bluuebunny/crossref_metadata_2025 huggingface dataset:
Simian Immunodeficiency Virus Evades a Dominant Epitope-Specific Cytotoxic T Lymphocyte Response Through a Mutation Resulting in the Accelerated Dissociation of Viral Peptide and MHC Class I
Authors: Chen Zheng W. , Craiu Abie , Shen Ling , Kuroda Marcelo J. , Iroku Ugonna C. , Watkins David I. , Voss Gerald , Letvin Norman L.
Published: April 2014

The ability of an AIDS virus to escape from immune containment by selective mutation away from recognition by CTL was explored in simian immunodeficiency virus of macaques (SIVmac)-infected rhesus monkeys. CTL recognition of a previously defined common viral mutation in an immunodominant SIVmac Gag epitope was evaluated. CTL were assessed for their ability to recognize a SIVmac Gag protein with a single residue 2 (T → A) replacement in the minimal epitope peptide bound by the MHC class I molecule Mamu-A*01. SIVmac Gag-specific CTL lysed Mamu-A*01+ target cells infected with recombinant vaccinia virus expressing the wild-type but not the mutant Gag protein. In addition, CTL recognized the mutant epitope peptide less efficiently than the wild-type virus peptide. In studies to determine the mechanism by which the mutant virus evaded CTL recognition, this peptide was shown to bind Mamu-A*01 in a manner that was indistinguishable from the wild-type peptide. However, experiments in which an increasing duration of delay was introduced between peptide sensitization of target cells and the assessment of these cells as targets in killing assays suggest that the mutant peptide with a T → A replacement had a higher off-rate from Mamu-A*01 than the wild-type peptide did. Therefore, these findings suggest that AIDS viruses can evade virus-specific CTL responses through the accelerated dissociation of mutant peptide from MHC class I.
Combined Effects of IL-12 and IL-18 on the Induction of Collagen-Induced Arthritis
Authors: Leung Bernard P. , McInnes Iain B. , Esfandiari Ehsan , Wei Xiao-Qing , Liew Foo Y.
Published: April 2014

IL-18 expression has recently been detected in rheumatoid arthritis (RA) synovial membrane. We investigated the mechanisms by which IL-18-induced collagen-induced arthritis in DBA/1 mice primed intradermally with type II bovine collagen in IFA and boosted i.p. 21 days later with CII in saline. Mice were injected i.p. with rIL-12, rIL-18, or both (100 ng) during days −1 to 4 and again on days 20–24. Control mice received PBS. Mice treated with IL-12 or IL-18 alone developed significantly higher incidence and more severe disease compared with controls. These were elevated further by combination treatment with IL-12 and IL-18. The cytokine treatments led to markedly enhanced synovial hyperplasia, cellular infiltration, and cartilage erosion compared with controls. Cytokine-treated mice produced significantly more IFN-γ, TNF-α, and IL-6 than the controls. Interestingly, IL-18-treated mice produced more TNF-α and IL-6, but less IFN-γ, compared with mice treated with IL-12. Furthermore, splenic macrophages from DBA/1 mice cultured in vitro with IL-18, but not IL-12, produced substantial amounts of TNF-α. Mice treated with IL-18 or IL-18 plus IL-12 produced markedly more IgG1 and IgG2a anti-collagen Ab compared with controls, whereas IL-12 treatment only led to an enhanced IgG2a response. Together these results demonstrate that IL-18 can promote collagen-induced inflammatory arthritis through mechanisms that may be distinct from those induced by IL-12.
Cyclic AMP Blocks Bacterial Lipopolysaccharide-Induced Myosin Light Chain Phosphorylation in Endothelial Cells Through Inhibition of Rho/Rho Kinase Signaling
Authors: Essler Markus , Staddon James M. , Weber Peter C. , Aepfelbacher Martin
Published: April 2014

During Gram-negative sepsis bacterial LPS induces endothelial cell contraction, actin reorganization, and loss of endothelial integrity by an unknown signal mechanism. In this study, we provide evidence that LPS-stimulation of endothelial cells (HUVEC) decreases myosin light chain (MLC) phosphatase, resulting in an increase in MLC phosphorylation followed by cell contraction. All of these LPS effects could be blocked by the Rho-GTPase inhibitor C3 transferase from Clostridium botulinum or the Rho kinase inhibitor Y-27632. These data suggest that LPS induces MLC phosphorylation via Rho/Rho kinase-mediated inhibition of MLC phosphatase in HUVEC. Furthermore, we observed that cAMP-elevating drugs, known to exert a vasoprotective function, mimicked the effects of C3 transferase and Y-27632, i.e., inhibited LPS-induced MLC phosphatase inactivation and MLC phosphorylation. cAMP elevation did not inhibit myosin phosphorylation induced by constitutively active V14Rho or the MLC phosphatase inhibitor calyculin and did not induce phosphorylation of RhoA in HUVEC, indicating inhibition of an upstream regulator of Rho/Rho kinase. Taken together, Rho/Rho kinase appears to be a central target for inflammatory mediators causing endothelial cell contraction such as bacterial toxins, but also for vasoprotective molecules elevating intracellular cAMP.
Recognition of Sulfamethoxazole and Its Reactive Metabolites by Drug-Specific CD4+ T Cells from Allergic Individuals
Authors: Schnyder Benno , Burkhart Christoph , Schnyder-Frutig Karin , von Greyerz Salome , Naisbitt Dean J. , Pirmohamed Munir , Park B. Kevin , Pichler Werner J.
Published: April 2014

The recognition of the antibiotic sulfamethoxazole (SMX) by T cells is usually explained with the hapten-carrier model. However, recent investigations have revealed a MHC-restricted but processing- and metabolism-independent pathway of drug presentation. This suggested a labile, low-affinity binding of SMX to MHC-peptide complexes on APC. To study the role of covalent vs noncovalent drug presentation in SMX allergy, we analyzed the proliferative response of PBMC and T cell clones from patients with SMX allergy to SMX and its reactive oxidative metabolites SMX-hydroxylamine and nitroso-SMX. Although the great majority of T cell clones were specific for noncovalently bound SMX, PBMC and a small fraction of clones responded to nitroso-SMX-modified cells or were cross-reactive. Rapid down-regulation of TCR expression in T cell clones upon stimulation indicated a processing-independent activation irrespective of specificity for covalently or noncovalently presented Ag. In conclusion, our data show that recognition of SMX presented in covalent and noncovalent bound form is possible by the same TCR but that the former is the exception rather than the rule. The scarcity of cross-reactivity between covalently and noncovalently bound SMX suggests that the primary stimulation may be directed to the noncovalently bound SMX.
Cutting Edge: Cytolytic Effector Function in Human Circulating CD8+ T Cells Closely Correlates with CD56 Surface Expression
Authors: Pittet Mikaël J. , Speiser Daniel E. , Valmori Danila , Cerottini Jean-Charles , Romero Pedro
Published: April 2014

Recent data suggest that human effector CD8+ T cells express a distinct CD27−CD45RAhigh (CD57+CD28−CD11ahigh) phenotype. Here, we propose that CTL effector function correlates better with CD56 (neuronal cell adhesion molecule (NCAM)) surface expression. CD56 was absent on cord blood CD8+ T cells, but was expressed by 4–30% of freshly isolated circulating CD8+ T cells from 15 adults. Dramatic oligoclonal expansions in 3/3 individuals were confined to the CD56+ subset of CD8+ T cells. The CD56+ subset generally contained high amounts of intracellular perforin and granzyme B. Finally, direct cytolytic capacity was closely restricted to the CD56+(CD45RAhigh) cells, better than to CD27−CD45RAhigh cells in 5/5 individuals analyzed. Thus, the phenotype corresponding to the circulating effector CD8+ T cell pool may be simplified and more precisely defined by the use of just two surface markers: CD8 and CD56.
Analysis of Immunomodulatory Activities of Aqueous Humor from Eyes of Mice with Experimental Autoimmune Uveitis
Authors: Ohta Kouichi , Wiggert Barbara , Yamagami Satoru , Taylor Andrew W. , Streilein J. Wayne
Published: April 2014

Aqueous humor (AqH) contains immunosuppressive factors, especially TGF-β2, that contribute to the immune privileged status of the anterior chamber. However, this may not be true when the blood-ocular barrier is compromised by ocular inflammation. To determine the immunosuppressive status of AqH from murine eyes afflicted with experimental autoimmune uveitis, B10.A mice were immunized with interphotoreceptor retinoid-binding protein. AqH was collected from eyes of affected mice periodically after immunization and then evaluated for content of TGF-β, proinflammatory cytokines, and the capacity to suppress anti-CD3-driven T cell proliferation. mRNA expression of selected cytokines in iris and ciliary body from inflamed eyes was analyzed by ribonuclease protection assay. We found that TGF-β levels were significantly increased in AqH from EAU eyes on days 11, 17, and 28. AqH collected on day 11 (onset of disease) failed to suppress T cell proliferation and contained large amounts of locally produced IL-6 that antagonized TGF-β. In contrast, AqH collected at 17 days (when ocular inflammation was progressively severe) re-expressed the ability to suppress T cell proliferation, in this case due to high levels of blood-derived TGF-β1 and eye-derived TGF-β2 in the absence of IL-6. Thus, during the onset of experimental autoimmune uveitis, the ocular microenvironment loses its immunosuppressive properties due to local production of IL-6. But as inflammation mounts, AqH IL-6 content falls, and the fluid reacquires sufficient TGF-β eventually to suppress immunogenic inflammation. The paradoxical roles of IL-6 in antagonizing TGF-β, while promoting TGF-β accumulation during ocular inflammation, is discussed.
Naive CD8+ T Cells Do Not Require Costimulation for Proliferation and Differentiation into Cytotoxic Effector Cells
Authors: Wang Bo , Maile Robert , Greenwood Roberta , Collins Edward J. , Frelinger Jeffrey A.
Published: April 2014

Most current models of T cell activation postulate a requirement for two distinct signals. One signal is delivered through the TCR by engagement with peptide/MHC complexes, and the second is delivered by interaction between costimulatory molecules such as CD28 and its ligands CD80 and CD86. Soluble peptide/MHC tetramers provide an opportunity to test whether naive CD8+ T cells can be activated via the signal generated through the TCR-αβ in the absence of any potential costimulatory molecules. Using T cells from two different TCR transgenic mice in vitro, we find that TCR engagement by peptide/MHC tetramers is sufficient for the activation of naive CD8+ T cells. Furthermore, these T cells proliferate, produce cytokines, and differentiate into cytolytic effectors. Under the conditions where anti-CD28 is able to enhance proliferation of normal B6 CD4+, CD8+, and TCR transgenic CD8+ T cells with anti-CD3, we see no effect of anti-CD28 on proliferation induced by tetramers. The results of this experiment argue that given a strong signal delivered through the TCR by an authentic ligand, no costimulation is required.
Phosphorylation of FADD/ MORT1 at Serine 194 and Association with a 70-kDa Cell Cycle-Regulated Protein Kinase
Authors: Scaffidi Carsten , Volkland Jörg , Blomberg Ida , Hoffmann Ingrid , Krammer Peter H. , Peter Marcus E.
Published: April 2014

The adapter molecule Fas-associated death domain protein (FADD)/mediator of receptor-induced toxicity-1 (MORT1) is essential for signal transduction of the apoptosis-inducing receptor CD95 (APO-1/Fas) as it connects the activated receptor with the effector caspase-8. FADD also plays a role in embryonic development and the cell cycle reentry of T cells. FADD is phosphorylated at serine residues. We now show that phosphorylation exclusively occurs at serine 194. The phosphorylation of FADD was found to correlate with the cell cycle. In cells arrested at the G2/M boundary with nocodazole, FADD was quantitatively phosphorylated, whereas only nonphosphorylated FADD was found in cells arrested in G1/S with hydroxyurea. In this context, we have identified a 70-kDa cell cycle-regulated kinase that specifically binds to the C-terminal half of FADD. Because CD95-mediated apoptosis is independent of the cell cycle, phosphorylation of FADD may regulate its apoptosis-independent functions.
CD5 Costimulation Up-Regulates the Signaling to Extracellular Signal-Regulated Kinase Activation in CD4+CD8+ Thymocytes and Supports Their Differentiation to the CD4 Lineage
Authors: Zhou Xu-Yu , Yashiro-Ohtani Yumi , Toyo-oka Kazuhito , Park Cheung-Seog , Tai Xu-Guang , Hamaoka Toshiyuki , Fujiwara Hiromi
Published: April 2014

CD5 positively costimulates TCR-stimulated mature T cells, whereas this molecule has been suggested to negatively regulate the activation of TCR-triggered thymocytes. We investigated the effect of CD5 costimulation on the differentiation of CD4+CD8+ thymocytes. Coligation of thymocytes with anti-CD3 and anti-CD5 induced enhanced tyrosine phosphorylation of LAT (linker for activation of T cells) and phospholipase C-γ (PLC-γ) compared with ligation with anti-CD3 alone. Despite increased phosphorylation of PLC-γ, this treatment down-regulated Ca2+ influx. In contrast, the phosphorylation of LAT and enhanced association with Grb2 led to activation of extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase. When CD3 and CD5 on CD4+CD8+ thymocytes in culture were coligated, they lost CD8, down-regulated CD4 expression, and induced CD69 expression, yielding a CD4+(dull)CD8−CD69+ population. An ERK inhibitor, PD98059, inhibited the generation of this population. The reduction of generation of CD4+CD8− cells resulted from decreased survival of these differentiating thymocytes. Consistent with this, PD98059 inhibited the anti-CD3/CD5-mediated Bcl-2 induction. These results indicate that CD5 down-regulates a branch of TCR signaling, whereas this molecule functions to support the differentiation of CD4+CD8+ thymocytes by up-regulating another branch of TCR signaling that leads to ERK activation.
Differential Regulation and Function of Fas Expression on Glial Cells
Authors: Lee Sung Joong , Zhou Tong , Choi Chulhee , Wang Zheng , Benveniste Etty N.
Published: April 2014

Fas/Apo-1 is a member of the TNF receptor superfamily that signals apoptotic cell death in susceptible target cells. Fas or Fas ligand (FasL)-deficient mice are relatively resistant to the induction of experimental allergic encephalomyelitis, implying the involvement of Fas/FasL in this disease process. We have examined the regulation and function of Fas expression in glial cells (astrocytes and microglia). Fas is constitutively expressed by primary murine microglia at a low level and significantly up-regulated by TNF-α or IFN-γ stimulation. Primary astrocytes express high constitutive levels of Fas, which are not further affected by cytokine treatment. In microglia, Fas expression is regulated at the level of mRNA expression; TNF-α and IFN-γ induced Fas mRNA by ∼20-fold. STAT-1α and NF-κB activation are involved in IFN-γ- or TNF-α-mediated Fas up-regulation in microglia, respectively. The cytokine TGF-β inhibits basal expression of Fas as well as cytokine-mediated Fas expression by microglia. Upon incubation of microglial cells with FasL-expressing cells, ∼20% of cells underwent Fas-mediated cell death, which increased to ∼60% when cells were pretreated with either TNF-α or IFN-γ. TGF-β treatment inhibited Fas-mediated cell death of TNF-α- or IFN-γ-stimulated microglial cells. In contrast, astrocytes are resistant to Fas-mediated cell death, however, ligation of Fas induces expression of the chemokines macrophage inflammatory protein-1β (MIP-1β), MIP-1α, and MIP-2. These data demonstrate that Fas transmits different signals in the two glial cell populations: a cytotoxic signal in microglia and an inflammatory signal in the astrocyte.
STAT1 Affects Lymphocyte Survival and Proliferation Partially Independent of Its Role Downstream of IFN-γ
Authors: Lee Chien-Kuo , Smith Eric , Gimeno Ramon , Gertner Rachel , Levy David E.
Published: April 2014

Lymphocytes derived from mice deficient in STAT1 showed reduced apoptosis and enhanced proliferation in vitro. To understand the involvement of STAT1 in the observed reduction in apoptosis, we examined the levels of caspase and bcl-2 family genes that are involved in cell survival and/or apoptosis. The levels of caspase 1 and 11, two enzymes involved in both cytokine protein processing and induction of apoptosis, were reduced in STAT1−/− cells compared with wild-type. However, the levels of bcl-2 genes were comparable in both mice. STAT1−/− cells also displayed an enhanced proliferation following TCR stimulation. This hyperproliferation could not be ascribed completely to the loss of IFN-γ-mediated antiproliferation. First, similar phenotypes were also observed in fibroblasts and pre-B cells derived from STAT1−/− mice, which do not produce IFN-γ. Second, comparisons with cells lacking the gene for IFN-γ or with cells treated with neutralizing Abs to IFN-γ only partially mimicked the STAT1−/− phenotype. Interestingly, the kinetics of degradation of p27kip1, a CDK inhibitor, following TCR ligation were faster, and, concomitantly, the up-regulation of CDK2 kinase activity and protein levels were increased in stimulated T cells of STAT1−/− mice relative to those of wild-type mice. Furthermore, STAT1−/− animals were more susceptible to carcinogen-induced thymic tumors, a possible consequence of altered T cell growth and/or survival. These results demonstrate an essential role for STAT1 for lymphocyte survival and proliferation that is only partially dependent on IFN-γ signaling.
The Nuclear Receptor PPARγ and Immunoregulation: PPARγ Mediates Inhibition of Helper T Cell Responses
Authors: Clark Robert B. , Bishop-Bailey David , Estrada-Hernandez Tatiana , Hla Timothy , Puddington Lynn , Padula Steven J.
Published: April 2014

The peroxisome proliferator-activated receptors (PPARs) are a family of transcription factors belonging to the nuclear receptor superfamily. Until recently, the genes regulated by PPARs were those believed to be predominantly associated with lipid metabolism. Recently, an immunomodulatory role for PPARγ has been described in cells critical to the innate immune system, the monocyte/macrophage. In addition, evidence for an antiinflammatory role of the PPARγ ligand, 15-deoxy-Δ12,14-PGJ2 (15d-PGJ2) has been found. In the present studies, we demonstrate, for the first time, that murine helper T cell clones and freshly isolated splenocytes express PPARγ 1. The PPARγ expressed is of functional significance in that two ligands for PPARγ, 15d-PGJ2 and a thiazolidinedione, ciglitazone, mediate significant inhibition of proliferative responses of both the T cell clones and the freshly isolated splenocytes. This inhibition is mediated directly at the level of the T cell and not at the level of the macrophage/APC. Finally, we demonstrate that the two ligands for PPARγ mediate inhibition of IL-2 secretion by the T cell clones while not inhibiting IL-2-induced proliferation of such clones. The demonstration of the expression and function of PPARγ in T cells reveals a new level of immunoregulatory control for PPARs and significantly increases the role and importance of PPARγ in immunoregulation.
Analyses of TCRB Rearrangements Substantiate a Profound Deficit in Recombination Signal Sequence Joining in SCID Foals: Implications for the Role of DNA-Dependent Protein Kinase in V(D)J Recombination
Authors: Shin Euy Kyun , Rijkers Tonnie , Pastink Albert , Meek Katheryn
Published: April 2014

We reported previously that the genetic SCID disease observed in Arabian foals is explained by a defect in V(D)J recombination that profoundly affects both coding and signal end joining. As in C.B-17 SCID mice, the molecular defect in SCID foals is in the catalytic subunit of the DNA-dependent protein kinase (DNA-PKCS); however, in SCID mice, signal end resolution remains relatively intact. Moreover, recent reports indicate that mice that completely lack DNA-PKCS also generate signal joints at levels that are indistinguishable from those observed in C.B-17 SCID mice, eliminating the possibility that a partially active version of DNA-PKCS facilitates signal end resolution in SCID mice. We have analyzed TCRB rearrangements and find that signal joints are reduced by ∼4 logs in equine SCID thymocytes as compared with normal horse thymocytes. A potential explanation for the differences between SCID mice and foals is that the mutant DNA-PKCS allele in SCID foals inhibits signal end resolution. We tested this hypothesis using DNA-PKCS expression vectors; in sum, we find no evidence of a dominant-negative effect by the mutant protein. These and other recent data are consistent with an emerging consensus: that in normal cells, DNA-PKCS participates in both coding and signal end resolution, but in the absence of DNA-PKCS an undefined end joining pathway (which is variably expressed in different species and cell types) can facilitate imperfect signal and coding end joining.
Liver CD4−CD8− NK1.1+ TCRαβ Intermediate Cells Increase During Experimental Malaria Infection and Are Able to Exhibit Inhibitory Activity Against the Parasite Liver Stage In Vitro
Authors: Pied Sylviane , Roland Jacques , Louise Anne , Voegtle Danièle , Soulard Valérie , Mazier Dominique , Cazenave Pierre-André
Published: April 2014

Experimental infection of C57BL/6 mice by Plasmodium yoelii sporozoites induced an increase of CD4−CD8− NK1.1+ TCRαβint cells and a down-regulation of CD4+ NK1.1+ TCRαβint cells in the liver during the acute phase of the infection. These cells showed an activated CD69+, CD122+, CD44high, and CD62Lhigh surface phenotype. Analysis of the expressed TCRVβ segment repertoire revealed that most of the expanded CD4−CD8− (double-negative) T cells presented a skewed TCRVβ repertoire and preferentially used Vβ2 and Vβ7 rather than Vβ8. To get an insight into the function of expanded NK1.1+ T cells, experiments were designed in vitro to study their activity against P. yoelii liver stage development. P. yoelii-primed CD3+ NK1.1+ intrahepatic lymphocytes inhibited parasite growth within the hepatocyte. The antiplasmodial effector function of the parasite-induced NK1.1+ liver T cells was almost totally reversed with an anti-CD3 Ab. Moreover, IFN-γ was in part involved in this antiparasite activity. These results suggest that up-regulation of CD4−CD8− NK1.1+ αβ T cells and down-regulation of CD4+ NK1.1+ TCRαβint cells may contribute to the early immune response induced by the Plasmodium during the prime infection.
Nitric Oxide Production by Human Intestinal Epithelial Cells and Competition for Arginine as Potential Determinants of Host Defense Against the Lumen-Dwelling Pathogen Giardia lamblia
Authors: Eckmann Lars , Laurent Fabrice , Langford T. Dianne , Hetsko Michael L. , Smith Jennifer R. , Kagnoff Martin F. , Gillin Frances D.
Published: April 2014

Giardia lamblia infection of the human small intestine is a common protozoan cause of diarrheal disease worldwide. Although infection is luminal and generally self-limiting, and secretory Abs are thought to be important in host defense, other defense mechanisms probably affect the duration of infection and the severity of symptoms. Because intestinal epithelial cells produce NO, and its stable end products, nitrite and nitrate, are detectable mainly on the apical side, we tested the hypothesis that NO production may constitute a host defense against G. lamblia. Several NO donors, but not their control compounds, inhibited giardial growth without affecting viability, suggesting that NO is cytostatic rather than cytotoxic for G. lamblia. NO donors also inhibited giardial differentiation induced by modeling crucial environmental factors, i.e., encystation induced by bile and alkaline pH, and excystation in response to gastric pH followed by alkaline pH and protease. Despite the potent antigiardial activity of NO, G. lamblia is not simply a passive target for host-produced NO, but has strategies to evade this potential host defense. Thus, in models of human intestinal epithelium, G. lamblia inhibited epithelial NO production by consuming arginine, the crucial substrate used by epithelial NO synthase to form NO. These studies define NO and arginine as central components in a novel cross-talk between a luminal pathogen and host intestinal epithelium.
C-C Chemokine Receptor 3 Antagonism by the β-Chemokine Macrophage Inflammatory Protein 4, a Property Strongly Enhanced by an Amino-Terminal Alanine-Methionine Swap
Authors: Nibbs Robert J. B. , Salcedo Theodora W. , Campbell John D. M. , Yao Xiao-Tao , Li Yuling , Nardelli Bernardetta , Olsen Henrik S. , Morris Tina S. , Proudfoot Amanda E. I. , Patel Vikram P. , Graham Gerard J.
Published: April 2014

Allergic reactions are characterized by the infiltration of tissues by activated eosinophils, Th2 lymphocytes, and basophils. The β-chemokine receptor CCR3, which recognizes the ligands eotaxin, eotaxin-2, monocyte chemotactic protein (MCP) 3, MCP4, and RANTES, plays a central role in this process, and antagonists to this receptor could have potential therapeutic use in the treatment of allergy. We describe here a potent and specific CCR3 antagonist, called Met-chemokine β 7 (Ckβ7), that prevents signaling through this receptor and, at concentrations as low as 1 nM, can block eosinophil chemotaxis induced by the most potent CCR3 ligands. Met-Ckβ7 is a more potent CCR3 antagonist than Met- and aminooxypentane (AOP)-RANTES and, unlike these proteins, exhibits no partial agonist activity and is highly specific for CCR3. Thus, this antagonist may be of use in ameliorating leukocyte infiltration associated with allergic inflammation. Met-Ckβ7 is a modified form of the β-chemokine macrophage inflammatory protein (MIP) 4 (alternatively called pulmonary and activation-regulated chemokine (PARC), alternative macrophage activation-associated C-C chemokine (AMAC) 1, or dendritic cell-derived C-C chemokine (DCCK) 1). Surprisingly, the unmodified MIP4 protein, which is known to act as a T cell chemoattractant, also exhibits this CCR3 antagonistic activity, although to a lesser extent than Met-Ckβ7, but to a level that may be of physiological relevance. MIP4 may therefore use chemokine receptor agonism and antagonism to control leukocyte movement in vivo. The enhanced activity of Met-Ckβ7 is due to the alteration of the extreme N-terminal residue from an alanine to a methionine.
A Soluble Recombinant Multimeric Anti-Rh(D) Single-Chain Fv/CR1 Molecule Restores the Immune Complex Binding Ability of CR1-Deficient Erythrocytes
Authors: Oudin S. , Libyh M. Tonye , Goossens D. , Dervillez X. , Philbert F. , Réveil B. , Bougy F. , Tabary T. , Rouger P. , Klatzmann D. , Cohen J. H. M.
Published: April 2014

CR1 (CD35, the C3b/C4b receptor) is a widely distributed membrane glycoprotein with a unique cluster conformation on the surface of erythrocytes (E). CR1 on E is responsible for the transport of immune complexes (IC) to liver and spleen. As a cofactor of the C3b cleavage by factor I, CR1 is also a potent inhibitor of C activation and inflammation. In some diseases (systemic lupus erythematosus, hemolytic anemia, AIDS, etc.) an acquired low level of CR1 on E has been observed, leading to an impaired clearance of IC. The aim of this study was to design a heterofunctional molecule that will bind to E and restore a normal or a supranormal CR1 density on E that could mimic the unique distribution pattern of CR1 on normal E. For that purpose a new multimerizing system based on the properties of the C-terminal part of the α-chain of the C4 binding protein (C4bp) was used. We first produced a multimeric soluble CR1 that proved to be a better inhibitor of in vitro C activation than the monomeric form of CR1, then a heteromultimeric molecule made of CR1 and single-chain Fv anti-Rh(D) valences able to attach E and providing E with as much as a 10-fold increase in CR1 density with the same CR1 distribution pattern as native E. CR1/single-chain Fv anti-Rh(D)-treated E were able in vitro to attach as many opsonized IC as native E. These data open the way for future use of multimeric and heteromultimeric forms of soluble recombinant CR1 as therapy of IC diseases.
Evidence for Common Autoimmune Disease Genes Controlling Onset, Severity, and Chronicity Based on Experimental Models for Multiple Sclerosis and Rheumatoid Arthritis
Authors: Bergsteinsdottir Kristin , Yang Hai-Tao , Pettersson Ulf , Holmdahl Rikard
Published: April 2014

The pathogenicity of multiple sclerosis is still poorly understood, but identification of susceptibility genes using the animal model experimental allergic encephalomyelitis (EAE) could provide leads. Certain genes may be shared between different autoimmune diseases, and identification of such genes is of obvious importance. To locate gene regions involved in the control of EAE and to compare the findings with the susceptibility loci recently identified in a model for rheumatoid arthritis (pristane-induced arthritis), we made crosses between the encephalomyelitis- and arthritis-susceptible rat strain DA and the resistant E3 strain. Genetic analysis of animals produced in a F2 intercross identified 11 loci associated with specific EAE-associated traits. Interestingly, five of these loci were situated at the same position as major loci controlling pristane-induced arthritis and showed similarities in inheritance pattern and subphenotype associations. Our results show that different phases of EAE are controlled by different sets of genes and that common genes are likely to be involved in different autoimmune diseases.
Viral IL-10 and Soluble TNF Receptor Act Synergistically to Inhibit Collagen-Induced Arthritis Following Adenovirus- Mediated Gene Transfer
Authors: Kim Kwang-Nam , Watanabe Shohei , Ma Yuhe , Thornton Sherry , Giannini Edward H. , Hirsch Raphael
Published: April 2014

Viral IL-10 (vIL-10) and soluble TNF receptor (sTNFR) are anti-inflammatory proteins that can suppress collagen-induced arthritis (CIA). These and related proteins have shown efficacy in the treatment of human rheumatoid arthritis; however, neither alone is able to completely suppress disease. Furthermore, they have short half-lives, necessitating frequent administration. To determine the ability of these proteins to act synergistically following gene transfer, arthritis was induced in DBA/1 male mice by immunization with type II collagen on days 0 and 21. Mice were injected i.v. either before disease onset (day 20) or after disease onset (day 28) with 1010 particles of adenovirus encoding vIL-10, a soluble TNF receptor-IgG1 fusion protein (sTNFR-Ig), a combination of both vectors, or a control vector lacking a transgene. Significant synergism was observed with the combination of vIL-10 and sTNFR-Ig, with a substantial reduction in both the incidence and severity of disease as well as inhibition of progression of established disease. sTNFR-Ig alone had no effect on CIA. vIL-10 alone inhibited disease when given before disease onset, but had minimal effect on established disease. Both proteins inhibited spleen cell proliferation and IFN-γ secretion in response to stimulation with type II collagen, but only vIL-10 reduced the synovial mRNA levels of the proinflammatory cytokines IL-1β, TNF-α, and IL-6. These findings demonstrate that vIL-10 and sTNFR-Ig act synergistically in suppressing CIA and suggest that gene transfer offers a potential therapeutic modality for the treatment of arthritis.
HIV-1-Specific Mucosal CD8+ Lymphocyte Responses in the Cervix of HIV-1-Resistant Prostitutes in Nairobi
Authors: Kaul Rupert , Plummer Francis A. , Kimani Joshua , Dong Tao , Kiama Peter , Rostron Timothy , Njagi Ephantus , MacDonald Kelly S. , Bwayo Job J. , McMichael Andrew J. , Rowland-Jones Sarah L.
Published: April 2014

Understanding how individuals with a high degree of HIV exposure avoid persistent infection is paramount to HIV vaccine design. Evidence suggests that mucosal immunity, particularly virus-specific CTL, could be critically important in protection against sexually acquired HIV infection. Therefore, we have looked for the presence of HIV-specific CD8+ T cells in cervical mononuclear cells from a subgroup of highly HIV-exposed but persistently seronegative female sex workers in Nairobi. An enzyme-linked immunospot assay was used to measure IFN-γ release in response to known class I HLA-restricted CTL epitope peptides using effector cells from the blood and cervix of HIV-1-resistant and -infected sex workers and from lower-risk uninfected controls. Eleven of 16 resistant sex workers had HIV-specific CD8+ T cells in the cervix, and a similar number had detectable responses in blood. Where both blood and cervical responses were detected in the same individual, the specificity of the responses was similar. Neither cervical nor blood responses were detected in lower-risk control donors. HIV-specific CD8+ T cell frequencies in the cervix of HIV-resistant sex workers were slightly higher than in blood, while in HIV-infected donor cervical response frequencies were markedly lower than blood, so that there was relative enrichment of cervical responses in HIV-resistant compared with HIV-infected donors. HIV-specific CD8+ T cell responses in the absence of detectable HIV infection in the genital mucosa of HIV-1-resistant sex workers may be playing an important part in protective immunity against heterosexual HIV-1 transmission.
Linear PADRE T Helper Epitope and Carbohydrate B Cell Epitope Conjugates Induce Specific High Titer IgG Antibody Responses
Authors: Alexander Jeff , del Guercio Marie-France , Maewal Ajesh , Qiao Lei , Fikes John , Chesnut Robert W. , Paulson James , Bundle David R. , DeFrees Shawn , Sette Alessandro
Published: April 2014

Linear carbohydrate-peptide constructs based on the 13 amino acid nonnatural pan DR epitope (PADRE) and carbohydrate B cell epitopes are demonstrated to be potent immunogens. These data support our belief that PADRE should be considered as an alternative to more complex carriers for use in prophylaxis and therapeutic vaccines. Two model carbohydrate-PADRE glycoconjugates were used to demonstrate that PADRE could effectively provide T cell help for carbohydrate-specific Ab responses. Conjugates of PADRE covalently linked to the human milk oligosaccharide, lacto-N-fucopentose II or a dodecasaccharide derived from Salmonella typhimurium O-Ag induced high titer IgG Ab responses in mice, which were comparable to glycoconjugates employing human serum albumin (HSA) as the carrier protein. Different adjuvants, in combination with PADRE conjugates, allowed for the modulation of the isotype profile with alum supporting an IgG1 profile; QS-21 an IgG2a, 2b profile, while an alum/QS-21 mixture generated a balanced IgG1/IgG2b isotype profile. As defined by binding to synthetic glycoconjugates, dodecasaccharide-specific Abs exhibited fine specificity similar to protective polyclonal Ab responses previously reported for dodecasaccharide-protein conjugates. The same Abs bound to intact S. typhimurium cells, suggesting that biologically relevant specificities were produced. The affinity of the dodecasaccharide-specific Abs was further shown to be comparable to that of a well-characterized, high affinity monoclonal anti-carbohydrate Ab recognizing the same epitope.
Cutting Edge: Lipoxin (LX) A4 and Aspirin-Triggered 15-Epi-LXA4 Block Allergen-Induced Eosinophil Trafficking
Authors: Bandeira-Melo Christianne , Bozza Patricia T. , Diaz Bruno L. , Cordeiro Renato S. B. , Jose Peter J. , Martins Marco A. , Serhan Charles N.
Published: April 2014

Tissue eosinophilia prevention represents one of the primary targets to new anti-allergic therapies. As lipoxin A4 (LXA4) and aspirin-triggered 15-epi-LXA4 (ATL) are emerging as endogenous “stop signals” produced in distinct pathologies including some eosinophil-related pulmonary disorders, we evaluated the impact of in situ LXA4/ATL metabolically stable analogues on allergen-induced eosinophilic pleurisy in sensitized rats. LXA4/ATL analogues dramatically blocked allergic pleural eosinophil influx, while concurrently increasing circulating eosinophilia, inhibiting the earlier edema and neutrophilia associated with allergic reaction. The mechanisms underlying this LXA4/ATL-driven allergic eosinophilia blockade was independent of mast cell degranulation and involved LXA4/ATL inhibition of both IL-5 and eotaxin generation, as well as platelet activating factor action. These findings reveal LXA4/ATL as a novel class of endogenous anti-allergic mediators, capable of preventing local eosinophilia.
TGF-β1 Reciprocally Controls Chemotaxis of Human Peripheral Blood Monocyte-Derived Dendritic Cells Via Chemokine Receptors
Authors: Sato Katsuaki , Kawasaki Hiroshi , Nagayama Hitomi , Enomoto Makoto , Morimoto Chikao , Tadokoro Kenji , Juji Takeo , Takahashi Tsuneo A.
Published: April 2014

We examined the effect of TGF-β1 on the chemotactic migratory ability of human monocyte-derived dendritic cells (DCs). Treatment of immature DCs with TGF-β1 resulted in increased expressions of CCR-1, CCR-3, CCR-5, CCR-6, and CXC chemokine receptor-4 (CXCR-4), which were concomitant with enhanced chemotactic migratory responses to their ligands, RANTES (for CCR-1, CCR-3, and CCR-5), macrophage-inflammatory protein-3α (MIP-3α) (for CCR-6), or stromal cell-derived growth factor-1α (for CXCR-4). Ligation by TNF-α resulted in down-modulation of cell surface expressions of CCR-1, CCR-3, CCR-5, CCR-6, and CXCR-4, and the chemotaxis for RANTES, MIP-3α, and stromal cell-derived growth factor-1α, whereas this stimulation up-regulated the expression of CCR-7 and the chemotactic ability for MIP-3β. Stimulation of mature DCs with TGF-β1 also enhanced TNF-α-induced down-regulation of the expressions of CCR-1, CCR-3, CCR-5, CCR-6, and CXCR-4, and chemotaxis to their respective ligands, while this stimulation suppressed TNF-α-induced expression of CCR-7 and chemotactic migratory ability to MIP-3β. Our findings suggest that TGF-β1 reversibly regulates chemotaxis of DCs via regulation of chemokine receptor expression.
IFN-α2b Reduces IL-2 Production and IL-2 Receptor Function in Primary CD4+ T Cells
Authors: Zella Davide , Romerio Fabio , Curreli Sabrina , Secchiero Paola , Cicala Claudia , Zagury Daniel , Gallo Robert C.
Published: April 2014

Initially described as an antiviral cytokine, IFN-α has been subsequently shown to affect several cellular functions, including cellular differentiation and proliferation. For these reasons, IFN-α is currently used in clinical practice for the treatment of viral infections and malignancies. In this manuscript, we show two novel mechanisms concomitantly responsible for the antiproliferative effect of IFN-α. First, long-term treatment with IFN-α of primary CD4+ T cells reduced surface expression of CD3 and CD28. These events resulted in decreased phosphorylation of the mitogen-activated extracellular signal-regulated activating kinase and its substrate extracellular signal-regulated kinase, leading to diminished production of IL-2. Second, IFN-α treatment of primary CD4+ T cells reduced proliferative response to stimulation in the presence of exogenous IL-2 by markedly decreasing mRNA synthesis and surface expression of CD25 (α-chain), a critical component of the IL-2R complex. These results may be relevant for the antitumor effects of IFN-α and may help us to better understand its detrimental role in the inhibition of proliferation of the bulk of CD4+ T cells (uninfected cells) in HIV-infected persons, who are known to overproduce IFN-α.
Stat6 Regulation of In Vivo IL-4 Responses
Authors: Finkelman Fred D. , Morris Suzanne C. , Orekhova Tatyana , Mori Masaaki , Donaldson Debra , Reiner Steven L. , Reilly Nancy L. , Schopf Lisa , Urban Joseph F.
Published: April 2014

Although in vitro development of a Th2 response from naive CD4+ T cells is Stat6 dependent, mice immunized with a goat Ab to mouse IgD have been reported to produce a normal primary IL-4 response in Stat6-deficient mice. Experiments have now been performed with mice immunized with more conventional Ags or inoculated with nematode parasites to account for this apparent discrepancy. The ability of an immunogen to induce a primary in vivo IL-4 response in Stat6-deficient mice was found to vary directly with its ability to induce a strong type 2 cytokine-biased response in normal mice. Even immunogens, however, that induce strong primary IL-4 responses in Stat6-deficient mice induce poor memory IL-4 responses in these mice. Consistent with this, Stat6-deficient CD4+ T cells make relatively normal IL-4 responses when stimulated in vitro for 3 days with anti-CD3 and anti-CD28, but poor IL-4 responses if they are later restimulated with anti-CD3. Thus, Stat6 signaling enhances primary IL-4 responses that are made as part of a type 0 cytokine response (mixed type 1 and type 2) and is required for normal development or survival of Th2 memory cells.
4-1BB Costimulation Is Required for Protective Anti-Viral Immunity After Peptide Vaccination
Authors: Tan Joyce T. , Whitmire Jason K. , Murali-Krishna Kaja , Ahmed Rafi , Altman John D. , Mittler Robert S. , Sette Alessandro , Pearson Thomas C. , Larsen Christian P.
Published: April 2014

Peptide vaccination induces T cell activation and cytotoxic T cell development. In an effort to understand what factors can improve immune responses to peptide vaccination, the role of 4-1BB (CD137) costimulation was examined, since 4-1BB has been shown to promote T cell responses in other systems. 4-1BBL-deficient (−/−) and wild-type (+/+) mice were immunized with a lipidated lymphocytic choriomeningitis virus (LCMV) peptide NP396–404. Analysis of peptide-specific responses early after immunization by CTL assay, intracellular IFN-γ staining, and IFN-γ enzyme-linked immunospot assay (ELISPOT) indicated that CD8 T cell responses were reduced 3- to 10-fold in the absence of 4-1BB costimulation. Moreover, when agonistic anti-4-1BB Ab was given, CD8 T cell responses in 4-1BBL−/− mice were augmented to levels similar to those in 4-1BBL+/+ mice. Two months after immunization, 4-1BBL+/+ mice still had epitope-specific cells and were protected against viral challenge, demonstrating that peptide vaccination can induce long-term protection. In fact, 70% of CD8 T cells were specific for the immunizing peptide after viral challenge, demonstrating that strong, epitope-specific CD8 T cell responses are generated after peptide vaccination. In contrast, peptide-immunized 4-1BBL−/− mice had fewer epitope-specific cells and were impaired in their ability to resolve the infection. These results show that immunization with a single LCMV peptide provides long term protection against LCMV infection and point to costimulatory molecules such as 4-1BB as important components for generating protective immunity after vaccination.
Heat Shock Protein 70 Is Able to Prevent Heat Shock-Induced Resistance of Target Cells to CTL
Authors: Dressel Ralf , Elsner Leslie , Quentin Thomas , Walter Lutz , Günther Eberhard
Published: April 2014

Heat shock or transfection with heat shock protein 70 (Hsp70) genes has been shown to protect tumor cell lines against immune mechanisms of cytotoxicity. We have reported previously that heat shock confers resistance to CTL in the rat myeloma cell line Y3 that is Hsp70 defective. Evidence is now presented that Hsp70 is able to prevent the induction of the resistant phenotype. In Con A-stimulated lymphocytes and in lymphocyte × Y3 somatic cell hybrid clones a severe, non-Hsp70-inducing heat shock elicits resistance to CTL in contrast to a heat shock that results in Hsp70 expression. Thus, Hsp70 expression appears to be negatively associated with the development of resistance. Furthermore, loading of Y3 cells with recombinant Hsp70 protein before heat shock is able to prevent resistance. Because apoptosis induced in Y3 cells by heat shock is not affected, Hsp70 appears to interfere selectively with the CTL-induced lethal pathway that is found to be calcium but not caspase dependent. It is suggested that after heat shock Hsp70 enhances the CTL-induced apoptotic pathway by chaperoning certain proteins in the target cell that are involved in the execution of cell death. Thus, although shown to confer protection against many cytotoxic mechanisms, Hsp70 does not appear to be generally cytoprotective. This observation could also be of relevance when interpreting the effectiveness of tumor immunity.
CpG-DNA Activates In Vivo T Cell Epitope Presenting Dendritic Cells to Trigger Protective Antiviral Cytotoxic T Cell Responses
Authors: Vabulas Ramunas M. , Pircher Hanspeter , Lipford Grayson B. , Häcker Hans , Wagner Hermann
Published: April 2014

MHC class I-restricted T cell epitopes lack immunogenicity unless aided by IFA or CFA. In an attempt to circumvent the known inflammatory side effects of IFA and CFA, we analyzed the ability of immunostimulatory CpG-DNA to act as an adjuvant for MHC class I-restricted peptide epitopes. Using the immunodominant CD8 T cell epitopes, SIINFEKL from OVA or KAVYNFATM (gp33) from lymphocytic choriomeningitis virus glycoprotein, we observed that CpG-DNA conveyed immunogenicity to these epitopes leading to primary induction of peptide-specific CTL. Furthermore, vaccination with the lymphocytic choriomeningitis virus gp33 peptide triggered not only CTL but also protective antiviral defense. We also showed that MHC class I-restricted peptides are constitutively presented by immature dendritic cells (DC) within the draining lymph nodes but failed to induce CTL responses. The use of CpG-DNA as an adjuvant, however, initiated peptide presenting immature DC progression to professional licensed APC. Activated DC induced cytolytic CD8 T cells in wild-type mice and also mice deficient of Th cells or CD40 ligand. CpG-DNA thus incites CTL responses toward MHC class I-restricted T cell epitopes in a Th cell-independent manner. Overall, these results provide new insights into CpG-DNA-mediated adjuvanticity and may influence future vaccination strategies for infectious and perhaps tumor diseases.
HLA-DR-Mediated Apoptosis Susceptibility Discriminates Differentiation Stages of Dendritic/Monocytic APC
Authors: Bertho Nicolas , Drénou Bernard , Laupeze Béatrice , Berre Claudine Le , Amiot Laurence , Grosset Jean-Marc , Fardel Olivier , Charron Dominique , Mooney Nuala , Fauchet Renée
Published: April 2014

Professional APC are characterized by their ability to present peptide via HLA class II in the presence of costimulatory molecules (CD40, CD80, and CD86). The efficiency of Ag presentation can be classed as follows: mature dendritic cells (DC) are most efficient, immature DC and macrophages are intermediate, and monocytes are considered poor APC. There is a large body of evidence demonstrating that HLA-DR transmits signals in the APC. In this study, we have addressed the question of the outcome of HLA-DR signals on APC of the monocyte/DC lineages throughout their differentiation from immature to mature APC. DC were generated from both monocytes and CD34+ cells of the same individual, macrophages were differentiated from monocytes. Immunophenotypical analysis clearly distinguished these populations. HLA-DR-mediated signals led to marked apoptosis in mature DC of either CD34 or monocytic origin. Significantly less apoptosis was observed in immature DC of either origin. Nonetheless, even immature DC were more susceptible to HLA-DR-mediated apoptosis than macrophages, whereas monocytes were resistant to HLA-DR-mediated apoptosis. The mechanism of HLA-DR-mediated apoptosis was independent of caspase activation. Taken together, these data lead to the notion that signals generated via HLA-DR lead to the demise of mature professional APC, thereby providing a means of limiting the immune response.