bluuebunny/crossref_metadata_2025_split huggingface dataset:
The β2-Adrenergic Agonist Salbutamol Is a Potent Suppressor of Established Collagen-Induced Arthritis: Mechanisms of Action
Authors: Malfait Anne-Marie , Malik Angela S. , Marinova-Mutafchieva Lilia , Butler Debra M. , Maini Ravinder N. , Feldmann Marc 
Published: January 2023

Abstract
The therapeutic potential of salbutamol, a β2-adrenergic agonist, was explored in collagen-induced arthritis. This study was based on a report that salbutamol, by elevating intracellular cAMP, inhibits IL-12 production by macrophages and dendritic cells, thus preventing Th1 development. Ten-week-old male DBA/1 mice were immunized by intradermal injection of type II collagen in CFA. Arthritis developed 15–30 days later and the mice were treated after onset of disease with salbutamol, 200 μg i.p. After 10 days, the mice were sacrificed, and the hind paws were evaluated histologically. Salbutamol, 200 μg daily or every other day, had a profound therapeutic effect on the clinical progression of arthritis, as assessed by clinical score and paw thickness. The therapeutic effect was dose dependent. Daily administration of 200 μg of salbutamol offered the best protection against joint damage, as assessed by histology. In vitro, salbutamol reduced IL-12 and TNF-α release by peritoneal macrophages in a dose-dependent manner, as well as TNF release by synovial cells from arthritic mice. Ex vivo, draining lymph node cells of the salbutamol-treated arthritic mice showed a diminished CII-specific IFN-γ production and proliferation. In vivo, salbutamol specifically blocked mast cell degranulation in joint tissues. In conclusion, salbutamol has important effects on the immunoinflammatory response and a significant therapeutic action in collagen-induced arthritis.
Cutting Edge: Contrasting Roles of TNF Receptor-Associated Factor 2 (TRAF2) and TRAF3 in CD40-Activated B Lymphocyte Differentiation
Authors: Hostager Bruce S. , Bishop Gail A. 
Published: January 2023

Abstract
In B lymphocytes, CD40 signals contribute to the activation of Ab secretion, isotype switching, T cell costimulation, and immunological memory. TRAF proteins appear to be important components of the CD40 signal transduction complex, but their roles in the activation of B cell effector functions are poorly understood. We examined the contributions of TNF receptor-associated factor 2 (TRAF2) and TRAF3 to CD40-activated differentiation in mouse B cells transfected with inducible TRAF and dominant-negative TRAF cDNAs. We find that binding of TRAF2 and TRAF3 to CD40 is not required for the induction of Ab secretion, but that both TRAF molecules can regulate the activation process. We demonstrate a negative regulatory role for TRAF3 and that this activity is dependent on the availability of an intact TRAF3-binding site in the cytoplasmic domain of CD40. In contrast, TRAF2 appears to play a positive role in B cell differentiation, and this activity is apparent even when its binding site on CD40 is disrupted.
Potential Role of CD4+ T Cell-Mediated Apoptosis of Activated Astrocytes in Theiler’s Virus-Induced Demyelination
Authors: Palma JoAnn P. , Yauch Robert L. , Lang Sharon , Kim Byung S. 
Published: January 2023

Abstract
Intracerebral inoculation of Theiler’s murine encephalomyelitis virus (TMEV) into susceptible mouse strains results in a chronic, immune-mediated demyelinating disease similar to human multiple sclerosis. Here, we examined the role of astrocytes as an APC population in TMEV-induced demyelination and assessed the potential consequences of T cell activation following Ag presentation. IFN-γ-pretreated astrocytes were able to process and present all the predominant T cell epitopes of TMEV to virus-specific T cell hybridomas, clones, as well as bulk T cells. Despite low levels of proliferation of T cells due to prostaglandins produced by astrocytes, such Ag presentation by activated astrocytes induced the production of IFN-γ, a representative proinflammatory cytokine, in TMEV-specific Th cell clones derived from the CNS of virus-infected mice. Furthermore, these Th cell clones mediate lysis of the astrocytes in vitro in a Fas-dependent mechanism. TUNEL staining of CNS tissue demonstrates the presence of apoptotic GFAP+ cells in the white matter of TMEV-infected mice. These results strongly suggest that astrocytes could play an important role in the pathogenesis of TMEV-induced demyelination by activating T cells, subsequently leading to T cell-mediated apoptosis of astrocytes and thereby compromising the blood-brain barrier.
ICAM-2 and a Peptide from Its Binding Domain Are Efficient Activators of Leukocyte Adhesion and Integrin Affinity
Authors: Kotovuori Annika , Pessa-Morikawa Tiina , Kotovuori Pekka , Nortamo Pekka , Gahmberg Carl G. 
Published: January 2023

AbstractCell adhesion mediated by the CD11/CD18 integrins and their ligands, the ICAMs, is required for many leukocyte functions. In resting cells the integrins are nonadhesive, but when activated they become adhesive for their ligands. Previous findings have shown that a peptide derived from the first Ig domain of ICAM-2 (P1) binds to LFA-1 (CD11a/CD18) and Mac-1 (CD11b/CD18) and activates leukocyte aggregation. Because its mechanism of action has remained poorly understood, we have now studied the peptide-induced ligand binding in detail. Here we show that P1 was able to induce CD11/CD18-dependent adhesion of human T lymphocytes to immobilized, purified ICAM-1, -2, and -3. The optimal peptide concentration was 150 μg/ml, whereas concentrations higher than 400 μg/ml did not have any stimulatory effect. The increase in adhesion was detectable within 10 min of treatment with the peptide; it was dependent on energy, divalent cations, temperature, and an intact cytoskeleton but was unaffected by protein kinase C and protein tyrosine kinase inhibitors. Peptide treatment resulted in strong stimulation of the binding of soluble, recombinant ICAMs to T lymphocytes, showing that the integrin affinity toward its ligands was increased. Importantly, soluble ICAM-2Fc was also able to induce T lymphocyte adhesion to purified ICAM-1, -2, and -3, and it was a more potent stimulatory molecule than ICAM-1Fc or ICAM-3Fc.
IL-12 Delivery from Recombinant Vaccinia Virus Attenuates the Vector and Enhances the Cellular Immune Response Against HIV-1 Env in a Dose-Dependent Manner
Authors: Gherardi M. Magdalena , Ramirez Juan C. , Rodríguez Dolores , Rodríguez Juan R. , Sano Gen-Ichiro , Zavala Fidel , Esteban Mariano 
Published: January 2023

Abstract
To develop vaccination strategies against HIV-1 infection aimed to specifically enhance the cell-mediated immunity (CMI), we have engineered vaccinia virus (VV) recombinants expressing HIV-1 Env (rVVenv) and murine IL-12 (rVVlucIL-12) genes or coexpressing both genes (rVVenvIL-12). In mice inoculated with rVVlucIL-12 there is a rapid clearance of the virus, and this correlates with the induction of high levels of IL-12 and IFN-γ in serum and spleen early after infection. Enzyme-linked immunospot analysis of mice inoculated with rVVlucIL-12, revealed a nearly 2-fold increase in the number of specific anti-VV CD8+ T cells compared with that in mice given control rVV, and the serum Ab response was biased in favor of a Th1 response. An enhancement of about 2-fold in the number of anti-gp160 IFN-γ-secreting CD8+ T cells was observed in mice inoculated with rVVenvIL-12, when a dose of 1 × 107 PFU/mouse was used, but this enhancement was not observed when mice were given 5 × 107 PFU. This variation with virus dosage was confirmed in mice immunized simultaneously with different multiplicities of rVV expressing singly the env or IL-12 genes. The highest specific CMI was obtained in mice coadministered a low dose (2 × 104 PFU) of rVVlucIL-12 and 1 × 107 PFU of rVVenv. Our findings provide evidence for specific enhancement of the CMI to HIV-1 Env by the differential expression of IL-12 and env genes delivered from VV recombinants. This approach can be of wide vaccination interest as a means to improve immune responses to other Ags.
Differential Regulation of the IL-12 p40 Promoter and of p40 Secretion by CpG DNA and Lipopolysaccharide
Authors: Cowdery John S. , Boerth Nancy J. , Norian Lyse A. , Myung Peggy S. , Koretzky Gary A. 
Published: January 2023

AbstractChallenge of macrophages with DNA containing an internal CpG motif results in IL-12 p40 secretion. In the presence of IFN-γ, CpG DNA induces more p40 secretion than does LPS. In the RAW 264 macrophage cell line, both CpG DNA and LPS activate a p40 promoter-reporter construct, and the promoter response to either agent is augmented 2- to 5-fold by IFN-γ. While either LPS or CpG DNA induces p40 promoter activity, only CpG DNA induces an increase in p40 mRNA or protein secretion. Even though IFN-γ augmented LPS-driven p40 promoter activity in RAW 264 cells, the combination of IFN-γ and LPS induced less p40 mRNA or protein secretion than the combination of IFN-γ and CpG DNA. The ability of IFN-γ to augment LPS or CpG DNA-induced p40 promoter activation was observed with truncation mutants of the IL-12 promoter containing as few as 250 bp 5′ of the TATA box. Although LPS alone is a poor inducer of p40 transcription, both LPS and CpG DNA induce similar nuclear translocation of NF-κB. This binding is not augmented by costimulation with IFN-γ. Thus, CpG DNA induces p40 transcription by a mechanism that includes NF-κB translocation; however, CpG DNA appears to induce other factor(s) necessary for p40 transcription. These results illustrate fundamental differences between CpG DNA and LPS with respect to activation of IL-12 p40 secretion.
Lysosomal Accumulation and Recycling of Lipopolysaccharide to the Cell Surface of Murine Macrophages, an In Vitro and In Vivo Study
Authors: Forestier Claire , Moreno Edgardo , Pizarro-Cerda Javier , Gorvel Jean-Pierre 
Published: January 2023

Abstract
In this study, we detailed in a time-dependent manner the trafficking, the recycling, and the structural fate of Brucella abortus LPS in murine peritoneal macrophages by immunofluorescence, ELISA, and biochemical analyses. The intracellular pathway of B. abortus LPS, a nonclassical endotoxin, was investigated both in vivo after LPS injection in the peritoneal cavity of mice and in vitro after LPS incubation with macrophages. We also followed LPS trafficking after infection of macrophages with B. abortus strain 19. After binding to the cell surface and internalization, Brucella LPS is routed from early endosomes to lysosomes with unusual slow kinetics. It accumulates there for at least 24 h. Later, LPS leaves lysosomes and reaches the macrophage cell surface. This recycling pathway is also observed for LPS released by Brucella S19 following in vitro infection. Indeed, by 72 h postinfection, bacteria are degraded by macrophages and LPS is located inside lysosomes dispersed at the cell periphery. From 72 h onward, LPS is gradually detected at the plasma membrane. In each case, the LPS present at the cell surface is found in large clusters with the O-chain facing the extracellular medium. Both the antigenicity and heterogenicity of the O-chain moiety are preserved during the intracellular trafficking. We demonstrate that LPS is not cleared by macrophages either in vitro or in vivo after 3 mo, exposing its immunogenic moiety toward the extracellular medium.
IL-18, a Novel Immunoregulatory Cytokine, Is Up-Regulated in Crohn’s Disease: Expression and Localization in Intestinal Mucosal Cells
Authors: Pizarro Theresa T. , Michie Miette H. , Bentz Marcia , Woraratanadharm Jan , Smith Michael F. , Foley Eugene , Moskaluk Christopher A. , Bickston Stephen J. , Cominelli Fabio 
Published: January 2023

Abstract
IL-18, a novel immunoregulatory cytokine with potent IFN-γ-inducing activities, may play an important role in Th1-mediated chronic inflammatory disorders. The aim of the present study was to characterize the expression and localization of IL-18 in colonic specimens and isolated mucosal cell populations from patients with Crohn’s disease (CD), a prototypic Th1-mediated disorder. Using a semiquantitative RT-PCR protocol, IL-18 mRNA transcripts were found to be increased in freshly isolated intestinal epithelial cells (IEC) and lamina propria mononuclear cells (LPMC) from CD compared with ulcerative colitis (UC) and noninflamed control (cont) patients, and were more abundant in IEC compared with LPMC. Immunohistochemical analysis of surgically resected colonic tissues localized IL-18 to both LPMC (specifically, macrophages and dendritic cells) as well as IEC. Staining was more intense in CD compared with UC and cont, and in involved (inv) vs noninvolved (n inv) areas. Western blot analysis revealed that an 18.3-kDa band, consistent with both recombinant and mature human IL-18 protein, was found predominantly in CD vs UC intestinal mucosal biopsies; a second band of 24 kDa, consistent with the inactive IL-18 precursor, was detected in n inv areas from both CD and UC biopsies and was the sole form found in noninflamed cont. To our knowledge, this report is the first describing increased expression of IL-18 in a human Th1-mediated chronic inflammatory disease. In addition, our studies further support the concept that IEC and dendritic cells may possess important immunoregulatory functions in both normal, as well as pathological, mucosal immunity.
Does IgE Bind to and Activate Eosinophils from Patients with Allergy?
Authors: Kita Hirohito , Kaneko Masayuki , Bartemes Kathleen R. , Weiler Deborah A. , Schimming Andrew W. , Reed Charles E. , Gleich Gerald J. 
Published: January 2023

Abstract
Human eosinophils have been reported to express both the mRNA and protein for the high affinity IgE receptor (FcεRI); it is speculated that this receptor plays a role in eosinophil mediator release in allergic diseases. However, questions still remain. How much of the FcεRI protein is actually expressed on the cell surface of the eosinophil? If they are present, are these IgE receptors associated with effector functions of eosinophils? To address these issues, we studied blood eosinophils from patients with ragweed hay fever. A high level of low affinity IgG receptor (FcγRII, CD32), but no expression of FcεRI, was detectable on the eosinophil surface by standard FACS analysis. However, after in vitro sensitization with biotinylated chimeric IgE (cIgE), cell-bound cIgE was detected by PE-conjugated streptavidin. This cIgE binding was partially inhibited by anti-FcεRI mAb, suggesting that eosinophils do express minimal amounts of FcεRI detectable only by a sensitive method. Indeed, FACS analysis of whole blood showed that eosinophils express ∼0.5% of the FcεRI that basophils express. When stimulated with human IgE or anti-human IgE, these eosinophils did not exert effector functions; there was neither production of leukotriene C4 or superoxide anion nor any detectable degranulation response. In contrast, eosinophils possessed membrane-bound human IgG and showed functional responses when stimulated with human IgG or anti-human IgG. Thus, IgG and/or cytokines, such as IL-5, appear to be more important for eosinophil activation in allergic diseases than IgE.
Cutting Edge: The CD45 Tyrosine Phosphatase Is an Inhibitor of Lck Activity in Thymocytes
Authors: D’Oro Ugo , Ashwell Jonathan D. 
Published: January 2023

AbstractA widely accepted model for regulation of the Lck tyrosine kinase is that it is activated by CD45-mediated dephosphorylation of its COOH-terminal negative regulatory tyrosine (Tyr505). Previous work from our laboratory, however, found that despite hyperphosphorylation of Tyr505, the activity of Lck from CD45− T cell lines was actually increased due to hyperphosphorylation of the positive regulatory tyrosine, residue 394. To avoid potential complications introduced by transformed cells, in this study we have characterized the effect of CD45 on Lck activity in normal cells. Lck in thymocytes from CD45−/− mice was hyperphosphorylated on tyrosine residues. Importantly, and in disagreement with the model that CD45 only activates Lck in vivo, the kinase activity of Lck from cells lacking CD45 was substantially increased. These results support a model in which CD45 dephosphorylates both Tyr505 and Tyr394, the net effect in normal thymocytes being a decrease in enzymatic activity.
Fas Ligand and Fas Are Expressed Constitutively in Human Astrocytes and the Expression Increases with IL-1, IL-6, TNF-α, or IFN-γ
Authors: Choi Chulhee , Park Joo Young , Lee Jeonggi , Lim Jung-Hee , Shin Eui-Cheol , Ahn Young Soo , Kim Chul-Hoon , Kim Se-Jong , Kim Joo-Deuk , Choi Il Saing , Choi In-Hong 
Published: January 2023

Abstract
Fas ligand (FasL) and Fas are mediators of apoptosis, which are implicated in the peripheral deletion of autoimmune cells, activation-induced T cell death, and cytotoxicity mediated by CD8+ T cells. Fas is also believed to be involved in several central nervous system diseases, but until now, the effector cells expressing FasL in the brain have not been identified. We investigated the expression levels of Fas and FasL with the stimulation of cytokines and the possible effector cells targeting Fas-bearing cells. Our data demonstrated that: 1) FasL is expressed constitutively on astrocytes taken from a fetus or an adult and that its expression increases when these cells are treated with IL-1, IL-6, or TNF-α in which the pretreatment of IFN-γ triggers astrocytes to express more FasL; 2) astrocytes induce apoptosis in MOLT-4 cells through FasL; 3) Fas is also expressed constitutively and is up-regulated by IL-1, IL-6, or TNF-α in which the pretreatment of IFN-γ triggers astrocytes to express more Fas; 4) apoptosis occurs when fetal astrocytes are treated with agonistic anti-Fas IgM Ab after culture with IFN-γ and TNF-α; and 5) TNF-related apoptosis inducing ligand is up-regulated in fetal astrocytes with stimuli of IL-1 or TNF-α. These findings suggest a possible role of astrocytes in the induction of apoptosis in central nervous system diseases.
The Role of CD80, CD86, and CTLA4 in Alloimmune Responses and the Induction of Long-Term Allograft Survival
Authors: Judge Thomas A. , Wu Zihou , Zheng Xiang-Guang , Sharpe Arlene H. , Sayegh Mohamed H. , Turka Laurence A. 
Published: January 2023

AbstractBlocking the interaction of the CD28 costimulatory receptor with its ligands, CD80 and CD86, inhibits in vivo immune responses, such as allograft rejection, and in some instances induces tolerance. Previously, we found that CTLA4Ig, which blocks the CD28/CTLA-4 (CD152) ligands CD80 and CD86, can be used to induce transplantation tolerance to vascularized allografts. Recent data suggest that an intact CD152-negative signaling pathway is essential for induction of tolerance to nominal Ags. Here, we show that blockade of CD152 using an anti-CD152 mAb at the time of transplantation prevents the induction of long-term allograft survival by agents that target CD80 and CD86. In contrast, CD152 signals are not required for the maintenance of established graft survival. We also report for the first time that blockade of CD86 alone can induce long-term graft survival. This requires that anti-CD86 mAb is given on the day of transplantation and also depends upon an intact CD152 pathway. This result, plus experiments using CD80-deficient mice, suggests a dominant role for CD80 molecules on donor cells as the relevant ligand for CD152. We additionally find that blockade of CD152 at the time of transplantation does not interfere with the effectiveness of anti-CD154 mAbs, suggesting distinct mechanisms for inhibition of graft rejection by blocking the CD28 vs CD154 pathways.
Identification of Tyrosine Phosphorylation Sites in the CD28 Cytoplasmic Domain and Their Role in the Costimulation of Jurkat T Cells
Authors: Sadra Ali , Cinek Tomas , Arellano Jerry L. , Shi Jia , Truitt Kenneth E. , Imboden John B. 
Published: January 2023

Abstract
The cytoplasmic domain of CD28 contains four tyrosine residues. Because signal transduction by CD28 appears to involve its tyrosine phosphorylation, we determined sites of CD28 tyrosine phosphorylation using mutants of mouse CD28 that retained tyrosine at one position, with the remaining three positions mutated to phenylalanine. When expressed in Jurkat cells and stimulated by mAb, only the mutants with tyrosine at position 170 or 188 were tyrosine phosphorylated. Phosphorylation of Tyr170 recruits phosphatidylinositol 3-kinase to CD28. Tyr188 has not been associated with any specific signaling event, but we found that ligation of CD28 by the natural ligand B7.2 also induced phosphorylation of Tyr188, suggesting that this event is of physiological importance. Consistent with that possibility, mutation of Tyr188 to phenylalanine severely impaired the ability of mouse CD28 to deliver a costimulus for the expression of CD69 and the production of IL-2. The functional consequences of the mutation of Tyr188 were unique; mutation of the other three tyrosines, individually or in combination, did not impair costimulation. Therefore, of the four CD28 tyrosine residues only Tyr188 is required for signaling in Jurkat cells, suggesting that its phosphorylation is a key event in the costimulation of T cells.
Evidence of Positive Cross-Regulation on Th1 by Th2 and Antigen-Presenting Cells: Effects on Th1 Induced by IL-4 and IL-12
Authors: Oriss Timothy B. , McCarthy Susan A. , Campana Martha A. K. , Morel Penelope A. 
Published: January 2023

AbstractThe response of Th cells to cytokines is normally strictly regulated, such that following antigenic stimulation, Th cells respond for only a short period of time, after which they become refractory to cytokine-mediated effects. IL-12, a costimulator of Th1 having no proliferation-inducing capacity of its own, allows Th1 clones and lines to respond to IL-4 when they would otherwise be unable to respond to this cytokine. Cells that have proliferated in response to IL-4 plus IL-12 are fully able to be subsequently activated by specific Ag and APC. Additionally, the response to IL-4 of Th1 effector cells derived from normal murine spleen is enhanced significantly by IL-12. Furthermore, in the presence of IL-12, stimulated Th2 can induce proliferation of Th1 via IL-4 production, in a dual chamber culture system. We hypothesize that the effects of IL-4 and IL-12 represent a novel, positive cross-regulatory pathway that acts on Th1, and is mediated by Th2 (the IL-4 source) and APC (the IL-12 source). We propose this as a way for a Th2 immune response to positively influence an ongoing or waning Th1 response.
Positive Recognition of MHC Class I Molecules by the Ly49D Receptor of Murine NK Cells
Authors: George Thaddeus C. , Mason Llewellyn H. , Ortaldo John R. , Kumar Vinay , Bennett Michael 
Published: January 2023

AbstractMembers of the murine Ly49 family of receptors have been shown to inhibit and activate NK cell function. Subsets of Ly49-expressing NK cells mediate the rejection of bone marrow cell allografts and the lysis of allogeneic lymphoblasts. In this report we have studied Ly49-mediated positive and negative signaling in an in vitro cytotoxicity assay using sorted NK cell subsets as effectors and a panel of 51Cr-labeled Con A lymphoblasts as targets in the presence or the absence of Abs to Ly49 and/or class I molecules. Our results demonstrate that the activating receptor Ly49D delivers stimulatory signals for target cell lysis upon interacting with H2-Dd, Dr, and Dsp2, but not H2b or H2k class I Ags. On the other hand, the inhibitory receptor Ly49G2 delivers negative signals for target cell lysis upon interacting with Dd, Dr, and H2k, but not H2b or Dsp2, class I Ags. Furthermore, Ly49-mediated negative signaling dominates Ly49D-mediated positive signaling. Thus, lysis of class I MHC-bearing targets by NK cells is not merely the consequence of the absence of an Ly49-mediated negative signal, but also requires positive recognition of class I molecules by certain Ly49 receptors. Activation of NK cells by nonself class I molecules was not predicted by the missing self hypothesis.