrate that this response is also mediated by the dimerization of MHC class II molecules through two binding sites. Furthermore, SED forms Zn2+-dependent homodimers that allow multiple modes of MHC class II clustering, including ligation of α-chains (α/α), β-chains (β/β), or the α- and β-chains of two different class II molecules. The β/β interaction following Zn2+-dependent SED/SED homodimer formation seems to be mediated by the appearance of a novel binding site on SED that interacts with histidine 81 of the MHC class II β-chain. The different modes of SED interactions also influence SED-induced T cell activation where simultaneous ligation of the α- and β-chains is essential for optimal response. These various modes of SED binding may be used to preserve bivalency regardless of variability in the MHC class II α/β/peptide complexes.
A Two-Step Mechanism for Recruitment of Pip by PU.1
Authors: Perkel Jeffrey M. , Atchison Michael L. 
Published: January 2023

Abstract
Transcription of the Ig κ light chain gene is controlled in part by the 3′ κ enhancer. Two of the proteins that bind to the 3′ enhancer, PU.1 and Pip, show tissue-restricted expression and may be responsible for the tissue specificity of 3′ enhancer activity. PU.1 alone can bind to DNA; however, Pip cannot bind to its 3′ enhancer site in electrophoretic mobility shift assays, unless recruited by PU.1. Previously, we showed that the PU.1 PEST domain (rich in the amino acids proline, glutamate, serine, and threonine; sequences 118–160) is necessary for Pip recruitment to DNA. Here we used detailed mutagenic analyzes of PU.1 to more precisely identify sequences required for Pip recruitment by electrophoretic mobility shift assay. We found that mutation of three segments within the PU.1 PEST domain (118–125, 133–139, and 141–147) modulated the efficiency of Pip recruitment, while mutation of sequences between residues 88–118 and 154–168 had no effect. Interestingly, we found that the PU.1 ETS domain (residues 170 to 255) is both necessary and sufficient for Pip interaction in solution and that other ETS domain proteins can physically interact with Pip as well. Our results suggest that Pip recruitment to DNA by PU.1 occurs via a two-step mechanism. First, a physical interaction that is not sufficient to recruit Pip occurs via the PU.1 ETS domain. Second, a conformational change in the PU.1 PEST domain, apparently mediated by serine phosphorylation, induces a conformational change in Pip enabling it to bind to DNA. We also show that the PU.1 PEST domain does not target PU.1 for rapid turnover.
The Role of Recombinant Murine IL-12 and IFN-γ in the Pathogenesis of a Murine Systemic Candida albicans Infection
Authors: Lavigne Liz M. , Schopf Lisa R. , Chung Charles L. , Maylor Rich , Sypek Joseph P. 
Published: January 2023

Abstract
Studies on murine candidiasis suggest that resistance to disease is linked to a Th1 response and production of IFN-γ, while failure to elicit protection is associated with a Th2 response and production of IL-4 and IL-10. Experimental infection of C57BL/6 mice, IL-12 treatment of these mice, or both infection and IL-12 treatment resulted in a characteristic Th1 cytokine mRNA profile as measured by quantitative competitive PCR. Specifically, little or no IL-4 transcripts were detected, while IFN-γ message was elevated, particularly with IL-12 treatment. Despite its role in driving increased IFN-γ expression and production, IL-12 treatment, paradoxically, promoted disease progression in our model. Therefore, we examined the effect of IFN-γ neutralization on IL-12-induced susceptibility to infection. None of the systemically infected mice receiving IL-12 alone survived, while IL-12- and anti-IFN-γ-treated mice had a 70% survival rate, similar to that after infection alone. These results suggested that IFN-γ induced by IL-12 treatment contributed to lethality. However, in separate studies, IFN-γ knockout mice were more susceptible to infection than their wild-type counterparts, suggesting that IFN-γ is required for resistance. Nonetheless, infected IFN-γ knockout mice treated with recombinant murine IL-12 exhibited enhanced resistance, suggesting that the toxicities observed with IL-12 are directly attributable to IFN-γ and that an optimal immune response to Candida infections necessitates a finely tuned balance of IFN-γ production. Thus, we propose that although IFN-γ can drive resistance, the overproduction of IFN-γ during candidiasis, mediated by IL-12 administration, leads to enhanced susceptibility.
Accommodation and T-Independent B Cell Tolerance in Rats With Long Term Surviving Hamster Heart Xenografts
Authors: Lin Yuan , Vandeputte Michel , Waer Mark 
Published: January 2023

Abstract
It was previously reported that treatment with leflunomide (LF; 10 mg/kg/day) together with cyclosporine (CsA; 10 mg/kg/day) resulted in long term survival of hamster heart xenografts (Xg) in rats and that LF could be withdrawn 2 to 4 wk after transplantation. To study the mechanisms allowing withdrawal of LF, second hamster heart Xgs were transplanted 6 wk after the first xenograft. Only the rats that received LF for 4 wk accepted second Xgs (&gt;30 days; n = 5). Hence, after 4 wk of LF, the rats developed partial B cell tolerance, as they were unable to produce T-independent (CsA-resistant) XAbs. Rejection of second Xgs (2–4 days; n = 5) in the 2-wk LF group resulted in the formation of IgM xenoantibodies (XAbs) localizing together with complement within rejected grafts. However, these XAbs did not affect first Xgs, suggesting that the latter Xgs became resistant to this IgM XAb-mediated rejection, a phenomenon referred to as accommodation. Accommodation was further confirmed as adoptive transfer of IgM XAbs, which resulted in hyperacute Xg rejection in naive rats (&lt;1 h; n = 5), did not cause rejection in long term survivors (&gt;30 days; n = 4). This was associated with a down-regulation of the expression on the graft endothelial cells of adhesion molecules (believed to be important expressers of xenogeneic epitopes), such as P- and E-selectins. Interestingly, these adhesion molecules reappeared after retransplanting the accommodated Xgs to naive recipients. In conclusion, depending on the duration of the LF treatment, long term survival of hamster hearts in CsA-treated rats is based in part on accommodation and in part on T-independent B cell tolerance.
Inhibition of Proinflammatory Molecule Production by Adenovirus-Mediated Expression of a Nuclear Factor κB Super-Repressor in Human Intestinal Epithelial Cells
Authors: Jobin Christian , Panja Asit , Hellerbrand Claus , Iimuro Yuji , Didonato Joseph , Brenner David A. , Sartor R. Balfour 
Published: January 2023

AbstractNF-κB plays a major role in the transcriptional regulation of many proinflammatory genes in multiple cell lineages, including intestinal epithelial cells (IEC). Activation of NF-κB requires both phosphorylation and degradation of its natural cytoplasmic inhibitor, IκB. We tested whether a super-repressor of NF-κB activity, which is a mutated nondegradable IκBα resistant to phosphorylation and degradation, could be delivered into IEC using an adenoviral vector (Ad5IκB) and determined the anti-inflammatory potential of this inhibitor following different stimuli. We showed for the first time that recombinant adenovirus efficiently infected (&gt;80%) transformed as well as primary IEC. Cytoplasmic levels of the NF-κB super-repressor protein were more than 50-fold higher than those of endogenous IκB, and this mutated IκB was resistant to IL-1β-induced degradation. Immunofluorescent RelA nuclear staining was strongly inhibited in Ad5IκB-infected IEC compared with control Ad5LacZ, and NF-κB, but not AP-1 binding activity, was reduced by more than 70% as measured by electrophoretic mobility shift assay (EMSA). Induction of inducible nitric-oxide synthase (iNOS), IL-1β, and IL-8 genes by IL-1β, TNF-α, or PMA was blocked in Ad5IκB-infected cells but not in Ad5LacZ controls as assayed by RT-PCR and ELISA. In addition, IL-1β-induced IL-8 secretion was totally inhibited by Ad5IκB in primary colonic IEC. We conclude that an adenoviral vector efficiently transfers a nondegradable IκB in both transformed and native IEC. The strong inhibition of NF-κB activity and the resulting down-regulation of multiple proinflammatory molecules by Ad5IκB suggests an exciting approach for in vivo intestinal gene therapy and illustrates the key role of NF-κB in transcriptional regulation of the inflammatory phenotype of IEC.