bluuebunny/crossref_metadata_2025_split huggingface dataset:
IL-12 Antagonism Enhances Apoptotic Death of T Cells Within Hepatic Allografts from Flt3 Ligand-Treated Donors and Promotes Graft Acceptance
Authors: Li Wei , Lu Lina , Wang Zhiliang , Wang Lianfu , Fung John J. , Thomson Angus W. , Qian Shiguang 
Published: April 2014

Abstract
Mouse livers are accepted across MHC barriers and induce donor-specific tolerance without immunosuppressive therapy. By contrast, livers from donors treated with Flt3 ligand, which dramatically increases hepatic interstitial dendritic cells, are rejected acutely (median survival time 5 days). This switch from tolerance to rejection is associated with a marked reduction in apoptotic activity of graft-infiltrating cells. We hypothesized that IL-12 production by enhanced numbers of donor APC might inhibit apoptosis, promote expansion of Th1 cells, and play a key role in liver rejection. Therefore, C3H (H2k) recipients of liver grafts from Flt3 ligand-treated B10 donors were given neutralizing anti-IL-12 mAb (200 or 500 μg) on days 0 and 2 after transplant. Graft survival was markedly prolonged at the higher mAb dose, with 50% of grafts surviving &gt;100 days. This effect was associated with reductions in IFN-γ gene transcripts within the graft-infiltrating cell population and with reductions in circulating IFN-γ and IL-10 levels, donor-specific CTL and NK cell activities, and circulating alloantibody levels. At the same time, there were marked increases in apoptotic (TUNEL+) CD4+ and especially CD8+ cells, both within the grafts and in spleens of anti-IL-12 mAb-treated mice. In vitro, exogenous IL-12 inhibited apoptotic death induced in naive allogeneic T cells by liver nonparenchymal cells. These findings suggest that suppression of rejection by IL-12 antagonism, linked to restoration of apoptotic activity within the peripheral alloreactive T cell population, is important for liver allograft survival and tolerance induction.
Epitope Mapping of Antibodies to the C-Terminal Region of the Integrin β2 Subunit Reveals Regions that Become Exposed Upon Receptor Activation
Authors: Lu Chafen , Ferzly Mazen , Takagi Junichi , Springer Timothy A. 
Published: April 2014

Abstract
The cysteine-rich repeats in the stalk region of integrin β subunits appear to convey signals impinging on the cytoplasmic domains to the ligand-binding headpiece of integrins. We have examined the functional properties of mAbs to the stalk region and mapped their epitopes, providing a structure-function map. Among a panel of 14 mAbs to the β2 subunit, one, KIM127, preferentially bound to αLβ2 that was activated by mutations in the cytoplasmic domains, and by Mn2+. KIM127 also bound preferentially to the free β2 subunit compared with resting αLβ2. Activating β2 mutations also greatly enhanced binding of KIM127 to integrins αMβ2 and αXβ2. Thus, the KIM127 epitope is shielded by the α subunit, and becomes reexposed upon receptor activation. Three other mAbs, CBR LFA-1/2, MEM48, and KIM185, activated αLβ2 and bound equally well to resting and activated αLβ2, differentially recognized resting αMβ2 and αXβ2, and bound fully to activated αMβ2 and αXβ2. The KIM127 epitope localizes within cysteine-rich repeat 2, to residues 504, 506, and 508. By contrast, the two activating mAbs CBR LFA-1/2 and MEM48 bind to overlapping epitopes involving residues 534, 536, 541, 543, and 546 in cysteine-rich repeat 3, and the activating mAb KIM185 maps near the end of cysteine-rich repeat 4. The nonactivating mAbs, 6.7 and CBR LFA-1/7, map more N-terminal, to subregions 344–432 and 432–487, respectively. We thus define five different β2 stalk subregions, mAb binding to which correlates with effect on activation, and define regions in an interface that becomes exposed upon integrin activation.
Mice Triallelic for the Ig Heavy Chain Locus: Implications for VHDJH Recombination
Authors: Barreto Vasco , Meo Tommaso , Cumano Ana 
Published: April 2014

Abstract
VHDJH recombination has been extensively studied in mice carrying an Ig heavy chain rearranged transgene. In most models, inhibition of endogenous Ig rearrangement occurs, consistently with the feedback model of IgH recombination. Nonetheless, an incomplete IgH allelic exclusion is a recurrent observation in these animals. Furthermore, transgene expression in ontogeny is likely to start before somatic recombination, thus limiting the use of Ig-transgenic mice to access the dynamics of VHDJH recombination. As an alternative approach, we challenged the regulation of somatic recombination with the introduction of an extra IgH locus in germline configuration. This was achieved by reconstitution of RAG2−/− mice with fetal liver cells trisomic for chromosome 12 (Ts12). We found that all three alleles can recombine and that the ratio of Ig allotype-expressing B cells follows the allotypic ratio in trisomic cells. Although these cells are able to rearrange the three alleles, the levels of Ig phenotypic allelic exclusion are not altered when compared with euploid cells. Likewise, we find that most VDJ rearrangements of the silenced allele are unable to encode a functional μ-chain, indicating that the majority of these cells are also genetically excluded. These results provide additional support for the feedback model of allelic exclusion.
Isoforms of Jun Kinase Are Differentially Expressed and Activated in Human Monocyte/Macrophage (THP-1) Cells
Authors: Dreskin Stephen C. , Thomas Gregory W. , Dale Sara N. , Heasley Lynn E. 
Published: April 2014

Abstract
Ten isoforms of c-jun N-terminal kinase (JNK) have been described that arise by differential mRNA splicing of three genes. In that the relative expression and function of these different JNK proteins in human monocytic cells is not known, we have examined the JNK isoforms in THP-1 monocyte/macrophage cells. Differentiation of THP-1 cells by exposure to 10−8 M PMA for 42–48 h enhances cellular responses to LPS, including enhanced activation of total JNK activity and increased phosphorylation of p54 JNK as well as p46 JNK. Examination of JNK proteins on Western blots reveals a predominance of p46 JNK1 and p54 JNK2 proteins. Clearing of lysates by immunoprecipitation of JNK1(99% effective) removes 46% of the JNK enzymatic activity (p &lt; 0.01), whereas clearing of JNK1 plus JNK2 (70% effective) depletes the sample of 72% of the JNK activity (p &lt; 0.01). Further analysis, undertaken with real-time RT-PCR, revealed that 98% of the JNK messages code for three isoforms: JNK1β1, JNK2α1, and JNK2α2. The p54 JNK that is phosphorylated in LPS-stimulated, PMA-differentiated THP-1 cells is most likely JNK2α2 because 97% of the p54 JNK-encoding messages code for JNK2α2. By analogous reasoning, the p46 JNKs that are not heavily phosphorylated, but account for approximately half of the N-terminal c-jun kinase enzymatic activity, are most likely either JNK1β1 or JNK2α1 because they account for 98% of the messages that can code for 46kDa JNKs.
ZAP-70 and SLP-76 Regulate Protein Kinase C-θ and NF-κB Activation in Response to Engagement of CD3 and CD28
Authors: Herndon Thomas M. , Shan Xiaochuan C. , Tsokos George C. , Wange Ronald L. 
Published: April 2014

Abstract
The transcription factor NF-κB is a critical regulator of T cell function that becomes strongly activated in response to coengagement of TCR and CD28. Although events immediately proximal to NF-κB activation are well understood, uncertainty remains over which upstream signaling pathways engaged by TCR and CD28 lead to NF-κB activation. By using Jurkat T cell lines that are deficient or replete for either the protein tyrosine kinase ZAP-70 or the cytosolic adapter molecule SLP-76, the role of these proteins in modulating NF-κB activation was examined. NF-κB was not activated in response to coengagement of TCR and CD28 in either the ZAP-70- or SLP-76-negative cells, whereas stimuli that bypass these receptors (PMA plus A23187, or TNF-α) activated NF-κB normally. Protein kinase C (PKC) θ activation, which is required for NF-κB activation, also was defective in these cells. Reexpression of ZAP-70 restored PKCθ and NF-κB activation in response to TCR and CD28 coengagement. p95vav (Vav)-1 tyrosine phosphorylation was largely unperturbed in the ZAP-70-negative cells; however, receptor-stimulated SLP-76/Vav-1 coassociation was greatly reduced. Wild-type SLP-76 fully restored PKCθ and NF-κB activation in the SLP-76-negative cells, whereas 3YF-SLP-76, which lacks the sites of tyrosine phosphorylation required for Vav-1 binding, only partially rescued signaling. These data illustrate the importance of the ZAP-70/SLP-76 signaling pathway in CD3/CD28-stimulated activation of PKC θ and NF-κB, and suggest that Vav-1 association with SLP-76 may be important in this pathway.
Protein Kinase C-θ Participates in the Activation of Cyclic AMP-Responsive Element-Binding Protein and Its Subsequent Binding to the −180 Site of the IL-2 Promoter in Normal Human T Lymphocytes
Authors: Solomou Elena E. , Juang Yuang-Taung , Tsokos George C. 
Published: April 2014

Abstract
IL-2 gene expression is regulated by the cooperative binding of discrete transcription factors to the IL-2 promoter/enhancer and is predominantly controlled at the transcriptional level. In this study, we show that in normal T cells, the −180 site (−164/−189) of the IL-2 promoter/enhancer is a p-cAMP-responsive element-binding protein (p-CREB) binding site. Following activation of the T cells through various membrane-initiated and membrane-independent pathways, protein kinase C (PKC)-θ phosphorylates CREB, which subsequently binds to the −180 site and associates with the transcriptional coactivator p300. Rottlerin, a specific PKC-θ inhibitor, diminished p-CREB protein levels when normal T cells were treated with it. Rottlerin also prevented the formation of p-CREB/p300 complexes and the DNA-CREB protein binding. Cotransfection of fresh normal T cells with luciferase reporter construct driven by two tandem −180 sites and a PKC-θ construct caused a significant increase in the transcription of the reporter gene, indicating that this site is functional and regulated by PKC-θ. Cotransfection of T cells with a luciferase construct driven by the −575/+57 region of the IL-2 promoter/enhancer and a PKC-θ construct caused a similar increase in the reporter gene transcription, which was significantly limited when two bases within the −180 site were mutated. These findings show that CREB plays a major role in the transcriptional regulation of IL-2 and that a major pathway for the activation of CREB and its subsequent binding to the IL-2 promoter/enhancer in normal T cells is mediated by PKC-θ.
Cloning, Expression, and Function of BLAME, a Novel Member of the CD2 Family
Authors: Kingsbury Gillian A. , Feeney Lee Ann , Nong Yuhua , Calandra Susan  A , Murphy Curran J. , Corcoran Justin M. , Wang Yanjun , Prabhu Das Mercy R. , Busfield Samantha J. , Fraser Christopher C. , Villeval Jean Luc 
Published: April 2014

Abstract
The CD2 family is a growing family of Ig domain-containing cell surface proteins involved in lymphocyte activation. Here we describe the cloning and expression analysis of a novel member of this family, B lymphocyte activator macrophage expressed (BLAME). BLAME shares the structural features of the CD2 family containing an IgV and IgC2 domain and clusters with the other family members on chromosome 1q21. Quantitative PCR and Northern blot analysis show BLAME to be expressed in lymphoid tissue and, more specifically, in some populations of professional APCs, activated monocytes, and DCs. Retroviral forced expression of BLAME in hematopoietic cells of transplanted mice showed an increase in B1 cells in the peripheral blood, spleen, lymph nodes, and, most strikingly, in the peritoneal cavity. These cells do not express CD5 and are CD23lowMac1low, characteristics of the B1b subset. BLAME may therefore play a role in B lineage commitment and/or modulation of signal through the B cell receptor.
Uncoordinated HLA-D Gene Expression in a RFXANK-Defective Patient with MHC Class II Deficiency
Authors: Lennon-Duménil Ana-Maria , Barbouche Mohamed-Ridha , Vedrenne Jocelyn , Prod’Homme Thomas , Béjaoui Mohamed , Ghariani Salma , Charron Dominique , Fellous Marc , Dellagi Koussay , Alcaïde-Loridan Catherine 
Published: April 2014

Abstract
We describe the analysis of a patient, JER, presenting classical immunological features of MHC class II deficiency. Unexpectedly, some HLA transcripts (HLA-DRA, HLA-DQA, and HLA-DMA) were found to be expressed in the JER cell line at nearly wild-type levels, while HLA-DPA and the HLA-D β-chain transcripts were not detected. Gene reporter experiments confirmed the differential transcriptional activities driven by the HLA-D promoters in the JER cells. A defect in RFXANK was first suggested by genetic complementation analyses, then assessed with the demonstration of a homozygous mutation affecting a splice donor site downstream exon 4 of RFXANK. Because the severe deletion of the resulting protein cannot account for the expression of certain HLA-D genes, minor alternative transcripts of the RFXANK gene were analyzed. We thereby showed the existence of a transcript lacking exon 4, encoding a 28-aa-deleted protein that retains a transcriptional activity. Altogether, we characterize a new type of mutation in the RFXANK gene in a MHC class II-defective patient leading to an uncoordinated expression of the HLA-D genes, and propose that this phenotype is ensured by severely limited amounts of an active, although truncated RFXANK protein.
Endotoxin-Induced Maturation of MyD88-Deficient Dendritic Cells
Authors: Kaisho Tsuneyasu , Takeuchi Osamu , Kawai Taro , Hoshino Katsuaki , Akira Shizuo 
Published: April 2014

Abstract
LPS, a major component of the cell wall of Gram-negative bacteria, can induce a variety of biological responses including cytokine production from macrophages, B cell proliferation, and endotoxin shock. All of them were completely abolished in MyD88-deficient mice, indicating the essential role of MyD88 in LPS signaling. However, MyD88-deficient cells still show activation of NF-κB and mitogen-activated protein kinase cascades, although the biological significance of this activation is not clear. In this study, we have examined the effects of LPS on dendritic cells (DCs) from wild-type and several mutant mice. LPS-induced cytokine production from DCs was dependent on MyD88. However, LPS could induce functional maturation of MyD88-deficient DCs, including up-regulation of costimulatory molecules and enhancement of APC activity. MyD88-deficient DCs could not maturate in response to bacterial DNA, the ligand for Toll-like receptor (TLR)9, indicating that MyD88 is differentially required for TLR family signaling. MyD88-dependent and -independent pathways originate at the intracytoplasmic region of TLR4, because both cytokine induction and functional maturation were abolished in DCs from C3H/HeJ mice carrying the point mutation in the region. Finally, in vivo analysis revealed that MyD88-, but not TLR4-, deficient splenic CD11c+ DCs could up-regulate their costimulatory molecule expression in response to LPS. Collectively, the present study provides the first evidence that the MyD88-independent pathway downstrem of TLR4 can lead to functional DC maturation, which is critical for a link between innate and adaptive immunity.
Human Cytomegalovirus pp65- and Immediate Early 1 Antigen-Specific HLA Class I-Restricted Cytotoxic T Cell Responses Induced by Cross-Presentation of Viral Antigens
Authors: Tabi Zsuzsanna , Moutaftsi Magdalena , Borysiewicz Leszek K. 
Published: April 2014

Abstract
Dendritic cells (DCs) play a pivotal role in the development of anti-viral CD8+ CTL responses. This is straightforward if they are directly infected with virus, but is less clear in response to viruses that cannot productively infect DCs. Human CMV (HCMV) shows strain-specific cell tropism: fibroblast (Fb)-adapted laboratory strains (AD169) and recent clinical isolates do not infect DCs, whereas endothelial cell-adapted strains (TB40/E) result in productive lytic DC infection. However, we show here that uninfected DCs induce CD8+ T cell cytotoxicity and IFN-γ production against HCMV pp65 and immediate early 1 Ags following in vitro coculture with HCMV-AD169-infected Fbs, regardless of the HLA type of these Fbs. CD8+ T cell stimulation was inhibited by pretreatment of DCs with cytochalasin B or brefeldin A, indicating a phagosome/endosome to cytosol pathway. HCMV-infected Fbs were not apoptotic as measured by annexin V binding, and induction of apoptosis of infected Fbs in vitro did not augment CTL induction by DCs, suggesting a mechanism other than apoptosis in the initiation of cross-presentation. Furthermore, HCMV-infected Fbs provided a maturation signal for immature DCs during coculture, as evidenced by increased CD83 and HLA class II expression. Cross-presentation of HCMV Ags by host DCs enables these professional APCs to bypass some of the evasion mechanisms HCMV has developed to avoid T cell recognition. It may also serve to explain the presence of immediate early 1 Ag-specific CTLs in the face of pp65-induced inhibition of Ag presentation at the level of the infected cell.
Neonatal T Cells in an Adult Lung Environment Are Competent to Resolve Pneumocystis carinii Pneumonia
Authors: Qureshi Mahboob H. , Garvy Beth A. 
Published: April 2014

Abstract
Initiation of the pulmonary inflammatory response to Pneumocystis carinii is delayed by 3 wk in mice infected as neonates compared with adults. There was no difference in the proliferative response of draining lymph node T cells from mice infected as neonates compared with adults when stimulated in vitro with either Con A or anti-CD3 mAb. However, TNF-α and IFN-γ mRNA expression in the lungs of P. carinii-infected neonates was significantly lower than in adults indicating a lack of appropriate activation signaling in the local environment. This may have been due to active suppression because TGF-β mRNA expression was significantly elevated in neonatal lungs compared with adults. To determine whether T cells from 10-day-old mice would effect resolution of P. carinii if harbored in an adult lung environment, cells were adoptively transferred to SCID mice with established P. carinii infections. There was no difference in the kinetics of T cell migration into the lungs or of clearance of P. carinii organisms when SCID mice were reconstituted with splenocytes from young mice as compared with adult mice. Furthermore, splenocytes from young mice stimulated both TNF-α and IFN-γ mRNA expression to levels that were similar to that in the lungs of SCID mice reconstituted with adult cells. These data indicate that neonatal lymphocytes are competent to resolve P. carinii infection when harbored in an adult lung environment, suggesting that the neonatal lung environment, and not the T cells, is ineffective at responding to P. carinii infection.
A Heritable Defect in IL-12 Signaling in B10.Q/J Mice. I. In Vitro Analysis
Authors: Ortmann Robert , Smeltz Ronald , Yap George , Sher Alan , Shevach Ethan M. 
Published: April 2014

Abstract
B10.Q mice are normally susceptible to the induction of collagen-induced arthritis. We noted that one subline of B10.Q mice, B10.Q/J, was completely resistant to disease induction when immunized with collagen in CFA. B10.Q/J mice have a global defect in the generation of Th1 responses, and Ag-specific T cells derived from this strain failed to produce IFN-γ. Because T cells from these mice could produce normal amounts of IFN-γ when activated by IL-12/IL-18-independent stimuli, the defect appeared to be a failure to respond to IL-12. This defect extended to NK cells, which also failed to produce IFN-γ when stimulated by IL-12. The capacity of NK cells, but not activated T cells, to produce IFN-γ in response to IL-12 could be partially restored by IL-18. The expression of the IL-12R β1- and β2-chains on T cells and NK cells from B10.Q/J mice was normal. However, activated T cells from B10.Q/J mice did not signal normally through the IL-12R and manifested a defect in their capacity to phosphorylate Stat4. This defect was partial in that it could be overcome by increasing both the concentration of IL-12 and the incubation times in the Stat4 phosphorylation assays. Because Stat4 function is apparently intact in B10.Q/J mice, the defect in IL-12 signaling can be localized between the IL-12R complex and Stat4. This subtle abnormality in IL-12 responsiveness results in a profound defect in the generation of Th1 cells and the development of autoimmune disease.
A Heritable Defect in IL-12 Signaling in B10.Q/J Mice. II. Effect on Acute Resistance to Toxoplasma gondii and Rescue by IL-18 Treatment
Authors: Yap George S. , Ortmann Robert , Shevach Ethan , Sher Alan 
Published: April 2014

Abstract
This study documents a defect in IL-12-dependent IFN-γ responses in a substrain (B10.Q-H2-q/SgJ) of B10.Q mice that manifests as an acute susceptibility to infection by the intracellular protozoan pathogen, Toxoplasma gondii. Despite robust systemic production of IL-12, infected B10.Q/J animals fail to mount an early IFN-γ response after parasite inoculation. Genetic experiments revealed that the host resistance and IFN-γ production defects are determined by a single autosomal recessive locus distinct from the Stat4 gene. Nonetheless, a delayed IL-12-mediated IFN-γ response emerges in later stages of acute infection but is unable to prevent host mortality. IL-18 administration restores, in an IL-12-dependent manner, the early IFN-γ response and host resistance of B10.Q/J animals. These in vivo studies indicate that the partially impaired IL-12 responsiveness in B10.Q/J mice can result in defective host resistance and demonstrate a therapeutic function for IL-18 in reversing a genetically based immunodeficiency in IL-12-dependent IFN-γ production.
A Role for CD44 in the Production of IFN-γ and Immunopathology During Infection with Toxoplasma gondii
Authors: Blass Sarah L. , Puré Ellen , Hunter Christopher A. 
Published: April 2014

Abstract
The interaction of activated CD44 with its ligand, low m.w. hyaluronan, is involved in inflammation, but no role has been identified for this interaction in the regulation of an immune response to infection. In these studies, infection of C57BL/6 mice with Toxoplasma gondii resulted in increased expression of CD44 on T cells, B cells, NK cells, and macrophages, and a small percentage of CD4+ T cells express an activated form of CD44. Administration of anti-CD44 to infected mice prevented the development of a CD4+ T cell-dependent, infection-induced inflammatory response in the small intestine characterized by the overproduction of IFN-γ. The protective effect of anti-CD44 treatment was associated with reduced production of IFN-γ, but not IL-12, in vivo and in vitro. Furthermore, the addition of low m.w. hyaluronan to cultures of splenocytes or purified CD4+ T cells from infected mice resulted in the production of high levels of IFN-γ, which was dependent on IL-12 and TCR stimulation. Together, these results identify a novel role for CD44 in the regulation of IFN-γ production by CD4+ T cells during infection and demonstrate a role for CD44 in the regulation of infection-induced immune pathology.