T cell activation

Whole body hyperthermia (WBH) has been used as an adjunct to radio-/chemotherapy in patients with various malignant diseases. Although clear evidence is still missing, it has been hypothesized that an activation of the immune system might contribute to the therapeutic effect of WBH. To examine whether a treatment with 60-minute 41.8°C WBH as an adjunct to chemotherapy (WBH-CT) induces an activation of T cells, blood samples were collected at numerous time points before and up to 48 h post-treatment. The aim of this study was to examine the effect of WBH-CT on the expression of a broad range of activation markers on peripheral blood lymphocytes (PBL), on serum cytokines and intracellular cytokine levels in T cells, and the capacity of these cells to proliferate. Immediately after 41.8°C WBH-CT treatment, a drastic increase in peripheral natural killer (NK) cells (P<0.05) and CD56 + cytotoxic T lymphocytes (CTL; P<0.01) in the patients' peripheral blood was observed. At 5 h post-treatment, the percentages of both effector cell types had returned to baseline levels. This transient phenomenon was accompanied by a short period of reduced T cell activity, indicated by diminished serum levels of soluble interleukin-2 receptors (sIL-2R) at 3 h post-WBH-CT (P<0.05) and decreased lymphocytic proliferation at the same point in time. This first phase was followed by a marked but short-lived increase in the patients' serum levels of interleukin-6 (IL-6; P<0.01) during the first 5 h following treatment, with a subsequent decrease to baseline levels at 24 h and significantly increased serum levels of tumor necrosis factor-a (TNF-a) at 0 h (P<0.01), 3 h (P<0.05), 5 h (P<0.05) and 24 h (P<0.01) post-WBH-CT. The third phase of the immunological consequences of WBH-CT consisted of an increase in the percentage of peripheral cytotoxic T lymphocytes (CTL) expressing CD56, reaching a maximum at 48 h post-WBH (P<0.01). Furthermore, the percentage of CD4 + T cells expressing the T cell activation marker CD69 increased nearly twofold over time, reaching its maximum at 48 h (P<0.05). As an additional marker for T cell activation, serum levels of sIL-2R increased markedly (P<0.01), reaching maximum levels at the same point in time. Elevated intracellular concentrations of interferon-gamma (IFN-c) and/or TNF-a in CD8 + T cells were found in 4 out of 5 patients at 24 h post-WBH-CT. Since similar changes were not observed in patients receiving chemotherapy alone, this is the first study to provide evidence for prolonged WBH-CT-induced activation of human T cells.

Glucocorticoids (GCs) are important steroid hormones with widespread activities in metabolism, development, and immune regulation. The adrenal glands are the major source of GCs and release these hormones in response to psychological and immunological stress. However, there is increasing evidence that GCs may also be synthesized by nonadrenal tissues. Here, we report that the intestinal mucosa expresses steroidogenic enzymes and releases the GC corticosterone in response to T cell activation. T cell activation causes an increase in the intestinal expression of the steroidogenic enzymes required for GC synthesis. In situ hybridization analysis revealed that these enzymes are confined to the crypt region of the intestinal epithelial layer. Surprisingly, in situ-produced GCs exhibit both an inhibitory and a costimulatory role on intestinal T cell activation. In the absence of intestinal GCs in vivo , activation by anti-CD3 injection resulted in reduced CD69 expression and interferon-␥ production by intestinal T cells, whereas activation by viral infection led to increased T cell activation. We conclude that the intestinal mucosa is a potent source of immunoregulatory GCs.

The mouse 4T1 mammary carcinoma is a BALB/c-derived tumor that spontaneously metastasizes and induces immune suppression. Although >95% of wild type BALB/c mice die from metastatic 4T1 tumor even if the primary mammary tumor is surgically removed, >65% of BALB/c mice with a deleted Signal Transducer Activator of Transcription 6 (STAT6) gene survive post-surgery. STAT6-deficiency also confers enhanced immunity against spontaneously developing breast cancer since NeuT +/) mice that are STAT6deficient develop mammary tumors later and survive longer than NeuT +/) mice that are STAT6-competent. Rejection of metastastic disease and survival of STAT6deficient mice after removal of primary tumor involve three mechanisms: (1) The generation of M1 type macrophages that produce nitric oxide and are tumoricidal; (2) A decrease to normal in the elevated levels of myeloid suppressor cells that accumulate during primary tumor growth; and (3) CD8 + tumor-specific T lymphocytes. STAT6-deficient, but not wild type BALB/c, mice generate nitric oxide producing macrophages because they lack the STAT6 transcription factor which is necessary for signaling through the type 2 IL-4Ra complex, and which induces the production of arginase instead of nitric oxide. Keywords Tumor-induced immune suppression AE Immune surveillance AE M1 macrophages AE Metastatic breast cancer AE Cell-mediated tumor immunity Signal Transducer Activator of Transcription 6 deficient (STAT6-/-) mice have enhanced immunity to transplanted tumors This article is a symposium paper from the conference ''Tumor Escape and Its Determinants''

The small chemical vacuolin-1 induces rapid formation of large vacuoles in various cell types. In epithelial cells, vacuolin-1 has been shown to inhibit Ca 2+ ionophore-induced exocytosis depending on experimental conditions used but had no effect on repair of damaged membranes. However, it is not known whether vacuolin-1 could inhibit exocytosis induced by immunoreceptor triggering in professional secretory cells and whether there is any correlation between effect of vacuolin-1 on exocytosis and membrane repair in such cells. Here we show that in rat basophilic leukemia (RBL-2H3) cells activated by the high-affinity IgE receptor (FcεRI) triggering vacuolin-1 enhanced exocytosis. Under identical conditions of activation, vacuolin-1 inhibited exocytosis in mouse bone marrow-derived mast cells (BMMCs). This inhibition was not reflected by decreased phosphorylation of the FcεRI α and β subunits, linker for activation of T cells, non-T cell activation linker, Akt and MAP kinase Erk, and uptake of extracellular Ca 2+ , indicating that early activation events are not affected. In both cell types vacuolin-1 led to formation of numerous vacuoles, a process which was inhibited by bafilomycin A1, an inhibitor of vacuolar H + -ATPase. Thapsigargin-or Ca 2+ ionophore A23187induced exocytosis also showed different sensitivity to the inhibitory effect of vacuolin-1. Pretreatment of the cells with vacuolin-1 followed by permeabilization with bacterial toxin streptolysin O enhanced Ca 2+dependent repair of plasma membrane lesions in RBL-2H3 cells but inhibited it in BMMCs. Our data indicate that lysosomal exocytosis exhibits different sensitivity to vacuolin-1 depending on the cell type analyzed and mode of activation. Furthermore, our results support the concept that lysosomal exocytosis is involved in the repair of injured plasma membranes.

Enteropathy defines abnormalities of the small intestinal mucosa of various etiologies in which nutrition has a causal or a therapeutic role. Breast milk is the gold-standard feeding during infancy for optimal nutrition in health and the majority of diseases. Therapeutic formulae have decreased the need for enteral or parenteral support. Gastrointestinal infections are worldwide the most frequent cause of enteropathy by increasing mucosal permeability, local expression of costimulatory molecules allowing antigen penetration in the mucosa, and T-cell activation leading sometimes to disruption of oral tolerance. Concomitant malnutrition impairs not only the immunologic response but also the recovery of damaged mucosa with secondary intestinal and pancreatic enzymatic reductions. Optimal nutritional rehabilitation is the cornerstone of the management of persisting diarrhea. Celiac disease and cow's milk protein allergy are examples of chronic enteropathy. Multiple food allergies, even during breast-feeding, are increasingly reported due to an impaired development of oral tolerance. The dietary approach to allergic disease is currently evolving from passive allergen avoidance to active modulation of the immune system to (re)establish tolerance. The gastrointestinal flora provides maturational signals for the lymphoid tissue, improves balance of inflammatory cytokines, reduces bacterial invasiveness and dietary antigen load, and normalizes gut permeability. The clinical effects of nucleotides and zinc merit further clinical evaluation. Major attention has recently focused on the immune effects of dietary lipids in terms of possible prevention of allergic sensitization by downregulating inflammatory response and protecting the epithelial barrier and host-microbe interactions modifying the adherence of microbes to the mucosa.

Manufacturing of cellular products for therapeutic purposes like stem cell or cancer therapy requires equipment with specific characteristics not always addressed by conventional technologies. A new integrated cell processing device is presented that can handle all current technical requirements for manufacturing cellular products by automation of the complete process in a GMP-compliant single-use tubing set. Its capabilities are exemplified in the presented study by successful processing of adult stem cells, natural killer cells, and several cell lines. Multiple cell processing workflows can be automated in a functionally closed environment: from cell separation through cell culture to formulation of the final product.

Introduction Immunodeficiency with hyper-IgM (HIGM) results from genetic defects in the CD40-CD40 ligand (CD40L) pathway or in the enzymes required for immunoglobulin class switch recombination and somatic hypermutation. HIGM can thus be associated with an impairment of both B-cell and T-cell activation. Results and discussions There are seven main subtypes of HIGM and the most frequent is X-linked HIGM, resulting from CD40L mutations. In addition to the susceptibility to recurrent and opportunistic infections, these patients are prone to autoimmune manifestations, especially hematologic abnormalities, arthritis, and inflammatory bowel disease. Furthermore, organ-specific autoantibodies are commonly found in HIGM patients. Conclusions The mechanisms by which HIGM associates to autoimmunity are not completely elucidated but a defective development of regulatory T cells, the presence of IgM autoantibodies and an impaired peripheral B-cell tolerance checkpoint have been implicated. This article reviews the main subtypes of HIGM syndrome, the clinical autoimmune manifestations found in these patients, and the possible mechanisms that would explain this association.

Although the pathogenesis of systemic sclerosis (SSc) remains unknown, cytokine production and release are key events in this autoimmune disease, characterized by T cell activation and auto-antibodies production leading to microvascular damage, inflammation and fibrosis. We review herein experimental and clinical data, aiming to analyze the relationship between cytokine release and SSc pathogenesis. Measurement of circulating or in situ cytokine levels provides evidence for a balance between "Th1/Th2" or "Th17/Treg" cytokines in the development of SSc. Indeed, the Th2 cytokine response, with the production of IL-4, IL-10 and TGF-β, leads to tissue fibrosis, whereas Th1 and Th17 cytokines promote inflammation in SSc patients. Thus, cytokine levels have been assessed as diagnostic or prognostic markers in SSc patients. Restoration of the Th1/Th2/Th17/Treg balance is one of the hallmarks of treatment effectiveness and development of cytokine modulators could be considered for new therapeutic approaches in SSc patients.

The heavy metal mercury (Hg) is known to have immunomodulatory properties affecting lymphocyte signal transduction, death receptor signaling and autoimmunity. In this study we tested the hypothesis that Hg exposure would attenuate T-cell activation and caspase 8 and 3 activity in response to antigenic stimuli. To test this hypothesis, BALB/cJ mice were exposed to 10 mg/L mercuric chloride (HgCl 2 ) in their drinking water for two weeks followed injection with 20μg of the Staphylococcal aureus enterotoxin B (SEB) superantigen. Eighteen hours after SEB challenge, there was a statistically significant reduction in caspase 8 and caspase 3 enzyme activity in the SEB reactive Vβ8+ T-cells. The attenuated caspase activity in Hg exposed mice persisted for 48 hours after exposure. Moreover, activation of caspase 8 and caspase 3 was reduced by more than 60% in CD95 deficient MRL/MpJ-Fas lpr mice demonstrating caspase 8 and 3 activation in response to SEB is CD95 dependent. In addition to the effects of Hg on caspase activity, expression of the T-cell activation marker CD69 was also attenuated in SEB reactive Vβ8 T-cells in Hg-exposed mice. Moreover, CD69 expression in MRL/MpJ-Fas lpr mice was also reduced. Taken together the caspase and CD69 data support a role for CD95 in promoting a proapoptotic and activated state in SEB responsive T-lymphocytes and this state is attenuated by the autoimmune potentiating environmental agent mercury.

Anergy is a major mechanism to ensure antigen-specific tolerance in T lymphocytes in the adult. In vivo, anergy has mainly been studied at the cellular level. In this study, we used the T-cell-activating superantigen staphylococcal enterotoxin A (SEA) to investigate molecular mechanisms of T-lymphocyte anergy in vivo. Injection of SEA to adult mice activates CD4+ T cells expressing certain T-cell

Vitamin D has received increased attention recently for its pleiotropic actions on many chronic diseases. The importance of vitamin D on the regulation of cells of the immune system has gained increased appreciation over the past decade with the discovery of the vitamin D receptor (VDR) and key vitamin D metabolizing enzymes expressed by cells of the immune system. Animal studies, early epidemiologic and clinical studies have supported a potential role for vitamin D in maintaining immune system balance. The hormonal form of vitamin D up-regulates anti-microbial peptides, namely cathelicidin, to enhance clearance of bacteria at various barrier sites and in immune cells. Vitamin D modulates the adaptive immune system by direct effects on T cell activation and on the phenotype and function of antigen-presenting cells (APCs), particularly of DCs. The purpose of this manuscript is to review the molecular and clinical evidence for vitamin D as a modulator of the innate and adaptive immune system.

The nuclear factor of activated Tcells (NFAT) is a transcription factor essential for cytokine production during T-cell activation and is the target of several immunosuppressive drugs. Andrographolide is a diterpenic labdane that possesses anti-inflammatory and immunomodulatory effects. Several studies propose that andrographolide can reduce the immune response through inhibition of the nuclear factor kappa B (NF-κB) and mitogen-activated protein kinases (MAPK) such as extracellular signal regulated kinase 1/2 (ERK1/2) pathways. Moreover, andrographolide reduces IFN-γ and IL-2 production induced by concanavalin A in murine T-cell. Nevertheless, the mechanisms involved in the decrease of cytokine production are unknown. In the present study, we determined that andrographolide reduced IL-2 production in Jurkat cells stimulated with phorbol myristate acetate and ionomycin (PMA/Ionomycin). We then showed that andrographolide reduced NFAT luciferase activity and interfered with its nuclear distribution, with these effects being linked to an increase in c-jun-N-terminal kinase (JNK) phosphorylation. Additionally, reduction of NF-κB activity in Jurkat cells treated with andrographolide was observed. Using Western blotting, we demonstrated that andrographolide decreased ERK1 and ERK5 phosphorylation induced by anti-CD3 or PMA/Ionomycin. Andrographolide did not affect cell viability at concentration of 10 and 50 μM; however, our results suggest that andrographolide increase early apoptosis at 100 μM. We concluded that andrographolide can exert immunomodulatory effects by interfering with NFAT activation and ERK1 and ERK5 phosphorylation in T-cells.

Inappropriate T cell responses cause the four classical types of hypersensitivity immune reactions. All of these can target the kidney and cause distinct forms of glomerulonephritis. CD4 + T cells can mediate glomerular immunopathology by cytokine secretion, by activating effector cells such as macrophages or by inducing autoantibodies or immune-complexes. Cytotoxic CD8 + T cell responses and failure of regulatory T cells may represent two additional types of anti-renal hypersensitivity. T cell activation is critically dependent on dendritic cells (DC), whose role in renal disease appears to be protective, but underlying mechanisms are largely unknown. In this paper, we summarized mechanistic information from rodent models on the roles of DC and T cells in glomerular immunopathology.

A series of novel chemically defined soluble oligomers of the human MHC class II protein HLA-DR1 was constructed to probe the molecular requirements for initiation of T cell activation. MHC dimers, trimers, and tetramers stimulated T cells, as measured by upregulation of the activation markers CD69 and CD25, and by internalization of activated T cell receptor subunits. Monomeric MHC-peptide complexes

We recently characterized Winnie mice carrying a missense mutation in Muc2 , leading to severe endoplasmic reticulum stress in intestinal goblet cells and spontaneous colitis. In this study, we characterized the immune responses due to this intestinal epithelial dysfunction. In Winnie , there was a fourfold increase in activated dendritic cells (DCs; CD11c + major histocompatibility complex (MHC) class II hi ) in the colonic lamina propria accompanied by decreased colonic secretion of an inhibitor of DC activation, thymic stromal lymphopoietin (TSLP). Winnie also displayed a significant increase in mRNA expression of the mucosal T H 17 signature genes Il17a , IL17f , Tgfb , and Ccr6 , particularly in the distal colon.

Ligand binding to cell-surface receptors can activate intracellular signal transduction pathways through a variety of mechanisms . In one common mechanism, typical of seven-transmembrane span receptors that activate G proteins 1 , ligand binding induces a structural change in the receptor that can be sensed by effector proteins within the cell. In another mechanism, binding of a multivalent ligand induces receptor colocalization. This mechanism is typical of receptor tyrosine kinases (RTKs), in which receptor clustering facilitates transphosphorylation by the cytoplasmic kinase domains 2 . Other mechanisms driven by mass action can be envisioned (see following text). In a third mechanism, ligand binding induces rearrangement of a receptor oligomer. One example of this mechanism is the bacterial aspartic acid receptor, in which ligand binding induces a helix reorganization that activates receptor-associated cytoplasmic signaling proteins 3 . For soluble ligands, several examples of each of these mechanisms are known. When both the ligand and receptor are cell-surface proteins, the situation can be complicated; for example, by redistribution of membrane components or interaction of other proteins in the juxtaposed membranes. Cases of both of these situations have been observed in the interaction of T-cell antigen receptors (TCRs) with their ligands -major histocompatibility complex (MHC) proteins that are displayed on the surface of antigen-presenting cells. Recent investigation of this cell-cell signaling system, using soluble oligomeric arrays of MHC proteins as mimics of the antigen-presenting cell, have provided insights into the mechanism of T-cell triggering. However, more work is necessary for us to understand the roles of membrane-proximal signaling events in the overall T-cell activation process.

The role of distinct regions of HLA class I molecules in regulating T-cell activation via the CD3-antigen receptor complex was investigated. Monoclonal antibodies (MoAbs) which recognize monomorphic and polymorphic epitopes on HLA Class I molecules were shown to inhibit T-cell proliferation to OKT3. These MoAbs have differential effects on the synthesis of interleukin-2 (IL-2) and IL-2 receptor expression. Cell cycle analysis demonstrated that these MoAbs function both in inhibiting cell cycle entry (Go-G, shift) and in blocking cell cycle progression (Gi-S shift) of activated T cells. Furthermore, these MoAbs have regulatory effects on the alternate pathway of T-cell activation via the CD2 molecule, T-cell activation induced by PHA, and activation induced by the phorbol ester PMA in conjunction with the calcium ionophore Ionomycin. Thus these MoAbs have different effects depending upon the pathway of T-cell activation. The results indicate that HLA class I molecules are selectively involved in the sequence of intracellular events leading to T-cell activation and proliferation. o 1988 Academic press, Inc.

Signal exchange between cells is a key feature of life from humble monads to human beings. Appropriate communication is of particular importance between cells of multi-cellular organisms. Various basic mechanisms of cellcell communication have evolved during phylogenesis, which were subject to organ, tissue and cell type-specific adaptation. These mechanisms range from long-distance communication via hormones to more and more local processes, e.g. via cytokines, chemokines or neuromodulators/neurotransmitters, and eventually direct physical interactions of molecules anchored at cell surfaces. Accordingly, highly specialized transient or stable cell-cell contact sites have evolved that mediate signaling between cells. With few exceptions (e.g. lipophilic hormones, gases) intercellular communication depends on specific signal detection devices at the cell surface coupled to a signal transduction apparatus that mediates the signal transfer across the cell membrane and activates intracellular effector systems, which generate intracellularly decipherable signals.

Bcl10 is a critical regulator of NF-kappa B activity in T and B cells, coupling antigen receptor signaling to NF-kappa B activation via protein kinase C (PKC). Here we show that PKC or T-cell receptor (TCR)/CD28 signaling results in downregulation of Bcl10 protein levels, thereby attenuating NF-kappa B transcriptional activity. Bcl10 degradation requires an intact caspase recruitment domain and is not observed after stimulation with tumor necrosis factor alpha or lipopolysaccharides. Bcl10 downregulation is not affected by proteasome inhibitors but is accompanied by transient localization to lysosomal vesicles, suggesting involvement of the lysosomal pathway rather than the proteasome. The HECT domain ubiquitin ligases NEDD4 and Itch promote ubiquitination and degradation of Bcl10, thus downmodulating NF-kappa B activation. Since CD3/CD28-induced activation of JNK is not affected by the decline of Bcl10, degradation of Bcl10 selectively terminates IKK/NF-kappa B signaling in respons...

Background: T cell receptor (TCR) engagement leads to formation of signaling microclusters and induction of rapid and dynamic changes in the actin cytoskeleton, although the exact mechanism by which the TCR initiates actin polymerization is incompletely understood. The Vav family of guanine nucleotide exchange factors (GEF) has been implicated in generation of TCR signals and immune synapse formation, however, it is currently not known if Vav's GEF activity is required in T cell activation by the TCR in general, and in actin polymerization downstream of the TCR in particular.

Ginger (Zingiber officinale Rosc) (Family: Zingiberaceae) is a herbaceous perennial, the rhizomes of which are used as a spice. Ginger is a plant which is well known for its medicinal applications. Recently EST-derived SNPs are a free by-product of the currently expanding EST (Expressed Sequence Tag) databases. The development of high-throughput methods for the detection of SNPs (Single Nucleotide Polymorphism) and small indels (insertion/deletion) has led to a revolution in their use as molecular markers. Available (38139) Ginger EST sequences were mined from dbEST of NCBI. CAP3 program was used to assemble EST sequences into contigs. Candidate SNPs and Indel polymorphisms were detected using the perl script AutoSNP version 1.0 which has used 31905 ESTs for detecting SNPs and Indel sites. We found 64026 SNP sites and 7034 indel polymorphisms with frequency of 0.84 SNPs / 100 bp. Among the three tissues from which the EST libraries had been generated, Rhizomes had high frequency of 1.08 SNPs/indels per 100 bp whereas the leaves had lowest frequency of 0.63 per 100 bp and root is showing relative frequency 0.82/100bp. Transitions and transversion ratio is 0.90. In overall detected SNP, transversion is high when compare to transition. These detected SNPs can be used as markers for genetic studies.

The first findings that depression is characterized by cell-mediated immune activation and inflammation were published between 1990. Recently, it was reported thatbased on meta-analysis resultsdepression is an inflammatory disorder because the plasma levels of two cytokines are increased, i.e. interleukin-(IL)-6 and tumor necrosis factor-α (TNFα). The same meta-analysis found that plasma IL-2 and interferon-(IFN)γ levels are not altered in depression, suggesting that there is no T cell activation in that illness. The present paper reviews the body of evidence that depression is accompanied by cell-mediated immune activation. The findings include: increased serum levels of the soluble IL-2 receptor (sIL-2R) and the sCD8 molecule; increased numbers and percentages of T cells bearing T cell activation markers, such as CD2+CD25+, CD3+CD25+, and HLA-DR+; increased stimulated production of IFNγ; higher neopterin and sTNFR-1 or sTNFR-2 levels; induction of indoleamine 2,3-dioxygenase (IDO) with lowered levels of plasma tryptophan and increased levels of tryptophan catabolites along the IDO pathway (TRYCATs); and glucocorticoid resistance in immune cells. Interferon-α (IFNα)-based immunotherapy shows that baseline and IFNα-induced activation of T cells, IDO activity and TRYCAT formation are related to the development of IFNα-induced depressive symptoms. Animal models of depression show that a cell-mediated immune response is related to the development of depression-like behavior. Antidepressants and mood stabilizers suppress different aspects of cell-mediated immunity and rather specifically target IFNγ production. This review shows that inflammation and cell-mediated immune activation are key factors in depression.

A major question in immunology is how DC can display limited amounts of individual peptide-MHC complexes and still induce cross-linking of T-cell receptors to initiate cellular responses. One suggested mechanism is that MHC exists at the cell surface in high avidity multimers, and tetraspanin proteins, known to laterally associate with both MHC classes I and II, promote MHC multimerisation. To validate this theory, we tested the ability of DC deficient in either one of two typical tetraspanin molecules: CD37 or CD151 to present peptide to Ag-specific T cells. Surprisingly, although they exhibited no developmental or maturation defects, DC lacking either CD37 or CD151 expression were hyperstimulatory to T cells. We demonstrate that CD37 and CD151 control DC-mediated T-cell activation by two different mechanisms: CD151 regulates co-stimulation whereas CD37 regulates peptide/MHC presentation. The implications of these results on the model suggesting that tetraspanins promote MHC multimerisation are discussed.

Mannose-terminating glycans have been identified as pathogen-associated molecular patterns that are essential for internalization of microbes by antigen-presenting cells, leading to presentation. Here, we have investigated the importance of exposed mannosylation on an immunogenic therapeutic self-protein, procoagulant human factor VIII (FVIII). Administration of therapeutic FVIII to hemophilia A patients induces inhibitory anti-FVIII antibodies in up to 30% of the cases. We demonstrate that entry of FVIII into human dendritic cells (DC) leading to T cell activation, is mediated by mannose-terminating glycans on FVIII. Further, we identified macrophage mannose receptor (CD206) as a candidate endocytic receptor for FVIII on DC. Saturation of mannose receptors on DC with mannan, and enzymatic removal of mannosylated glycans from FVIII lead to reduced T cell activation. The interaction between FVIII and CD206 was blocked by VWF, suggesting that, under physiological conditions, the intrinsic mannose-dependent immunogenicity of FVIII is quenched by endogenous immunochaperones. These data provide a link between the mannosylation of therapeutic self-proteins and their iatrogenic immunogenicity. Such a link would be of special relevance in the context of replacement therapy where mechanisms of central and peripheral tolerance have not been established during ontogeny because of the absence of the antigen. dendritic cells ͉ factor VIII ͉ hemophilia ͉ mannose receptor ͉ mannosylated glycans

Although there has been extensive analysis on the capacity of MHC-peptide tetramers to bind antigen-specific TCR, there have been comparatively few studies regarding the role of the CD4 and CD8 co-receptors in binding and activation by these multimeric molecules. Here, we start from the observation that different antibodies against human CD8 exert opposite effects on MHC-peptide tetramer binding to the TCR: tetramer staining was enhanced by OKT8 antibody, while it was blocked with SK1 antibody. We used these different anti-CD8 antibodies to modulate CD8 function during tetramer staining of Melan-A/MART1-specific CTL clones. We show that CD8 action could be variably modulated during all the phases of interaction, indicating that CD8 participates in both the initial association of the TCR with MHC-peptide tetramers and the stability of this interaction. While the blocking effect of anti-CD8 antibodies was mostly exerted during the initial binding of the TCR with MHC-peptide tetramers, the enhancing effect was exerted by augmenting the duration of this interaction. Blocking anti-CD8 antibodies were also capable of preventing tetramer-mediated T cell activation. The possibility of variably affecting MHC-peptide tetramer binding and T cell activation using anti-CD8 antibodies confirms the critical role exerted by the CD8 co-receptor in this interaction and supports the notion that TCR engagement by MHC-peptide ligands typically involves CD8.

(cross-priming) but also, and importantly, T-cell tolerance contingent upon the activation/ maturation status of DC. Artificial enhancement of tumor antigen cross-presentation and control of the immune-promoting status of the antigen-presenting DC will have important therapeutic implications in the near future.

T cell activation is controlled by the coordination of stimulatory and negative regulatory signals which are not completely defined. In this study we tested for a possible direct effect of CD14 on the regulation of T cell activation and function. We show that soluble CD14 (sCD14) induces inhibition of antigen-mediated peripheral blood mononuclear cells (PBMC) proliferation and anti-CD3-mediated proliferation of CD4 + CD8 − , CD4 − CD8 + and CD4 + CD8 + T cell clones. This effect is not due to cell death, but results from a marked inhibition of IL-2 production. Proliferation of T cell clones due to exogenous IL-2 is not affected by sCD14. We also found that sCD14 inhibits production of another Th1-like cytokine, IFN-+ and a Th2-like cytokine, IL-4. Importantly, sCD14 induces a progressive accumulation of the inhibitory protein I O B- § . We show that sCD14 binds to activated T cells. Following cell activation, biotinylated sCD14 stains CD3 + PBMC, as well as human T cell clones with varying intensity. The binding is saturable, can be inhibited by excess of unlabeled sCD14 and, following binding, sCD14 is internalized. Collectively, these findings reveal a previously unrecognized function of sCD14, namely its capacity to negatively regulate T lymphocyte activation and function by interacting directly with activated T cells.

T cell receptor (TCR) activation and signaling precede immunological synapse formation and are sustained for hours after initiation. However, the precise physical sites of the initial and sustained TCR signaling are not definitively known. We report here that T cell activation was initiated and sustained in TCR-containing microclusters generated at the initial contact sites and the periphery of the mature immunological synapse. Microclusters containing TCRs, the tyrosine kinase Zap70 and the adaptor molecule SLP-76 were continuously generated at the periphery. TCR microclusters migrated toward the central supramolecular cluster, whereas Zap70 and SLP-76 dissociated from these microclusters before the microclusters coalesced with the TCR-rich central supramolecular cluster. Tyrosine phosphorylation and calcium influx were induced as microclusters formed at the initial contact sites. Inhibition of signaling prevented recruitment of Zap70 into the microclusters. These results indicated that TCRrich microclusters initiate and sustain TCR signaling.

Abbreviations: AAV -ANCA-associated vasculitides, ANCAanti-neutrophil cytoplasmic autoantibodies, DTH -delayed-type hypersensitivity, IFN -interferon, IL -interleukin, MPA -microscopic polyangiitis, MPO -myeloperoxidase, PR3 -proteinase 3, RLV -renal-limited vasculitis, TCR -T-cell receptor, TEM -effector memory T cells, WG -Wegener's granulomatosis.

Tribodies are multifunctional recombinant antibody derivatives, which utilize the natural in vivo heterodimerization of the heavy chain (Fd fragment) and light chain (L) of a Fab fragment, to form a scaffold, upon which additional functions can be incorporated, such as additional binders -e.g. scFv binding domains.

MuS110 is a BiTE antibody bispeciWc for murine EpCAM (CD326) and murine CD3. A recent study has shown that muS110 has signiWcant anti tumor activity at well-tolerated doses as low as 5 g/kg in orthotopic breast and lung cancer models (Amann et al. in Cancer Res 68:143-151, 2008). Here, we have explored the safety pro-Wle of muS110 at higher doses. Escalation to 50 g/kg muS110 caused in mice transient loss of body weight, and transient piloerection, hypomotility, hypothermia and diarrhoea. These clinical signs coincided with serum peaks of TNF-, IL-6, IL-2, IFN-and IL-4, and an increase of surface markers for T cell activation. Because activation of T cells in response to BiTE antibodies is typically dependent on target cells, we analyzed mouse blood for the presence of EpCAM + cells. Various mouse strains presented with a subpopulation of 2-3% EpCAM + blood cells, mostly B and T lymphocytes, which was not detected in human blood samples. In vitro experiments in which the number of EpCAM + cells in blood samples was either reduced or increased suggested that both T cell activation and cytokine release in response to muS110 was dependent on the number of target-expressing cells. In support for a role of EpCAM + lymphocytes in the observed side eVects, reduction of EpCAM + blood cells in mice via a low-dose pre treatment with muS110 dramatically increased the tolerability of animals up to at least 500 g/kg of the BiTE antibody. This high tolerability to muS110 occurred in the presence of non-compromised T cells. No damage to EpCAM + epithelial tissues was evident from histopathological examination of animals daily injected with 100 g/kg muS110 for 28 days. In summary, these observations suggest that side eVects of muS110 in mice were largely caused by an acute T cell activation that was triggered by a subpopulation of EpCAM + lymphocytes. Because humans have extremely low numbers of EpCAM + cells in blood, this toxicity of an EpCAM-speciWc BiTE may be speciWc for mice.

T Cells: + 2 δ 9V γ Reactive V tuberculosis-Mycobacterium Macrophages to Phosphoantigen Presentation by http://iai.asm.org/content/70/8/4019 Updated information and services can be found at: These include: REFERENCES http://iai.asm.org/content/70/8/4019#ref-list-1 at: This article cites 54 articles, 29 of which can be accessed free CONTENT ALERTS more» articles cite this article), Receive: RSS Feeds, eTOCs, free email alerts (when new http://journals.asm.org/site/misc/reprints.xhtml Information about commercial reprint orders: http://journals.asm.org/site/subscriptions/ To subscribe to to another ASM Journal go to: on April 23, 2014 by guest http://iai.asm.org/ Downloaded from on April 23, 2014 by guest V␥9V␦2 ؉ T

T cell antigen recognition is accompanied by cytoskeletal polarization towards the APC and large-scale redistribution of cell surface molecules into 'supramolecular activation ( ) clusters' SMACs , forming an organized contact interface ( ) termed the 'immunological synapse' IS . Molecules are arranged in the IS in a micrometer scale bull's eye pattern ( with a central accumulation of TCR r peptide᎐MHC the ) cSMAC surrounded by a peripheral ring of adhesion ( ) molecules the pSMAC

Two monoclonal antibodies (mAb) recognizing different CD2 epitopes each inhibited anti-CD3-induced proliferation and anti-CD3-induced increase in surface CD2 expression. The magnitude of inhibition by either anti-CD2 mAb was dependent upon which anti-CD3 mAb was used as the stimulus, being more pronounced when the anti-CD3 mAb 454 was used as the stimulus than when either anti-CD3 mAb 147 or 446 was the stimulus. The effects of neuraminidase-treated sheep erythrocytes (which bind to CD2) were also more pronounced on mAb 454-induced proliferation than on mAb 147- or 446-induced proliferation. Furthermore, the effects of preincubation with anti-CD2 mAb depended upon the responder status of the donor to IgG1 anti-CD3 mAb. Preincubation of high-responder cells with anti-CD2 mAb had little effect on subsequent IgG1 anti-CD3-induced proliferation. In contrast, preincubation of low-responder cells with anti-CD2 mAb usually augmented the otherwise small proliferative response to IgG1 anti-C...

IgA deficiency (IgAD) and common variable immunodeficiency (CVID) often co-occur in families, associating with chronic inflammatory diseases such as celiac disease (CD). ICOS (inducible co-stimulator) and CTLA4 (cytotoxic T-lymphocyteassociated protein-4) may be important in both disorders, as ICOS is necessary for Ig class-switching and CTLA4 negatively regulates T-cell activation. Linkage and association of CD with CTLA4-ICOS is well documented, we thus aimed to further pinpoint CD susceptibility by haplotype-tagging analysis. We genotyped 663 CD families from Finland and Hungary, 575 additional CD patients from Finland, Hungary and Italy; 275 Swedish and Finnish IgAD individuals and 87 CVID individuals for 14-18 genetic markers in CTLA4-ICOS. Association was found between CTLA4-ICOS and both IgAD (P ¼ 0.0015) and CVID (P ¼ 0.0064). We confirmed linkage of CTLA4-ICOS with CD (LOD 2.38, P ¼ 0.0005) and found association of CTLA4-ICOS with CD (P ¼ 0.0009). Meta-analysis of the IgAD, CVID and CD materials revealed intergenic association (P ¼ 0.0005). Disease-associated markers were associated with lower ICOS and higher CTLA4 expression, indicating that the risk haplotypes contain functional variants. In summary, we identified a novel shared risk locus for IgAD, CVID and CD, the first report of association between CTLA4-ICOS and IgAD. Association between CD and CTLA4-ICOS was also confirmed in a large European data set.

Dendritic cells (DC) are professional antigen-presenting cells that are specialized in the uptake of antigens and their transport from peripheral tissues to the lymphoid organs. Because of their capacity to stimulate naïve T cells, DC have a central role in the initiation of primary immune responses and are considered promising tools and targets for immunotherapy. Investigating the signals that regulate the function of DC, the sentinels of the immune system, is also critical to understanding the role of DC in the regulation of immune response. We have assessed the effects of natural antibodies in the form of therapeutic intravenous immunoglobulins (IVIg) on the regulation of the function of DC. We have observed that DC are the primary targets for the immunosuppressive effects of IVIg on T-cell activation. This review focuses on the effects of IVIg on DC development and function.

T cells are activated by binding of the T cell receptor (TCR) to a peptide-major histocompatibility complex (MHC) complex (pMHC) expressed on the surface of antigen presenting cells. Various models have predicted that activation is limited to a narrow window of affinities (or dissociation rates) for the TCR–pMHC interaction and that above or below this window, T cells will fail to undergo activation. However, to date there have not been TCRs with sufficiently high affinities in order to test this hypothesis. In this report we examined the activity of a CD8-negative T cell line transfected with a high affinity mutant TCR (KD = 10 nM) derived from cytotoxic T lymphocyte clone 2C by in vitro engineering. The results show that despite a 300-fold higher affinity and a 45-fold longer off-rate compared with the wild-type TCR, T cells that expressed the mutant TCRs were activated by peptide. In fact, activation could be detected at significantly lower peptide concentrations than with T cell...

In its simplest form, effective T cell-mediated immunity emanates from the expansion of specific T cells activated in response to antigen. In establishing and maintaining the peripheral T-cell pool, the thymus plays a critical role. It does so by providing a microenvironment within which T-cell precursors proliferate, differentiate and undergo selection processes to create a fully functional population of major histocompatibility complex restricted, selftolerant T cells. The control of the thymic function involves intrathymic, as well as sympathetic nervous and endocrine system signalling. In addition to postganglionic noradrenergic fibres, both thymic lymphoid and nonlymphoid cells, including epithelial cells and macrophages, have been demonstrated to express tyrosine hydroxylase (TH), and suggested to form a local non-neural catecholaminergic cell network. A higher level of noradrenaline has been found in male than in female rat thymi, and a role of gonadal hormones in providing this dimorphism has been demonstrated. In addition, thymic lymphoid and non-lymphoid cells, including those expressing TH, have been found to bear βand α 1 -adrenoceptors (ARs) and a role of gonadal hormones in regulation of, at least, β-AR density and signalling has been suggested. These findings have also entailed conclusion that catecholamines (CAs) influence T-cell development, not only via neurocrine/endocrine, but also via autocrine/paracrine action. Generally, CAs have been shown to exert an inhibitory influence on thymopoiesis. Role of α 1 -and β-AR-mediated mechanisms in maintaining thymic homeostasis and in fine tuning of both conventional and regulatory T-cell development is discussed in the manuscript.