Search

Macrophages are crucial effectors of innate immunity against the pathogenic bacterium Listeria monocytogenes. The pro-inflammatory cytokine tumour necrosis factor-α (TNF) has been shown to be crucial for resistance to L. monocytogenes and mice deficient in TNF signalling succumb quickly after infection. However, the mechanisms underlying TNF-mediated defence against L. monocytogenes infection have not been completely elucidated. Here, we demonstrate that TNF concurrently functions to support a pro-inflammatory M1 phenotype while actively blocking macrophage polarization to the M2 phenotype. Compared to WT mice, peritoneal macrophages in TNF-deficient mice inoculated with L. monocytogenes respond with M2 polarization by upregulating Arg1. Consistently, TNF blockade in vitro resulted in M2 polarization in peritoneal macrophages during L. monocytogenes infection. Additionally, TNF promotes the transition from M2 to M1 polarization in peritoneal macrophages. Further investigation of peritoneal macrophage polarization suggested the NF-κB pathway is involved in the TNF-dependent M2 to M1 shift. Conversely, treatment of peritoneal macrophage with a PPARγ agonist blunted the expression of M1 genes induced by TNF and reduced NF-κB signalling pathway activation. Competing signalling mechanisms therefore play an essential role in the ability of peritoneal macrophage to resolve L. monocytogenes infections with TNF playing an essential role in driving M1 polarization.Abbreviations: LPM: large peritoneal macrophage; SPM: small peritoneal macrophage; LLO: listeriolysin O; iNOS: inducible nitric oxide synthase; DCs: dendritic cells.

T helper cells play an important role in the aetiology of Multiple Sclerosis (MS). Vitamin D has an anti-inflammatory effect on T helper cells and can affect onset and pathogenesis of MS. Two genes of the metabolic Vitamin D pathway expressed by activated T helper (Th) cells have been identified as MS risk genes by genome-wide association studies, CYP27B1 (25(OH)D

Traditionally, chemokine immunobiology has focused on chemotaxis and the positioning of cells at sites of inflammation and within lymphoid organs. More recently, however, regulation of intricate immune responses has emerged as a function attributed to chemokines and their receptors. One such pair, CCR6 and its chemokine ligand CCL20, has been receiving interest for its potential role in the coordination and regulation of humoral immune responses and in particular, memory responses, at the cellular level. B cells up-regulate CCR6 after activation in secondary lymphoid organs; however, its function is still unclear. In an important insight, the CCR6-CCL20 chemokine axis has been implicated in the regulation of effective humoral responses - disruption of this pair led to an increased number of ineffective T-B cell conjugates and poorer quality antibodies. Interestingly, follicular helper T cells and their precursors also up-regulate CCR6; though, again, the precise purpose of this is yet to be discovered. The chemokine axis in relation to secondary lymphoid organ (SLO) structures will be briefly reviewed as well. With the implication of CCR6 and CCL20 in the pathogenesis of autoantibody-driven autoimmune diseases such as systemic lupus erythematosus, understanding the intricacies of this chemokine pair would be conducive to the development of appropriate, targeted therapeutic strategies.

The chronic inflammatory autoimmune disease rheumatoid arthritis (RA) is characterized by an infiltration of activated proinflammatory immune cells into the joint that is accompanied by an overproduction of various mediators, leading to destruction of cartilage and bone erosion. Angiotensin II type 2 receptor (AT2R) is involved in antioxidative, anti-inflammatory, and antifibrotic responses. Synovial macrophages (SMs) are a type of tissue macrophages that are derived from bone marrow cells. SMs plays a central role in synovial regional immunization, which is significantly increased in both collagen-induced mice with arthritis mice and RA patients. AT2R activation caused a reversal of the polarization of SMs in the joint from the proinflammatory M1 SM to the tolerogenic, benign M2 SM. In consequence, this switch resulted in an attenuated form of the joint pathology in a rat model of collagen-induced arthritis. These results were mechanistically linked to the observation that GRK2 was translocated into cytoplasm, and ERK1/2 and NF-κB activation were inhibited. These findings open the way to a new therapeutic approach using an activation of AT2R to subvert joint inflammation in RA.

The CC chemokine receptor 6 (CCR6) and its sole chemokine ligand, CCL20, are an intriguing pair that have been implicated in a growing number of inflammatory, autoimmune and malignant disease processes. Recent observations have also highlighted this chemokine axis in the regulation of humoral immune responses. Through this review article, we explore the emerging links of CCR6-CCL20 with an archetypal autoimmune disease of humoral dysregulation: systemic lupus erythematosus (SLE). CCR6 is expressed prominently on several immune cells involved in the pathogenesis of SLE, such as dendritic cells and T-helper 17 cells. CCR6's expression is correlated with disease activity and serological markers of disease severity, suggesting a possible role in disease pathogenesis. However, there are numerous holes in our understanding of the functions of CCR6 and CCL20, and future studies are required to determine if there are any diagnostic, prognostic or monitoring roles for these important molecules.

The angiotensin II type 1 receptor (AT1R) antagonist losartan has been confirmed to have a moderate anti-inflammatory effect in vitro and in vivo. However, how it affects immune cells in Rheumatoid Arthritis (RA) is still unknown. We found that in human synovial tissues, AT1R is significantly expressed on T cells and B cells. Treatment with losartan (15 mg/kg) alone and in combination with a low dose of methotrexate (MTX 0.25 mg/kg/3 days) significantly suppressed the progression of CIA. Secondary paw swelling, joint destruction and the presence of pro-inflammatory cytokines (TNF-α and IFN-γ) in the serum were alleviated after treatment. The therapeutic effects of losartan were based on reduced T-cell and B-cell activation, specifically by decreased cell vitality and pro-inflammatory cytokine production. In addition, losartan combined with a low dose of MTX achieved a similar therapeutic effect, while protecting liver and kidney from MTX damage. Mechanistically, losartan inhibits the production of pro-inflammatory mediators, reduces the phosphorylation of p38, ERK, and p65, p50 nuclear transposition in T cells and B cells. Phosphorylation of JNK is not affected by losartan in the CIA rat model. losartan can be used as an effective RA treatment, which exhibits anti-arthritic effects potentially through down-regulating the phosphorylation of p38, ERK and signaling through NF-κB. While achieving similar anti-rheumatic effects, a combination therapy of losartan with a low dose of MTX, can protect from liver and renal damage caused by giving a high dose of MTX.

The confirmation of developmental differences between tissue macrophages and peripheral monocytes has changed our view of the functions and dynamics of these two important components of the innate immune system. It has been demonstrated conclusively that homeostasis of tissue resident macrophages is maintained by a low proliferative turn over. During an inflammatory response, bone marrow derived monocytes enter the tissue in large numbers and take part in the defense against the pathogens. After the destruction of invading pathogens, these cells disappear and tissue resident macrophages can be detected again. This new appreciation of the innate immune response has not only answered many outstanding questions regarding the role of the different myeloid cell types in inflammation, but also opened up new areas of research relating to the tissue- and pathogen-specific fate of the inflammatory macrophages or dendritic cells (DCs), and the transfer of this knowledge from mouse models to the human immune system. Nevertheless, there is still confusion in infection models, and especially in studies of human infections, as to what extent these recent observations and findings influence previous interpretations of data. This review will focus on insights from mouse models, summarize the literature on the ontogeny of macrophages and monocytes, explain the role of frequently used monocyte markers and effector molecules, and finally, discuss the role of inflammatory monocytes/macrophages/DCs in two experimental parasitic diseases.

Improved understanding of the molecular mechanisms underlying dysregulated inflammatory responses in severe infection and septic shock is urgently needed to improve patient management and identify new therapeutic opportunities. The WNT signaling pathway has been implicated as a novel constituent of the immune response to infection, but its contribution to the host response in septic shock is unknown. Although individual WNT proteins have been ascribed pro- or anti-inflammatory functions, their concerted contributions to inflammation in vivo remain to be clearly defined. Here we report differential expression of multiple WNT ligands in whole blood of patients with septic shock and reveal significant correlations with inflammatory cytokines. Systemic challenge of mice with lipopolysaccharide (LPS) similarly elicited differential expression of multiple WNT ligands with correlations between WNT and cytokine expression that partially overlap with the findings in human blood. Molecular regulators of WNT expression during microbial encounter in vivo are largely unexplored. Analyses in gene-deficient mice revealed differential contributions of Toll-like receptor signaling adaptors, a positive role for tumor necrosis factor, but a negative regulatory role for interleukin (IL)-12/23p40 in the LPS-induced expression of Wnt5b, Wnt10a, Wnt10b, and Wnt11. Pharmacologic targeting of bottlenecks of the WNT network, WNT acylation and β-catenin activity, diminished IL-6, tumor necrosis factor, and IL-12/23p40 in serum of LPS-challenged mice and cultured splenocytes, whereas IL-10 production remained largely unaffected. Taken together, our data support the conclusion that the concerted action of WNT proteins during severe infection and septic shock promotes inflammation, and that this is, at least in part, mediated by WNT/β-catenin signaling.

Excessive tumor necrosis factor (TNF) is known to cause significant pathology. Paradoxically, deficiency in TNF (TNF(−/−)) also caused substantial pathology during respiratory ectromelia virus (ECTV) infection, a surrogate model for smallpox. TNF(−/−) mice succumbed to fulminant disease whereas wild-type mice, and those engineered to express only transmembrane TNF (mTNF), fully recovered. TNF deficiency did not affect viral load or leukocyte recruitment but caused severe lung pathology and excessive production of the cytokines interleukin (IL)-6, IL-10, transforming growth factor beta (TGF-β), and interferon gamma (IFN-γ). Short-term blockade of these cytokines significantly reduced lung pathology in TNF(−/−) mice concomitant with induction of protein inhibitor of activated STAT3 (PIAS3) and/or suppressor of cytokine signaling 3 (SOCS3), factors that inhibit STAT3 activation. Consequently, inhibition of STAT3 activation with an inhibitor reduced lung pathology. Long-term neutralization of IL-6 or TGF-β protected TNF(−/−) mice from an otherwise lethal infection. Thus, mTNF alone is necessary and sufficient to regulate lung inflammation but it has no direct antiviral activity against ECTV. The data indicate that targeting specific cytokines or cytokine-signaling pathways to reduce or ameliorate lung inflammation during respiratory viral infections is possible but that the timing and duration of the interventive measure are critical.

Primary Sjogren's syndrome (pSS) is an autoimmune disease of unresolved aetiology that affects the exocrine glands. Clinical symptoms frequently also involve skin, liver, kidney and neurovascular components. The pathogenesis of pSS is still unclear but B cell hyperactivity has been identified as a hallmark of pSS. Currently, a curative therapeutic agent is lacking. In this study, we explored whether paeoniflorin-6′-O-benzene (CP-25) exerted therapeutic effects through regulating B lymphocyte migration via CXCR5-GRK2-MAPK mediated signaling pathways in a mouse model of antigen-induced, experimental Sjogren's syndrome (ESS). We found that CP-25 increased the salivary flow and alleviated the histopathology of ESS. Furthermore, CP-25 reduced the viability of B lymphocyte and limited the target organs index. In the peripheral blood and salivary gland of ESS mice, CP-25 down-regulated the proportion of total B cells, CXCR5+ B cells and PDCA1 + CD19- and limited the presence of phosphorylated (p-) p38 and ERK (p-ERK). Besides, CP-25 increased the percentage of memory B cells in the peripheral blood and reduced it in salivary gland. Furthermore, in vitro, CP-25 down-regulated p-p38, p-ERK, CXCR5 and membrane GRK2, and increased cytoplasm GRK2 in Maver-1 cells, a mantle cell lymphoma cell line, causing a lower migration ability of Maver-1 cells. Thus, we define CP-25 as a novel compound that is a potent therapeutic agent for pSS which modulates B lymphocyte subsets and impacts the migration of B lymphocytes through regulating the CXCR5-GRK2-ERK/p38 signaling pathway.

Macrophages have emerged as a key component of the innate immune system that emigrates to peripheral tissues during gestation and in the adult organism. Their complex pathway to maturity, their unique plasticity and their various roles as effector and regulatory cells during an immune response have been the focus of intense research. A class of surface molecules, the G-Protein coupled receptors (GPCRs) play important roles in many immune processes. They have drawn attention in regard to these functions and the potential for therapeutic targets that can modulate the response of immune cells in pathologies such as diabetes, atherosclerosis, and chronic inflammatory diseases. Of the more than 800 GPCRs identified, ~100 are currently targeted with drugs which have had their activity investigated in vivo. Macrophages express a number of GPCRs which have central roles during cell differentiation and in the regulation of their functions. While some macrophage GPCRs such as chemokine receptors have been studied in great detail, the roles of other receptors of this large family are still not well understood. This review summarizes new insights into macrophage biology, differences of human, and mouse macrophages and gives details of some of the GPCRs expressed by this cell type.

Interleukin-34 (IL-34) is a recently discovered cytokine that promotes tissue macrophage maturation and differentiation. We previously found that 1α,25-Dihydroxyvitamin D3 up-regulated IL-34 expression in SH-SY5Y neural cells. However, whether microRNA regulates IL-34 expression is not completely clear. By using on-line TargetScan and MiRanda software, we found that there was only one conserved microRNA-31 (miR-31) binding site in the 3' untranslated region (3'UTR) of IL-34 mRNA. Intriguingly, using qPCR we demonstrated that miR-31 levels were negatively correlated to IL-34 mRNA levels in different cell lines. By examining the effect of miR-31 on IL-34 3' UTR reporter luciferase activity and on IL-34 mRNA and argonaute RISC catalytic component 2 (AGO2) binding, it was found that miR-31 bound directly to IL-34 3'UTR and regulated the post-transcriptional expression of IL-34 in MGC-803 cells. Moreover, a miR-31 mimic significantly reduced IL-34 expression levels while a miR-31 inhibitor up-regulated IL-34 expression in KYSE-45 and HT-29 cells. Taken together, these results show that miR-31 negatively regulates IL-34 expression by directly binding to the IL-34 3' UTR in vitro.

A super-enhancer (SE) is a cluster of enhancers with a relatively high density of particular chromatin features. SEs typically regulate key genes that can determine cell identity and differentiation. Identifying SEs and their effects may be critical in predicting key regulatory genes, such as master transcription factor genes or oncogenes. Signal inducible SEs are dense stretches of signal terminal transcription factor (TF) binding regions, and may modulate the interaction between environmental factors (e.g., Vitamin D) and genetic factors (i.e., risk variants) in complex diseases such as multiple sclerosis (MS). As a complex autoimmune disease, the etiology and progression of MS, including the interaction between Vitamin D and MS risk variants, is still unclear and can be explored from the aspect of signal SEs. Vitamin D [with its active form: 1,25(OH)2D3], is an environmental risk factor for MS. It binds the Vitamin D receptor (VDR) and regulates gene expression. This study explores the association between VDR super-enhancers (VSEs) and MS risk variants. Firstly, we reanalyse public ChIP-seq and RNA-seq data to classify VSEs into three categories according to their combinations of persistent and secondary VDR binding. Secondly, we indicate the genes with VSE regions that are near MS risk variants. Furthermore, we find that MS risk variants are enriched in VSE regions, and we indicate some genes with a VSE overlapping MS risk variant for further exploration. We also find two clusters of genes from the set of genes showing correlation of expression patterns with the MS risk gene ZMIZ1 that appear to be regulated by VSEs in THP-1 cells. It is the first time that VSEs have been analyzed, and we directly connect the genetic risk factors for MS risk with Vitamin D based on VSEs.

Introduction: Macrophage phagocytosis of pathogens and tumour cells is an important early event in protection against infectious disease and cancer. As tumour necrosis factor α (TNF) is an important cytokine in macrophage activation, we investigated the involvement of TNF in macrophage phagocytosis of tumour cells. Methods: We used Devil Facial Tumour Disease (DFTD) cancer cells as the target tumour cells. The Tasmanian devil (Sarcophilus harrisii) population is threatened by the transmissible DFTD. Using DFTD cells provided the opportunity to determine if these cells can be phagocytosed and investigate requirement for TNF. As effector cells, bone marrow derived macrophages (BMDMs), generated from C57BL/6 wild type (B6.WT) and C57BL/6 TNF-/- (B6.TNF-/-) mice were used. Phagocytosis of DFTD cells was investigated by confocal microscopy and flow cytometry. Results: DFTD cells were consistently phagocytosed by B6.WT and B6.TNF-/- BMDMs with similar efficiency in vitro. Consequently the DFTD cells are not resistant to phagocytosis. Following activation by exposure to IFNγ and LPS or LPS alone, B6.TNF-/- BMDMs had higher phagocytic efficiency and lower nitric oxide (NO) production compared to wild-type controls. In addition, NO seems to be unlikely to be the involved in phagocytosis efficiency in IFNγ and LPS activated B6.TNF-/- macrophages and consequences thereof. Conclusion: Our results indicate that TNF is not required for IFNγ and LPS or LPS alone activation of macrophage phagocytosis. TNF may negatively regulate macrophage phagocytosis of tumour cells.

If the body's immune system is disordered and begins to attack "self" and therefore, its own tissues this is considered to be an autoimmune pathology. The specific mechanisms vary between the different diseases and have not always been elucidated but chronic, non-resolving inflammation is a common theme in the pathogenesis of autoimmune diseases. Interestingly, it has been shown that development and occurrence of various inflammatory responses are closely correlated to endoplasmic reticulum stress. Therefore, this review discusses the current progress of research about the relationship between autoimmune diseases and endoplasmic reticulum stress, specifically the unfolded protein response (UPR).

The interaction between T cell and dendritic cells (DCs) that leads to T cell activation affects the progression of the immune response including autoimmune diseases. Antigen presentation on immune cell surface, formation of an immunological synapse (IS), and specific identification of complex by T cells including two activating signals are necessary steps that lead to T cell activation. The formation of stimulatory IS involves the inclusion of costimulatory molecules, such as ICAM-1/LFA-1 and CD28/B7-1, and so on. Some fusion proteins and monoclonal antibodies targeting costimulatory molecules have been developed and approved to treat autoimmune diseases, including rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), multiple sclerosis (MS), type I diabetes (T1D), inflammatory bowel disease (IBD), and psoriasis. These biological agents, including CTLA-4- and LFA-3-Ig, anti-CD3 monoclonal antibody, could prevent the successful engagement of DCs by T cell with significant efficacy and safety profile. In this article, we reviewed the molecular mechanisms of T cell activation during the interaction between T cells and DCs, and summarized some biological agents that target costimulatory molecules involved in the regulation of T cell activation.

Fibroblast-like synoviocytes (FLS) and macrophage-like synoviocytes (MLS) are the two main cellular components of the synovium. It has been widely reported that FLS and MLS play essential roles in the joint pathology of rheumatoid arthritis (RA). Although various studies have analyzed both human and animal tissues and have shown that both cell types are involved in different stages of RA, ontology, and specific functions of both cell populations and their interactions are not well understood. In this review, we will summarize recent research on FLS and MLS in RA and focus on the development and function of two predominant synovial cell types. In addition, we will discuss the communication between FLS or MLS and highlight potential treatments for RA that involve synoviocytes.

A low level of inflammation is an integral part of the balance between the immune system and the microbiota in the high antigen environment of the gastrointestinal tract and maintains homeostasis. A failure of this balance can lead to chronic intestinal inflammation and increase the chances to develop colorectal cancer significantly. The underlying mechanisms that link inflammation and carcinogenesis are not clear but the molecular platforms of the inflammasomes have been implicated. Inflammasomes are molecule complexes that are assembled in response to microbial components or cellular danger signals and facilitate the production of bioactive pro-inflammatory cytokines. One inflammasome in particular, NLRP3, has been analysed extensively in its contribution to colitis and has been shown to be associated with the development of colitis-associated colorectal cancer. This review will summarise the role of NLRP3 in intestinal inflammation, discuss some of the triggers of inflammation in the gastrointestinal tract such as diet and introduce some opportunities to use this inflammasome as therapeutic target for the treatment of colitis and colitis-associated colorectal cancer.

Vitamin D has a plethora of functions that are important for the maintenance of general health and in particular, the functional integrity of the immune system, such as promoting an anti-inflammatory cytokine profile and reducing the Treg/Th17 ratio. Multiple sclerosis (MS) is a chronic, inflammatory, and neurodegenerative central nervous system (CNS) disorder of probable autoimmune origin. MS is characterized by recurring or progressive demyelination and degeneration of the CNS due in part to a misguided immune response to as yet undefined (CNS) antigens, potentially including myelin basic protein and proteolipid protein. MS has also been shown to be associated significantly with environmental factors such as the lack of vitamin D. The role of vitamin D in the pathogenesis and progression of MS is complex. Recent genetic studies have shown that various common MS-associated risk-single-nucleotide polymorphisms (SNPs) are located within or in the vicinity of genes associated with the complex metabolism of vitamin D. The functional aspects of these genetic associations may be explained either by a direct SNP-associated loss- or gain-of-function in a vitamin D-associated gene or due to a change in the regulation of gene expression in certain immune cell types. The development of new genetic tools using next-generation sequencing: e.g., chromatin immunoprecipitation sequencing (ChIP-seq) and the accompanying rapid progress of epigenomics has made it possible to recognize that the association between vitamin D and MS could be based on the extensive and characteristic genomic binding of the vitamin D receptor (VDR). Therefore, it is important to analyze comprehensively the spatiotemporal VDR binding patterns that have been identified using ChIP-seq in multiple immune cell types to reveal an integral profile of genomic VDR interaction. In summary, the aim of this review is to connect genomic effects vitamin D has on immune cells with MS and thus, to contribute to a better understanding of the influence of vitamin D on the etiology and the pathogenesis of this complex autoimmune disease.

Chemokines and their receptors are vital for the trafficking of immune cells. In an orchestrated fashion, up- and down-regulation of chemokines and their receptors contribute to both immune system homeostasis as well as inflammation. The CC chemokine, CCL20 and its cognate receptor, CCR6, are described as one of the few chemokine-receptor pairs that show exclusivity. In our review, we analyze observations which indicate that CCR6 does not have CCL20 as an exclusive ligand as once appreciated. For example, attempts to study the pair, utilizing mainly CCR6-deficient mice, are confounded by a family of non-chemokine ligands known as β-defensins that can bind to CCR6 and potentially can activate the cell. Therefore, a review of the activities of other potential binding partners of CCR6 is essential for interpretation of the current literature on this matter and for an understanding of their involvement in basic immunology and pathology.

The absence of tumor necrosis factor (TNF) causes lethal infection by Leishmania major in normally resistant C57BL/6J (B6.WT) mice. The underlying pathogenic mechanism of this fatal disease has so far remained elusive. We found that B6.WT mice deficient for the tnf gene (B6.TNF−/−) displayed not only a non-healing cutaneous lesion but also a serious infection of the liver upon L. major inoculation. Infected B6.TNF−/− mice developed an enlarged liver that showed increased inflammation. Furthermore, we detected an accumulating monocyte-derived macrophage population (CD45+F4/80+CD11bhiLy6Clow) that displayed a M2 macrophage phenotype with high expression of CD206, arginase-1, and IL-6, supporting the notion that IL-6 could be involved in M2 differentiation. In in vitro experiments, we demonstrated that IL-6 upregulated M-CSF receptor expression and skewed monocyte differentiation from dendritic cells to macrophages. This was countered by the addition of TNF. Furthermore, TNF interfered with the activation of IL-6-induced gp130-signal transducer and activator of transcription (STAT) 3 and IL-4-STAT6 signaling, thereby abrogating IL-6-facilitated M2 macrophage polarization. Therefore, our results support the notion of a general role of TNF in the inflammatory activation of macrophages and define a new role of IL-6 signaling in macrophage polarization downstream of TNF.

The CC chemokine receptor 6 (CCR6) and its sole chemokine ligand CC chemokine ligand 20 (CCL20) display an emerging role in the coordination of humoral immune responses. Recent studies demonstrate a role of this chemokine axis in the migration of B cells to key immunological sites during an immune response, and facilitating the generation of high-quality antibodies. Very little, however, is known about CCL20 and its role in these functions. We undertook a preliminary investigation into the expression and function of CCL20 and demonstrate its well-noted upregulation in the spleen during immunization. Furthermore, we show that most follicular T helper (Tfh) cells can be CCR6+ and can produce CCL20. Surprisingly, CCL20 cannot only be found in the cytoplasm but also on the surface of these cells and their precursors. Analysis of T–B-cell conjugates revealed that mature Tfh cells, but not their precursors, are highly enriched in the conjugates. Further functional studies are needed to unravel the precise role of CCL20 in coordinating T and B cell interactions during the humoral immune response.

The absence of tumor necrosis factor (TNF) causes lethal infection by Leishmania major in normally resistant C57BL/6J (B6.WT) mice. The underlying pathogenic mechanism of this fatal disease has so far remained elusive. We found that B6.WT mice deficient for the tnf gene (B6.TNF−/−) displayed not only a non-healing cutaneous lesion but also a serious infection of the liver upon L. major inoculation. Infected B6.TNF−/− mice developed an enlarged liver that showed increased inflammation. Furthermore, we detected an accumulating monocyte-derived macrophage population (CD45+F4/80+CD11bhiLy6Clow) that displayed a M2 macrophage phenotype with high expression of CD206, arginase-1, and IL-6, supporting the notion that IL-6 could be involved in M2 differentiation. In in vitro experiments, we demonstrated that IL-6 upregulated M-CSF receptor expression and skewed monocyte differentiation from dendritic cells to macrophages. This was countered by the addition of TNF. Furthermore, TNF interfered with the activation of IL-6-induced gp130-signal transducer and activator of transcription (STAT) 3 and IL-4-STAT6 signaling, thereby abrogating IL-6-facilitated M2 macrophage polarization. Therefore, our results support the notion of a general role of TNF in the inflammatory activation of macrophages and define a new role of IL-6 signaling in macrophage polarization downstream of TNF.

During an immune response inflammatory macrophages with their wide variety of effector mechanisms including the expression of inducible nitric oxide synthase play an important part in the defense against invading pathogens. The inflammatory phenotype requires the presence of TNF which suppresses alternative activation. In the bacterial Listeria monocytogenes infection model inflammatory macrophages are crucial for protection. After infection, TNF-deficient hosts have a similar number of splenic macrophages but die rapidly. A more detailed analysis of these cells showed that while inducible nitric oxide synthase is expressed at a comparable level TNF-deficient macrophages show an increased expression of Arginase 1.

The phenotypical consequences of a combined deficiency of the Fas-Fas Ligand (FasL) and one or both Tumor Necrosis Factor (TNF) signaling pathways were investigated. Mice, which expressed a non-functional FasL suffered from a pathological accumulation of both B and T cells leading to splenomegaly and lymphadenopathy and, depending on the genetic background, pathogenic self-reactive antibodies (generalized lymphoproliferative disorder (gld)-phenotype). If mice additionally lacked TNF, they displayed a significantly ameliorated gld-phenotype while TNF Receptor-1-deficient gld mice (B6.gld.TNFR1−/−) displayed a more severe phenotype. To complement this combination, we also generated TNF Receptor-2-deficient gld mice (B6.gld.TNFR2−/−). Both double knockouts followed in their splenic structure the respective TNFR contribution to the phenotype. TNFR1−/− mice showed an absence of B cell follicles in the spleen while TNFR2−/− mice were comparable to WT mice. In general, we demonstrated a strong contribution of both TNFR signaling pathways to the symptoms of gld with the notable exception of splenomegaly where only TNFR1−/− played a role.

Recent studies in human immunodeficiency virus (HIV) have garnered interest for the role of CC chemokine receptor 6 (CCR6) and its known ligands, CC chemokine ligand 20 (CCL20) and human β-defensins, in viral entry, dissemination and antiviral immunity. Several studies have suggested that CCR6 may also act as a weak co-receptor of HIV entry, in addition to the canonical CXC chemokine receptor 4 (CXCR4) and CCR5. However, the pathogenic significance has yet to be demonstrated as the observations for preferential infection of CD4+CCR6+ over CD4+CCR6- T cells appear to be independent of CCR6 expression. This indicates means for preferential infection other than CCR6 co-receptor use. Attention has also turned to the inadvertent role of the CCR6/CCL20 axis in attracting key immune cells, including TH17 cells and dendritic cells, to sites of infection and propagating the virus to other sites of the body. This review article will summarize the latest evidence that the CCR6/CCL20 chemokine axis is playing an important role in HIV pathogenesis and immunity. Further work with in vivo studies is needed to establish the biological and, hence, therapeutic significance of these findings.

B cells are known to play a dominant pathogenic role in autoimmune conditions such as systemic lupus erythematosus (SLE) and rheumatoid arthritis. In recent times, the chemokine receptor CCR6 and its cognate ligand CCL20 have been shown to play a role in the fundamental kinetics of germinal centres and B cell responses. As CCR6 is found on B cells and is upregulated after activation, we investigated the expression of CCR6 on naive, pre-germinal centre (GC), GC/plasma cell and memory B cells in peripheral B cells of SLE patients and healthy controls using flow cytometry. Pre-germinal centre B cells are found in lower proportions and the expression of CCR6 is increased on B cells of SLE patients, suggesting a role for the chemokine pair in the pathogenesis of the disease. Further studies are needed to explore these preliminary results.

Tumor necrosis factor (TNF) is a key cytokine in the effector phase of graft-versus-host disease (GVHD) after bone marrow transplantation, and TNF inhibitors have shown efficacy in clinical and experimental GVHD. TNF signals through the TNF receptors (TNFR), which also bind soluble lymphotoxin (LTα3), a TNF family member with a previously unexamined role in GVHD pathogenesis. We have used preclinical models to investigate the role of LT in GVHD. We confirm that grafts deficient in LTα have an attenuated capacity to induce GVHD equal to that seen when grafts lack TNF. This is not associated with other defects in cytokine production or T-cell function, suggesting that LTα3 exerts its pathogenic activity directly via TNFR signaling. We confirm that donor-derived LTα is required for graft-versus-leukemia (GVL) effects, with equal impairment in leukemic clearance seen in recipients of LTα- and TNF-deficient grafts. Further impairment in tumor clearance was seen using Tnf/Lta−/− donors, suggesting that these molecules play nonredundant roles in GVL. Importantly, donor TNF/LTα were only required for GVL where the recipient leukemia was susceptible to apoptosis via p55 TNFR signaling. These data suggest that antagonists neutralizing both TNF and LTα3 may be effective for treatment of GVHD, particularly if residual leukemia lacks the p55 TNFR.

Anti-beta 2 glycoprotein 1 (anti-β2GP1) antibodies are commonly found in patients with autoimmune diseases such as the antiphospholipid syndrome (APS) and systemic lupus erythematosus (SLE). Their presence is highly associated with increased risk of vascular thrombosis and/or recurrent pregnancy-related complications. Although they are a subtype of anti-phospholipid (APL) antibody, anti-β2GP1 antibodies form complexes with β2GP1 before binding to different receptors associated with anionic phospholipids on structures such as platelets and endothelial cells. β2GP1 consists of five short consensus repeat termed “sushi” domains. It has three interchangeable conformations with a cryptic epitope at domain 1 within the molecule. Anti-β2GP1 antibodies against this cryptic epitope are referred to as ‘type A’ antibodies, and have been suggested to be more strongly associated with both vascular and obstetric complications. In contrast, ‘type B’ antibodies, directed against other domains of β2GP1, are more likely to be benign antibodies found in asymptomatic patients and healthy individuals. Although the interactions between anti-β2GP1 antibodies, β2GP1, and platelets have been investigated, the actual targeted metabolic pathway(s) and/or receptor(s) involved remain to be clearly elucidated. This review will discuss the current understanding of the interaction between anti-β2GP1 antibodies and β2GP1, with platelet receptors and associated signalling pathways.

IL-17A, IL-17F, and IL-25 are ligands for IL-17RA. In the current study, we demonstrated that IL-25–deficient mice—but not IL-17A–, IL-17F–, IL-17A/F–, IL-23p19–, or retinoic acid-related orphan receptor (ROR)-γt–deficient mice—showed significant suppression of 1) the number of eosinophils and the levels of proinflammatory mediators in bronchoalveolar lavage fluids, 2) airway hyperresponsiveness to methacholine, and 3) OVA-specific IgG1 and IgE levels in the serum during OVA-induced Th2-type/eosinophilic airway inflammation. The IL-25 deficiency did not affect lung dendritic cell migration or Ag-specific memory–Th2 cell expansion during Ag sensitization. Adoptive transfer of T cells, mast cells, or bone marrow cells from IL-25–deficient mice revealed that induction of Th2-type/eosinophilic airway inflammation was dependent on activation of lung epithelial cells and eosinophils by IL-25 produced by airway structural cells such as epithelial cells but not by such hematopoietic stem-cell-origin immune cells as T cells and mast cells. Therefore, airway structural cell-derived IL-25—rather than Th17 cell-derived IL-17A and IL-17F—is responsible for induction of local inflammation by promoting activation of lung epithelial cells and eosinophils in the elicitation phase of Th2-type/eosinophilic airway inflammation. It is not required for Ag-specific Th2 cell differentiation in the sensitization phase.

The inhibition of TNF with therapeutic monoclonal antibodies or antibody/receptor fusion proteins in rheumatoid arthritis still constitutes the benchmark for a successful intervention in an ongoing auto-immune-inflammatory disease and underlines the importance of this cytokine. TNF plays a central role in the defence against intracellular infections and is responsible for the promotion of different aspects of the innate immune response such as inflammatory cell recruitment and cell differentiation. While this cytokine generally displays pro-inflammatory activities supporting the early stages of the inflammatory response, it has been demonstrated to be especially important during infection with intracellular pathogens and, consequently, leishmaniasis of TNF(-/-) mice ends fatally. However, the specific activities of TNF that confer protection are not yet fully understood. This review will summarize the current understanding of TNF function and signalling, and will discuss recent work in the models of malaria, toxoplasmosis, trypanosomiasis and leishmaniasis with particular emphasis on work with gene-deficient mouse models.

Development of NKT cells was shown to depend on lymphotoxin (LT) and IL-15 signaling pathways as well as on cytokine receptor common gamma chain. After positive selection, NKT-cell precursors transit through progressive maturation stages including proliferative expansion within the NK1.1(-) window. The transcription factors that integrate different signaling pathways into different stages of NKT-cell development are not well characterized. Here, we show that the Rel/NF-kappaB family member RelA regulates the NK1.1(-) to NK1.1(+) transition during NKT-cell development. RelA is also required for both IL-15- and IL-7-induced proliferation of CD44(hi)NK1.1(-) NKT-cell precursors. Activation of the invariant NKT-cell receptor induces both IL-15 receptor alpha and gamma chains' expression in an NF-kappaB-dependent manner, suggesting a molecular mechanism by which NF-kappaB regulates NKT-cell development. NF-kappaB also regulates TCR-induced expression of LT-alpha and LT-beta within NKT cells. In contrast to previous reports, however, we show that LT signaling is dispensable for thymic NKT-cell development but is essential for their colonization of peripheral organs such as liver.

The generalized lymphoproliferative disorder (gld) mouse strain is characterized by severe splenomegaly/lymphadenopathy, the production of autoimmune antibodies, and the appearance of CD4/CD8-negative T cells. An additional TNF deficiency of gld/gld mice attenuates the course of the disorder through a yet-unknown mechanism. In this study, we could demonstrate that the reduced splenomegaly and lymphadenopathy in B6.gld/gld.TNF−/− mice were correlated with a decreased peripheral T cell proliferation rate and a delayed polyclonal activation. A comparative analysis of naïve T cells and memory/effector T cells showed an age-dependent difference in the T cell activation pattern in the spleen of B6.gld/gld and B6.gld/gld.TNF−/− mice. T cells from B6.gld/gld.TNF−/− spleens and lymph nodes showed significantly higher levels of CCR7 and CD62 ligand on their surface compared with B6.gld/gld mice when mice of the same age were compared. Additionally, we found an increased titer of the Th1 cytokine IFN-γ in the serum of B6.gld/gld mice, whereas the concentration of IFN-γ was markedly reduced in the serum of B6.gld/gld.TNF−/− mice. These findings support the hypothesis that increased T cell activation and proliferation in the presence of TNF contribute to the exacerbation of the gld syndrome.

Induction of inducible nitric oxide synthase in mononuclear phagocytes by IFN-γ and innate tumor necrosis factor (TNF) provide the basis for an effective immune response to the intracellular parasite Leishmania (L.) major. In previous experiments, we observed a fatal visceral form of leishmaniasis in L. major-infected C57BL/6 TNF(-/-) mice. To further delineate the protective function of TNF and its receptor requirements, we comparatively assessed L. major-infected C57BL/6 mice that were either deficient for membrane and soluble TNF (Tnf (-) (/) (-)), for soluble TNF alone (memTnf(Δ/Δ) ), or the TNF receptors type 1 (Tnfr1 (-) (/) (-)) or type 2 (Tnfr2 (-) (/) (-)). We detected locally and systemically increased levels of the cytokine IFN-γ in the absence of the TNF-TNFR1-signaling pathway. An analysis of transcription factors and cytokines revealed that activated Tnf (-) (/) (-) CD4(+) T cells displayed a highly active Th1 phenotype with a strong usage of the T cell receptor Vβ5.1/2. From these data we conclude that the fatal outcome of L. major infection in Tnf (-) (/) (-) mice does not result from a skewed or deficient Th1 differentiation.

The CC-chemokine receptor 6 (CCR6) can be detected on naive and activated B cells. Counterintuitively, its absence accelerates the appearance of germinal centres (GCs) and increases the production of low-affinity antibodies. The detailed mechanism of CCR6 function during the humoral response has remained elusive, but previously we identified a distinct CCR6high B-cell population in vivo early after antigenic challenge. In this study, we defined this population specifically as early, activated pre-GC B cells. In accordance, we show that CCR6 is upregulated rapidly within hours on the protein or mRNA level after activation in vitro. In addition, only activated B cells migrated specifically towards CCL20, the specific ligand for CCR6. Lack of CCR6 increased the dark zone/light zone ratio of GC and led to decreased antigen-specific IgG1 and IgG2a antibody generation in a B-cell intrinsic manner in mixed bone marrow chimeras. In contrast, antigen-specific IgM responses were normal. Hence, CCR6 negatively regulates entry of activated, antigen-specific pre-GC B cells into the GC reaction.

Cytotoxic lymphocytes are critical for fighting viral and certain bacterial infections. Therefore, assessing the quality of cytotoxic T cell responses might have important clinical implications. TCR-pMHC binding (avidity) is a key determinant of T cell quality. Here we review currently available technologies for the measurement of TCR-pMHC avidity and their potential relevance for translational applications.

Ontogeny and differentiation of cells of the monocyte/macrophage lineage are currently subjects of intense research. The concept of the macrophage as “simple” phagocytic cell has undergone profound changes. It has now been established that this myeloid lineage of cells is phenotypically and functionally much more diverse and exerts a much wider influence on the immune response than previously thought. How have these new findings changed our perception of the role of monocytes and macrophages in parasitic diseases? There is now strong evidence that macrophages fulfill organ-specific differential functions, exert activating as well as inhibitory effects on the adaptive immune response, and exist in a whole range of different activation statuses which show a high degree of plasticity. In the present chapter, we will not only review the pertinent literature on these new developments but also bring it into relation to the anti-infectious immune response, focusing on four parasitic diseases (trypanosomiasis, toxoplasmosis, malaria, and leishmaniasis) as examples.

Although chemokines are sufficient for chemotaxis of various cells, increasing evidence exists for their fine-tuning by selective proteolytic processing. Using a model of immune cell chemotaxis into the CNS (experimental autoimmune encephalomyelitis [EAE]) that permits precise localization of immigrating leukocytes at the blood-brain barrier, we show that, whereas chemokines are required for leukocyte migration into the CNS, additional MMP-2/9 activities specifically at the border of the CNS parenchyma strongly enhance this transmigration process. Cytokines derived from infiltrating leukocytes regulate MMP-2/9 activity at the parenchymal border, which in turn promotes astrocyte secretion of chemokines and differentially modulates the activity of different chemokines at the CNS border, thereby promoting leukocyte migration out of the cuff. Hence, cytokines, chemokines, and cytokine-induced MMP-2/9 activity specifically at the inflammatory border collectively act to accelerate leukocyte chemotaxis across the parenchymal border. publisher: Elsevier articletitle: Focal MMP-2 and MMP-9 Activity at the Blood-Brain Barrier Promotes Chemokine-Induced Leukocyte Migration journaltitle: Cell Reports articlelink: http://dx.doi.org/10.1016/j.celrep.2015.01.037 content_type: article copyright: Copyright © 2015 The Authors. Published by Elsevier Inc. ispartof: Cell Reports vol:10 issue:7 pages:1040-54 ispartof: location:United States status: published

Cerebral listeriosis is a life-threatening disease. However, little is known about the bacterial virulence factors responsible for the severe course of disease and the factors of the immune system contributing to the control of Listeria monocytogenes (LM) or even to the damage of the brain. To analyze the importance of the actA gene of LM, which mediates cell-to-cell spread of intracellular LM, the function of TNF in murine cerebral listeriosis was studied. C57BL/6 mice survived an intracerebral (i.c.) infection with actA-deficient LM, but succumbed to infection with wild-type (WT) LM. Upon infection with actA-deficient LM, macrophages and microglial cells rapidly, and later LM-specific CD4 and CD8 T cells, produced TNF. In contrast to WT mice, TNF-deficient animals succumbed to the infection within 4 days due to failure of control of LM. Histology identified a more severe meningoencephalitis, brain edema, and neuronal damage, but a reduced inducible NO synthase expression in TNF-deficient mice. Reciprocal bone marrow chimeras between WT and TNF-deficient mice revealed that hematogenously derived TNF was essential for survival, whereas TNF produced by brain-resident cells was less important. Death of TNF-deficient mice could be prevented by LM-specific T cells induced by an active immunization before i.c. infection. However, brain pathology and inflammation of immunized TNF-deficient mice were still more severe. In conclusion, these findings identify a crucial role of TNF for the i.c. control of LM and survival of cerebral listeriosis, whereas TNF was not responsible for the destruction of brain tissue.

The pro-inflammatory cytokine TNF is essential for a protective immune response to some but not all strains of Leishmania major. TNF-deficient mice of a resistant genetic background succumbed rapidly to an infection with L. major BNI. Another member of the TNF superfamily, Fas ligand (FasL), has also been reported to be critical for the immune response to L. major. To test the relative importance of TNF versus FasL for the control of L. major BNI, we infected wildtype C57BL/6 (B6.WT), B6.TNF(-/-), B6.gld and C57BL/6.gld x TNF(-/-) (B6.gld.TNF(-/-)) double-negative mice. Visceral, fatal disease was only observed in B6.TNF(-/-) mice, but not in B6 gld mice. The course of infection and the immune response of B6.gld.TNF(-/-) mice were similar to those of B6.TNF(-/-) mice. B6.gld.TNF(-/-) mice had a high tissue parasite burden and expressed prominent amounts of inducible nitric oxide synthase (iNOS) in the skin, the lymph nodes (LN) and the spleen as previously reported for B6.TNF(-/-) mice, whereas the tissue parasite load and the iNOS expression of B6.gld mice resembled that of B6.WT controls. Neither the TNF- nor the FasL-deficiency exerted a detectable intrinsic effect on the proliferation of T cells. Thus, TNF, but not FasL is essential for the control of L. major BNI. The discrepancy between these and other published data are most likely due to the use of different strains of the pathogen.

Tumor necrosis factor-alpha (TNF-alpha) is a central mediator of the immune response to pathogens, but may also exert neurotoxic effects, thereby contributing to immunopathology. To define the role of TNF during the course of brain abscess, TNF-deficient (TNF(0/0) mice were stereotaxically infected with Staphylococcus (S.) aureus-laden agarose beads. In comparison to 100% survival of wild type (WT) mice, TNF(0/0) mice displayed high mortality rates (54%) in the initial phase of abscess development as well as significantly increased morbidity in the course of the disease. The worse clinical outcome was due to an increased intracerebral (i.c.) bacterial load in TNF(0/0) mice as compared to WT mice. The impaired control of S. aureus was associated with reduced inductible nitric oxide synthase (iNOS) mRNA and protein expression in TNF(0/0)mice. Similarly, numbers of inflammatory leukocytes, cytokine expression of IL-6, IL-12p40, IFNgamma IL-beta mRNA, and brain edema were significantly increased in TNF(0/0)mice as compared to WT animals. In addition, resolution of i.c. infiltrates was delayed in TNF(0/0)mice correlating with reduced apoptosis of inflammatory leukocytes and formation of a fibrous abscess capsule. Collectively, these data demonstrate that TNF is of key importance for the control of S. aureus-induced brain abscess and regulates the ensuing host immune response.

About 40% of bone marrow-derived dendritic cells (BM-DCs) generated from stem cells of C57BL/6 (B6.WT) mice differentiate in the presence of granulocyte macrophage-colony stimulating factor (GM-CSF) without further stimuli to mature DCs. These cells are characterized by high levels of major histocompatibility complex class II, CD40, and CD86 on their surface. Recent studies have revealed that tumor necrosis factor (TNF) is crucial for maturation of BM-DCs. However, once matured, the phenotype of mature TNF-negative C57BL/6 (B6.TNF−/−) and B6.WT BM-DCs is comparable. Both expressed high levels of CD40 and CD86 and were positive for mRNA of the chemokine receptor (CCR)7. To extend our studies, we generated a monoclonal antibody (mAb) specific for mouse CCR7. This mAb allowed us to analyze the surface expression of CCR7 during maturation of B6.WT and B6.TNF−/− BM-DCs in the presence of GM-CSF and stimulated with TNF or lipopolysaccharide (LPS) and to compare it with the CCR7 expression on ex vivo-isolated splenic DCs with or without additional stimulation. Our results showed that CCR7 expression on murine BM-DCs is an indication of cell maturity. Incubation with LPS induced the maturation of all BM-DCs in culture but increased the number of mature CCR7+ splenic DCs only marginally.

In the early phase of leishmaniasis three types of potential antigen-presenting cells, including epidermal Langerhans cells (LC), dermal dendritic cells (DC) and inflammatory DC, are localized at the site of infection. Therefore, it has been a central question which cell type is responsible for the initiation of a protective immune response. In the early stage of an anti-Leishmania immune response, detectable Leishmania major antigen was localized in the paracortex of the draining lymph nodes (LN). Characterization of antigen-positive cells showed that L. major co-localized with DC of a CD11c(+) CD8 alpha(-) Langerin(-) phenotype. To determine the area of antigen uptake, dermis or epidermis, and to further define the type of antigen-transporting cells, L. major was inoculated subcutaneously and concurrently LC were mobilized with fluorescein isothiocyanate (FITC). After 3 days, DC carrying L. major antigen were always FITC(-), indicating a dermal and not an epidermal origin. Moreover, addition of L. major antigen to ex vivo isolated CD8 alpha(-) and CD8 alpha(+) DC from the draining LN of L. major-infected C57BL/6 mice demonstrated that both DC subpopulations were able to stimulate antigen-specific T cell proliferation in vitro. Without addition of exogenous antigen only the CD8 alpha(-) Langerin(-) DC were capable of stimulating antigen-specific T cell proliferation. Thus, we demonstrate that CD8 alpha(-) Langerin(-) DC and not LC are the basis of the protective immune response to intracellular L. major parasites in vivo.

Previous studies provided evidence that the role of TNF in the control of Leishmania (Leishmania) major might vary with the parasite strain. Here, we analyzed the development and outcome of cutaneous leishmaniasis in C57BL/6 wild-type (B6.WT) and TNF-deficient (B6. TNF(-/-)) mice infected with two different isolates of L. (L.) major (FRIEDLIN vs. BNI). L. (L.) major BNI caused progressive, fatal disease in B6.TNF(-/-) mice. In contrast, B6.TNF(-/-) mice infected with the L. (L.) major FRIEDLIN strain exhibited partial resistance characterized by chronic, non-healing skin lesions without lethality. Analysis of the tissue parasite numbers showed that the numbers of L. (L.) major FRIEDLIN and BNI parasites were comparable in footpads and lymph nodes of B6.TNF(-/-) mice, whereas in the spleen the parasite numbers were strikingly lower in the case of L. (L.) major FRIEDLIN. In vitro, cytokine-activated inflammatory macrophages killed L. (L.) major FRIEDLIN more efficiently than L. (L.) major BNI. These results suggest that in the absence of TNF, the course of leishmaniasis depends on the biology of the inoculated L. (L.) major strain, which most likely explains the previously published discrepant results on the role of TNF in leishmaniasis.

Chemokines are thought to control lymphocyte recruitment to the inflamed endothelium. To dissect chemokine-mediated adhesion, binding of ex vivo isolated splenocytes to tumor necrosis factor (TNF)–activated endothelial cells was analyzed under shear stress. We observed specific adhesion of naive follicular B cells, which could be blocked by pertussis toxin. This indicated a G protein–mediated binding and pointed at a contribution of chemokine receptors to B-cell adhesion. Analysis of chemokines expressed by TNF-activated endothelial cells showed that CC chemokine ligand 2 (CCL2), CCL17, and CCL20 were up-regulated. Only on follicular B cells was the cognate receptor for CCL20, CC chemokine receptor 6 (CCR6), expressed strongly, and a functional transmigration assay with CCR6-negative B cells demonstrated conclusively the sole signaling of CCL20 through CCR6. Desensitization of CCR6 on naive B cells with CCL20 resulted in receptor down-regulation and reduced B-cell adhesion. We conclude that CCL20 plays a vital role in B-cell adhesion to the inflamed endothelium.

Immunity to Toxoplasma gondii critically depends on TNFR type I-mediated immune reactions, but the precise role of the individual ligands of TNFR1, TNF and lymphotoxin-alpha (LTalpha), is still unknown. Upon oral infection with T. gondii, TNF(-/-), LTalpha(-/-), and TNF/LTalpha(-/-) mice failed to control intracerebral T. gondii and succumbed to an acute necrotizing Toxoplasma encephalitis, whereas wild-type (WT) mice survived. Intracerebral inducible NO synthase expression and-early after infection-splenic NO levels were reduced. Additionally, peritoneal macrophages produced reduced levels of NO upon infection with T. gondii and had significantly reduced toxoplasmastatic activity in TNF(-/-), LTalpha(-/-), and TNF/LTalpha(-/-) mice as compared with WT animals. Frequencies of parasite-specific IFN-gamma-producing T cells, intracerebral and splenic IFN-gamma production, and T. gondii-specific IgM and IgG titers in LTalpha(-/-) and TNF/LTalpha(-/-) mice were reduced only early after infection. In contrast, intracerebral IL-10 and IL-12p40 mRNA expression and splenic IL-2, IL-4, and IL-12 production were identical in all genotypes. In addition, TNF(-/-), LTalpha(-/-), and TNF/LTalpha(-/-), but not WT, mice succumbed to infection with the highly attenuated ts-4 strain of T. gondii or to a subsequent challenge infection with virulent RH toxoplasms, although they had identical frequencies of IFN-gamma-producing T cells as compared with WT mice. Generation and infection of bone marrow reconstitution chimeras demonstrated an exclusive role of hematogeneously produced TNF and LTalpha for survival of toxoplasmosis. These findings demonstrate the crucial role of both LTalpha and TNF for control of intracerebral toxoplasms.

Background Enoxaparin, a low-molecular-weight heparin, is known to possess anti-inflammatory properties. However, its clinical exploitation as an anti-inflammatory agent is hampered by its anticoagulant effect and the associated risk of bleeding. Objective The aim of the current study was to examine the ability of non-anticoagulant fractions of enoxaparin to inhibit the release of key inflammatory cytokines in primed peripheral blood mononuclear cells derived from allergic mild asthmatics. Methods Peripheral blood mononuclear cells from allergic asthmatics were activated with phytohaemag glutinin (PHA), concanavalin-A (ConA) or phorbol 12-myristate 13-acetate (PMA) in the presence or absence of enoxaparin fractions before cytokine levels were quantified using specific cytokine bead arrays. Together with nuclear magnetic resonance analysis,time-dependent and target-specific effects of enoxaparin fractions were used to elucidate structural determinants for their anti-inflammatory effect and gain mechanistic insights into their anti-inflammatory activity. Results Two non-anticoagulant fractions of enoxaparin were identified that significantly inhibited T-cell activation. A disaccharide fraction of enoxaparin inhibited the release of IL-4, IL-5, IL-13 and TNF-α by more than 57% while a tetrasaccharide fraction was found to inhibit the release of tested cytokines by more than 68%. Our data suggest that the observed response is likely to be due to an interaction of 6-O-sulfated tetrasaccharide with cellular receptor(s). Conclusion and Clinical Relevance The two identified anti-inflammatory fractions lacked anticoagulant activity and are therefore not associated with risk of bleeding. The findings highlight the potential therapeutic use of enoxaparin-derived fractions, in particular tetrasaccharide, in patients with chronic inflammatory disorders.

Activation of resting T cells usually leads to their proliferation and differentiation into effector cells and a subsequent decline following elimination of the antigen. A situation of excessive antigen density may result in T cell receptor (TCR)-induced deletion of T effector cells, a process termed antigen-induced cell death (AgICD). Previous studies indicate that AgICD of cytotoxic T cells may be induced by either of the two key cytotoxic processes, granule exocytosis, including perforin and granzymes, or the Fas ligand (FasL)/Fas pathway. By using in vitro-polyclonally activated or ex vivo-derived virus-induced T cell populations from mice with mutations or targeted gene defects in one or more components of the two key cytolytic pathways we now show that TCR-induced apoptosis is only impaired in the absence of FasL and/or Fas, but not in the absence of perforin and/or granzymes. Furthermore, antibody-blockage of FasL alone is sufficient to inhibit early T cell death. Inhibition of both, FasL and tumor necrosis factor (TNF-alpha) is required to abrogate late apoptosis by AgICD. The fact that antibodies to IFN-gamma also inhibit AgICD suggests that the perforin plus granzyme-independent and FaSL and/or TNF-alpha facilitated process of AgICD of T effector cells is tightly regulated by endogenous IFN-gamma.