bluuebunny/crossref_metadata_2025_split huggingface dataset:
Enterocyte TLR4 Mediates Phagocytosis and Translocation of Bacteria Across the Intestinal Barrier
Authors: Neal Matthew D. , Leaphart Cynthia , Levy Ryan , Prince Jose , Billiar Timothy R. , Watkins Simon , Li Jun , Cetin Selma , Ford Henri , Schreiber Alan , Hackam David J. 
Published: April 2014

Abstract
Translocation of bacteria across the intestinal barrier is important in the pathogenesis of systemic sepsis, although the mechanisms by which bacterial translocation occurs remain largely unknown. We hypothesized that bacterial translocation across the intact barrier occurs after internalization of the bacteria by enterocytes in a process resembling phagocytosis and that TLR4 is required for this process. We now show that FcγRIIa-transfected enterocytes can internalize IgG-opsonized erythrocytes into actin-rich cups, confirming that these enterocytes have the molecular machinery required for phagocytosis. We further show that enterocytes can internalize Escherichia coli into phagosomes, that the bacteria remain viable intracellularly, and that TLR4 is required for this process to occur. TLR4 signaling was found to be necessary and sufficient for phagocytosis by epithelial cells, because IEC-6 intestinal epithelial cells were able to internalize LPS-coated, but not uncoated, latex particles and because MD2/TLR4-transfected human endothelial kidney (HEK)-293 cells acquired the capacity to internalize E. coli, whereas nontransfected HEK-293 cells and HEK-293 cells transfected with dominant-negative TLR4 bearing a P712H mutation did not. LPS did not induce membrane ruffling or macropinocytosis in enterocytes, excluding their role in bacterial internalization. Strikingly, the internalization of Gram-negative bacteria into enterocytes in vivo and the translocation of bacteria across the intestinal epithelium to mesenteric lymph nodes were significantly greater in wild-type mice as compared with mice having mutations in TLR4. These data suggest a novel mechanism by which bacterial translocation occurs and suggest a critical role for TLR4 in the phagocytosis of bacteria by enterocytes in this process.
Mycobacterium bovis Bacillus Calmette-Guérin Induces TLR2-Mediated Formation of Lipid Bodies: Intracellular Domains for Eicosanoid Synthesis In Vivo
Authors: D’Avila Heloisa , Melo Rossana C. N. , Parreira Gleydes G. , Werneck-Barroso Eduardo , Castro-Faria-Neto Hugo C. , Bozza Patrícia T. 
Published: April 2014

Abstract
Differentiation of macrophages into foamy (lipid-laden) macrophages is a common pathological observation in tuberculous granulomas both in experimental settings as well as in clinical conditions; however, the mechanisms that regulate intracellular lipid accumulation in the course of mycobacterial infection and their significance to pathophysiology of tuberculosis are not well understood. In this study, we investigated the mechanisms of formation and function of lipid-laden macrophages in a murine model of tuberculosis. Mycobacterium bovis bacillus Calmette-Guérin (BCG), but not Mycobacterium smegmatis, induced a dose- and time-dependent increase in lipid body-inducible nonmembrane-bound cytoplasmic lipid domain size and numbers. Lipid body formation was drastically inhibited in TLR2-, but not in TLR4-deficient mice, indicating a role for TLR2 in BCG recognition and signaling to form lipid bodies. Increase in lipid bodies during infection correlated with increased generation of PGE2 and localization of cyclooxygenase-2 within lipid bodies. Moreover, we demonstrated by intracellular immunofluorescent localization of newly formed eicosanoid that lipid bodies were the predominant sites of PGE2 synthesis in activated macrophages. Our findings demonstrated that BCG-induced lipid body formation is TLR2 mediated and these structures function as signaling platforms in inflammatory mediator production, because compartmentalization of substrate and key enzymes within lipid bodies has impact on the capacity of activated leukocytes to generate increased amounts of eicosanoids during experimental infection by BCG.
Vascular Endothelial Growth Factor-Induced Signaling Pathways in Endothelial Cells That Mediate Overexpression of the Chemokine IFN-γ-Inducible Protein of 10 kDa In Vitro and In Vivo
Authors: Boulday Gwénola , Haskova Zdenka , Reinders Marlies E. J. , Pal Soumitro , Briscoe David M. 
Published: April 2014

Abstract
Vascular endothelial growth factor (VEGF), an angiogenesis factor, has recently been found to have potent proinflammatory properties in vivo. However, the mechanism by which it mediates inflammation is poorly understood. In this study, we have evaluated the function of VEGF on the induced expression and function of the T cell chemoattractant chemokine IFN-γ-inducible protein of 10 kDa (IP-10). In vitro, we find that VEGF augments the effect of IFN-γ on the induction of IP-10 mRNA and protein expression in endothelial cells. Moreover, we show that VEGF and IFN-γ regulate the activation of the IP-10 promoter, and that the kinases PI3K, phosphoinositide-dependent kinase 1, and Akt act as intermediary signaling molecules for cytokine-inducible IP-10 transcriptional activation in endothelial cells. To examine whether VEGF is functional for IP-10 expression in vivo, Chinese hamster ovary cells that were designed to secrete VEGF were injected s.c. into the skin of nude mice and were found to mediate a time-dependent increase in IP-10 mRNA. This response was reduced in animals treated systemically with the PI3K inhibitor wortmannin. When the Chinese hamster ovary cells expressing VEGF plasmid were injected s.c. into C57BL/6 wild-type or CXCR3−/− mice, they elicited an inflammatory reaction in wild-type but not in CXCR3−/− mice. Collectively, these findings indicate that VEGF-induced augmentation of IP-10 expression is a major mechanism underlying its proinflammatory function.
Renal Ischemia-Reperfusion Injury and Adenosine 2A Receptor-Mediated Tissue Protection: The Role of CD4+ T Cells and IFN-γ
Authors: Day Yuan-Ji , Huang Liping , Ye Hong , Li Li , Linden Joel , Okusa Mark D. 
Published: April 2014

Abstract
A2A adenosine receptor (A2AR)-expressing bone marrow (BM)-derived cells contribute to the renal protective effect of A2A agonists in renal ischemia-reperfusion injury (IRI). We performed IRI in mice lacking T and B cells to determine whether A2AR expressed in CD4+ cells mediate protection from IRI. Rag-1 knockout (KO) mice were protected in comparison to wild-type (WT) mice when subjected to IRI. ATL146e, a selective A2A agonist, did not confer additional protection. IFN-γ is an important early signal in IRI and is thought to contribute to reperfusion injury. Because IFN-γ is produced by kidney cells and T cells we performed IRI in BM chimeras in which the BM of WT mice was reconstituted with BM from IFN-γ KO mice (IFN-γ KO→WT chimera). We observed marked reduction in IRI in comparison to WT→WT chimeras providing additional indirect support for the role of T cells. To confirm the role of CD4+ A2AR in mediating protection from IRI, Rag-1 KO mice were subjected to ischemia-reperfusion. The protection observed in Rag-1 KO mice was reversed in Rag-1 KO mice that were adoptively transferred WT CD4+ cells (WT CD4+→Rag-1 KO) or A2A KO CD4+ cells (A2A KO CD4+→Rag-1 KO). ATL146e reduced injury in WT CD4+→Rag-1 KO mice but not in A2A KO CD4+→Rag-1 KO mice. Rag-1 KO mice reconstituted with CD4+ cells derived from IFN-γ KO mice (IFN-γ CD4+→Rag-1 KO) were protected from IRI; ATL146e conferred no additional protection. These studies demonstrate that CD4+ IFN-γ contributes to IRI and that A2A agonists mediate protection from IRI through action on CD4+ cells.
Exacerbation of Experimental Autoimmune Encephalomyelitis in P2X7R−/− Mice: Evidence for Loss of Apoptotic Activity in Lymphocytes
Authors: Chen Lanfen , Brosnan Celia F. 
Published: April 2014

Abstract
The purinergic receptor P2X7R is a nucleotide-gated ion channel that has been proposed to function as a major regulator of inflammation. In this study we examined the role of this receptor in regulating inflammation in the CNS by determining the effects of the loss of this receptor (P2X7R−/−) on experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis. We show here that P2X7R−/− mice developed more severe clinical and pathological expression of EAE than wild type (WT) controls and that spleen and lymph node cells from P2X7R−/− mice proliferated more vigorously to Ag in vitro. Bone marrow (BM) radiation chimeras revealed that enhanced susceptibility to EAE was detected in chimeric mice of WT host engrafted with P2X7R−/− BM cells, indicating that the genotype of the BM cells regulated disease susceptibility. Coculture of P2X7R−/− macrophages with WT lymphocytes and vice versa showed that enhanced proliferative activity resided within the P2X7R−/− lymphocyte population and correlated with reduced levels of IFN-γ and NO and apoptosis of lymphocytes. mRNA and protein for IFN-γ were also significantly reduced in the CNS of P2X7R−/− mice with EAE. FACS analysis of cells isolated from the CNS showed significantly fewer annexin V/propidium iodide-positive lymphocytes in the CNS of P2X7R−/− mice early in the disease, and TUNEL staining of inflamed CNS tissues supported this result. From these data we conclude that enhanced susceptibility of P2X7R−/− mice to EAE reflects a loss of apoptotic activity in lymphocytes, supporting an important role for this receptor in lymphocyte homeostasis.
The Human Specific CCR1 Antagonist CP-481,715 Inhibits Cell Infiltration and Inflammatory Responses in Human CCR1 Transgenic Mice
Authors: Gladue Ronald P. , Cole Susan H. , Roach Marsha L. , Tylaska Laurie A. , Nelson Robin T. , Shepard Richard M. , McNeish John D. , Ogborne Kevin T. , Neote Kuldeep S. 
Published: April 2014

Abstract
We previously described the in vitro characteristics of the potent and selective CCR1 antagonist, CP-481,715. In addition to being selective for CCR1 vs other chemokine receptors, CP-481,715 is also specific for human CCR1 (hCCR1), preventing its evaluation in classical animal models. To address this, we generated mice whereby murine CCR1 was replaced by hCCR1 (knockin) and used these animals to assess the anti-inflammatory properties of CP-481,715. Cells isolated from hCCR1 knockin mice were shown to express hCCR1 and migrate in response to both murine CCR1 and hCCR1 ligands. Furthermore, this migration is inhibited by CP-481,715 at dose levels comparable to those obtained with human cells. In animal models of cell infiltration, CP-481,715 inhibited CCL3-induced neutrophil infiltration into skin or into an air pouch with an ED50 of 0.2 mg/kg. CP-481,715 did not inhibit cell infiltration in wild-type animals expressing murine CCR1. In a more generalized model of inflammation, delayed-type hypersensitivity, CP-481,715 significantly inhibited footpad swelling and decreased the amount of IFN-γ and IL-2 produced by isolated spleen cells from sensitized animals. It did not, however, induce tolerance to a subsequent challenge. These studies illustrate the utility of hCCR1 knockin animals to assess the activity of human specific CCR1 antagonists; demonstrate the ability of the CCR1 antagonist CP-481,715 to inhibit cell infiltration, inflammation, and Th1 cytokine responses in these animals; and suggest that CP-481,715 may be useful to modulate inflammatory responses in human disease.
The B12 Anti-Tryptase Monoclonal Antibody Disrupts the Tetrameric Structure of Heparin-Stabilized β-Tryptase to Form Monomers That Are Inactive at Neutral pH and Active at Acidic pH
Authors: Fukuoka Yoshihiro , Schwartz Lawrence B. 
Published: April 2014

AbstractThe novel tetrameric structure of human β-tryptase faces each active site into the central pore, thereby restricting access of most biologic protease inhibitors. The mechanism by which the anti-tryptase mAb B12 inhibits human β-tryptase peptidase and proteolytic activities at neutral pH, but augments proteolytic activity at acidic pH, was examined. At neutral pH, B12-β-tryptase complexes are inactive. At acidic pH, B12 (intact and Fab) minimally affects peptidase activity when added to β-tryptase tetramers, but does induce susceptibility to inhibition by soybean trypsin inhibitor and antithrombin III. Surprisingly, B12 Fab-β-tryptase complexes formed at both neutral and acidic pH exhibit the apparent molecular mass of a complex with 1 β-tryptase monomer and 1 Fab by gel filtration. B12 does not compete with heparin for binding to tryptase at either neutral or acidic pH. Thus, B12 directly disrupts β-tryptase tetramers to monomers that are inactive at neutral pH, whereas at acidic pH, are active and more accessible to protein inhibitors and substrates.
c-Jun Kinase Is a Critical Signaling Molecule in a Neonatal Model of Group B Streptococcal Sepsis
Authors: Kenzel Sybille , Mancuso Guiseppe , Malley Richard , Teti Guiseppe , Golenbock Douglas T. , Henneke Philipp 
Published: April 2014

Abstract
Group B streptococcus (GBS) is the major cause of sepsis in newborn infants. In vitro, inactivated GBS stimulates macrophages to produce inflammatory proteins via the TLR adapter protein MyD88. Furthermore, inflammatory cytokine release in response to GBS greatly exceeds that following stimulation with pneumococci. In this study, we attempted to unravel signaling events that are involved in GBS-, but not Streptococcus pneumoniae-stimulated phagocytes to identify molecular targets for adjunctive sepsis therapy. We found that inactivated GBS and S. pneumoniae differed in the activation of the MAPK JNK, but not IκB kinase. Furthermore, JNK was essential for the transcriptional activation of inflammatory cytokine genes in response to GBS. Inhibition of JNK by the anthrapyrazolone SP600125 abrogated GBS-induced cytokine formation via an AP-1- and NF-κB-dependent mechanism without impairing antibacterial properties such as phagocytosis of GBS and the formation of intracellular oxidative species. In contrast, inhibition of the MAPK p38 impaired both antibacterial processes. In a neonatal mouse model of GBS sepsis SP600125 inhibited the inflammatory response and improved survival. In conclusion, JNK plays a major role in the inflammatory, but not in the direct antibacterial response to inactivated GBS, and may thus serve as a rational target for an adjunctive GBS sepsis therapy.
Pulmonary Lipopolysaccharide (LPS)-Binding Protein Inhibits the LPS-Induced Lung Inflammation In Vivo
Authors: Knapp Sylvia , Florquin Sandrine , Golenbock Douglas T. , van der Poll Tom 
Published: April 2014

Abstract
LPS-binding protein (LBP) facilitates the interaction of the Gram-negative cell wall component LPS with CD14, thereby enhancing the immune response to LPS. Although lung epithelial cells have been reported to produce LBP in vitro, knowledge of the in vivo role of pulmonary LBP is limited. Therefore, in the present study we sought to determine the function of pulmonary LBP in lung inflammation induced by intranasal administration of LPS in vivo. Using LBP-deficient (LBP−/−) and normal wild-type mice, we show that the contribution of LBP to pulmonary LPS responsiveness depended entirely on the LPS dose. Although the inflammatory response to low dose (1 ng) LPS was attenuated in LBP−/− mice, neutrophil influx and cytokine/chemokine concentrations in the bronchoalveolar compartment were enhanced in LBP−/− mice treated with higher (&gt;10 ng) LPS doses. This finding was specific for LBP, because the exogenous administration of LBP to LBP−/− mice reversed this phenotype and reduced the local inflammatory response to higher LPS doses. Our results indicate that pulmonary LBP acts as an important modulator of the LPS response in the respiratory tract in vivo. This newly identified function of pulmonary LBP might prove beneficial by enabling a protective immune response to low LPS doses while preventing an overwhelming, potentially harmful immune response to higher doses of LPS.
Fibulin-4 Is a Target of Autoimmunity Predominantly in Patients with Osteoarthritis
Authors: Xiang Yang , Sekine Taichi , Nakamura Hiroshi , Imajoh-Ohmi Shinobu , Fukuda Hiroyuki , Yudoh Kazuo , Masuko-Hongo Kayo , Nishioka Kusuki , Kato Tomohiro 
Published: April 2014

Abstract
Autoimmunity to chondrocyte-producing proteins has been reported in patients with osteoarthritis (OA) as well as in those with rheumatoid arthritis (RA). To answer whether or not OA-specific autoimmunity exist, we performed screening of chondrocyte-producing autoantigens by two-dimensional electrophoresis and Western blotting with each of 20 OA and 20 RA serum samples. We identified an apparently OA-specific autoantigen spot with a molecular mass of 52 kDa and a Isoelectric point of 4.1 as fibulin-4 by mass fingerprinting. By preparing recombinant proteins of fibulin-4, we determined prevalence of the autoantibodies to fibulin-4 in 92 patients with OA, 67 patients with RA, 40 patients with systemic lupus erythematosus, and 43 patients with systemic scleroderma. As a result, the IgG type anti-fibulin-4 autoantibodies were detected in 23.9% of sera from patients with OA, in 8.9% of sera from patients with RA, in 2.5% of sera from patients with systemic lupus erythematosus, and in 9.3% of sera from patients with systemic scleroderma. Furthermore, we immunized DBA/1J, ICR, BALB/c, and C57BL/6 mice with the recombinant fibulin-4 proteins to investigate arthritogenecity of fibulin-4. As a result, mild synovitis was detected in all of the four strains. In addition, we demonstrated expression of fibulin-4 in chondrocytes at both mRNA and protein levels in vivo and in vitro by RT-PCR, Western blotting, and immunohistochemistry. Taken together, fibulin-4, expressed in chondrocytes and recognized as an autoantigen mainly in OA rather than in RA, may play pathogenic roles in OA.
Inhibition of the Transcription Factor Foxp3 Converts Desmoglein 3-Specific Type 1 Regulatory T Cells into Th2-Like Cells
Authors: Veldman Christian , Pahl Andreas , Beissert Stefan , Hansen Wiebke , Buer Jan , Dieckmann Detlef , Schuler Gerold , Hertl Michael 
Published: April 2014

Abstract
Pemphigus vulgaris (PV) is a severe autoimmune bullous skin disorder and is associated with autoantibodies against desmoglein (Dsg)3 that are regulated by Th2 cells. Recently, Dsg3-specific type 1 regulatory T cells (Tr1) were identified that are presumably critical for the maintenance of tolerance against Dsg3 because there is a much lower Dsg3-specific Tr1:Th2 ratio in the PV patients than in healthy individuals. The aim of this study was to down-regulate the transcription factor Foxp3 in Dsg3-specific Tr1 using antisense oligonucleotides because Foxp3 is constitutively expressed by the Dsg3-specific Tr1. Antisense-treated Dsg3-specific Tr1 clones lost expression of Foxp3, glucocorticoid-induced TNFR family-related receptor, and CTLA-4, and started to secrete IL-2, whereas the secretion of IL-5, TGF-β, and IL-10 remained unchanged. Moreover, antisense treatment induced a proliferative response to Dsg3 of the formerly anergic Tr1 and abrogated their suppressor activity on Dsg3-specific Th2 cell clones. Thus, inhibition of Foxp3 mRNA expression in the Tr1 induced a Th2-like phenotype. In conclusion, Foxp3 expression is inherent to Tr1 function, and modulation of Foxp3 expression in autoaggressive Th2 cells may provide a novel therapeutic approach aimed at restoring tolerance against Dsg3 in PV.
Visualization of p53264–272/HLA-A*0201 Complexes Naturally Presented on Tumor Cell Surface by a Multimeric Soluble Single-Chain T Cell Receptor
Authors: Zhu Xiaoyun , Belmont Heather J. , Price-Schiavi Shari , Liu Bai , Lee Hyung-il , Fernandez Marilyn , Wong Richard L. , Builes Janette , Rhode Peter R. , Wong Hing C. 
Published: April 2014

AbstractIntracellular Ags are processed into small peptides that are presented on cell surfaces in the context of HLA class I molecules. These peptides are recognized by TCRs displayed by CD8+ T lymphocytes (T cells). To date, direct identification and quantitation of these peptides has relied primarily on mass spectrometry analysis, which is expensive and requires large quantities of diseased tissues to obtain useful results. Here we demonstrate that multimerization of a soluble single-chain TCR (scTCR), recognizing a peptide from p53 presented in the context of HLA-A2.1, could be used to directly visualize and quantitate peptide/MHC complexes on unmanipulated human tumor cells. Tumor cells displaying as few as 500 peptide/MHC complexes were readily detectable by flow cytometry. The scTCR/multimers exhibited exquisite recognition capability and could distinguish peptides differing in as little as a single amino acid. We also demonstrate that scTCR/multimers could specifically stain human tumors generated in mice, as well as tumors obtained from patient biopsies. Thus, scTCR/multimers represent a novel class of immunostaining reagents that could be used to validate, quantitate, or monitor epitope presentation by cancer cells.
The Lupus-Susceptibility Locus, Sle3, Mediates Enhanced Resistance to Bacterial Infections
Authors: Mehrad Borna , Park Stacy J. , Akangire Gangaram , Standiford Theodore J. , Wu Tianfu , Zhu Jiankun , Mohan Chandra 
Published: April 2014

Abstract
The genetic predisposition to many autoimmune diseases is inherited as a polygenic trait. It is conceivable that some of the causative alleles in these diseases became prevalent in the population by conferring a survival benefit against environmental assaults, such as infections. We used mice cogenic for genetic loci predisposing to systemic lupus erythomatosus to test the hypothesis that some of these genetic loci protect the host from bacterial infections. Mice with the Sle3 lupus-susceptibility locus on a wild-type background were found to have enhanced antibacterial responses in the context of pneumonia and intra-abdominal sepsis than wild-type animals. This was associated with markedly augmented accumulation of neutrophils in infected tissues, and was bone marrow transferable and dependent on the presence of neutrophils, but not lymphocytes. There was no difference in in vitro leukocyte killing of bacteria nor influx of phagocytes between lupus-susceptible and wild-type animals, but neutrophils from lupus-susceptible mice displayed markedly reduced rate of apoptosis, associated with altered expression of Bcl-2 family proteins, contributing to their greater accumulation. Importantly, deliberate inhibition of apoptosis in wild-type animals significantly boosted the accumulation of neutrophils at the site of infection and resulted in an enhanced antimicrobial response. These observations support the concept that some of the genetic loci that mediate autoimmunity may also confer augmented antimicrobial innate immunity.