In addition to antibody secretion, B cells have recently been recognized to function as antigen-presenting/immune-modulatory cells. The present study was designed to evaluate the efficacy of B cell depletion by anti-mouse (m) CD20 monoclonal antibody (mAb) on Graves’ hyperthyroidism in a mouse model involving repeated injection of adenovirus expressing TSHR A-subunit (Ad-TSHR289). We observe that a single injection of 250 µg/mouse anti-mCD20 mAb eliminated B cells efficiently from the periphery and spleen and to a lesser

extent from the peritoneum for more than FG-4592 3 weeks. B cell depletion before immunization suppressed an increase in serum immunoglobulin (Ig)G levels, TSHR-specific splenocyte secretion of interferon (IFN)-γ, anti-TSHR antibody production and development of hyperthyroidism. B cell depletion 2 weeks after the first immunization, Doxorubicin price a time-point at which T cells were primed but antibody production was not observed, was still effective at inhibiting antibody production and disease development without inhibiting splenocyte secretion of IFN-γ. By contrast, B cell depletion in hyperthyroid mice was therapeutically ineffective. Together, these data demonstrate that B cells are critical not only as antibody-producing cells

but also as antigen-presenting/immune-modulatory cells in the early phase of the induction of experimental Graves’ hyperthyroidism and, although therapeutically less effective, B cell depletion is highly efficient for preventing disease development. Organ-specific autoimmune diseases result from abnormal B and T cell recognition of self-autoantigen. Some of these diseases are mediated largely by humoral immune responses producing pathogenic autoantibodies, and others by cellular immune responses ever leading to destruction of target tissues by cytotoxic T cells. Graves’ disease is representative of the former, characterized by stimulatory autoantibodies against the thyrotrophin receptor [thyroid stimulating hormone receptor (TSHR)] (thyroid stimulating antibody,

TSAb), which cause overproduction of thyroid hormones and thyroid hyperplasia [1]. As antibody producing cells, B cells are crucial immune cells in the pathogenesis of Graves’ disease. In addition, other important aspects of B cell function in immune reactions have been clarified recently, including antigen presentation, proinflammatory cytokine production, co-stimulatory molecule expression (CD80 and CD86), alterations in dendritic cell function, etc. [2]. Indeed, previous studies with mice genetically deficient for B cells [B cell knock-out (KO) mice] showed the requirement of B cells for development of autoimmune thyroiditis, type 1 diabetes and systemic lupus erythematosus (SLE) [3–5].

In addition, IL-17 can directly induce tissue injury by upregulating the expression of matrix metalloproteinases. In patients with SLE, IL-17-producing T cells have been shown to infiltrate the Selleck LY2109761 lungs, skin, and kidneys [20, 25, 26], most likely contributing to end organ damage by the mechanisms mentioned above. Systemic autoimmune diseases such as SLE are characterized by the overexpression

of type I IFN-stimulated genes, referred to as the IFN signature [79, 80]. Results from phase I trials with anti-IFN-α antibody (Sifalimumab) treatment of SLE patients have demonstrated a decrease in the expression of IFN signature genes in whole blood and skin lesions and improvement in disease activity suggesting that these genes are directly involved in SLE [81, 82]. Furthermore, IFN-α chemotherapy of cancer patients induces a transient lupus-like disease in 5–20% of patients, indicating that type I IFNs are sufficient to drive SLE

[83, 84]. www.selleckchem.com/products/crenolanib-cp-868596.html Lately, a role for Th17 cells and IL-17-driven responses in the pathogenesis of SLE, rather than the previously identified type I IFN response, has been suggested and is supported by the findings that high levels of IL-17 and uncontrolled IL-17-driven inflammation can promote autoreactive B-cell responses with production of autoantibodies and induce lupus-like features in the BXD2 and Trim21−/− mice, respectively [43, 48, 85]. Interestingly, both strains also express increased levels of type I interferons; either spontaneously (BXD2) or after TLR stimulation (BXD2, Trim21−/−) [48, 85]. Therefore, although systemic Selleck Pomalidomide autoimmune diseases and SLE in particular have been described as type I IFN-driven diseases, we propose that IL-17 and type I IFN constitute a dangerous combination by acting in concert to sustain the chronic

inflammatory and autoimmune responses as discussed below. Type I IFN produced by dendritic cells (DCs) and plasmacytoid DCs (pDCs) stimulated by TLR7 agonists has been shown to support Th17 responses and IL-17 production [86, 87]. These data are particularly relevant in SLE pathogenesis since pDCs have been shown to produce type I IFN in response to stimulation by the DNA- or RNA-containing immune complexes found in sera from SLE patients [88]. In contrast, type I IFN has been shown to limit Th17-cell development by inducing the cytokine IL-27 [89]. These seemingly paradoxical actions of type I IFNs could be due to an often underappreciated role of noncanonical IFNAR signaling [90, 91]. Canonical signaling induced by type I IFNs consists of phosphorylation of STAT1 and STAT2 followed by the formation of STAT1:STAT2:IRF9 heterotrimers and STAT1:STAT1 homodimers leading to the activation of genes with ISRE- and GAS-containing promoters, respectively [92-94]. In addition to STAT1 and STAT2, noncanonical IFNAR signaling can also activate STAT3-STAT6 in immune cells.

Recombinant Tax1 and Tax2A (subtype A) proteins were purified as described recently in detail [24, 25]. Truncated Tax2A/NH2term-His tagged sequence aa 1–198 (Tax2A/1–198) (MRGSHHHHHHGS AHFPGFGQSL LYGYPVYVFG DCVQADWCPV SGGLCSTRLH RHALLATCPE HQL TWDPIDG RVVSSPLQYL IPRLPSFPTQ RTSRTLKVLT PPTTPVSPKV PPAFFQSMRK HTPYRNGCLE

PTLGDQLPSL AFPEPGLRPQ NIYTTWGKTV VCLYLYQLSP PMTWPLIPHV IFCHPRQLGA FLTKVPLKRL EELLYKM LDLQPSLIS) and truncated Tax2A/COOHterm-His tagged sequence aa 135–331 (Tax2A/135–331) (MRGSHHHHHHGS EPGLRPQNIY TTWGKTVVCL YLYQLSPPMT WPLIPHVIFC HPRQLGAFLT KVPLKRLEEL LYKMFLHTGT VIVLPEDDLP TTMFQPVRAP CIQTAWCTGL LPYHSILTTP Small molecule library high throughput GLIWTFNDGS PMISGPCPKA GQPSLVVQSS LLIFEKFQTK AFHPSYLLSH QLIQYSSFHN LHLLFDEYTN IPVSILFNKE EADDNGD LDLQPSLIS) fragments of Tax2A protein containing NF-κB domains [28, 29] were subcloned from PET29a-Tax2-H6 [30] to pQE-30 (Amp-resistant) vector and transformed into the Esherichia coli BL21(DE3) strain (subcloning generation and protein production serviced by Promab Biotechnologies, Inc., Richmond, this website CA, USA). An extract was prepared in an identical manner from E. coli

cells containing the empty vector for use as a mock control. Determination of protein concentrations was performed using a bicinchoninic acid (BCA) protein assay kit (Pierce, Rockford, IL, USA). Endotoxin concentration for all protein recombinants at the concentration (100 pM) used in the in-vitro experiments were found to be endotoxin-free, as determined PtdIns(3,4)P2 by the limulus amoebocyte lysate test (E-TOXATE; Sigma) [24]. Recombinant replication-deficient adenoviruses expressing Tax2B (subtype B) or green fluorescent protein (GFP), used to control the efficiency of transduction (Ad-Tax2B or Ad-GFP, respectively) [31], were propagated and titrated as described recently [25]. The recombinant adenovirus containing the dominant negative mutant of IκBα with serine to alanine substitutions at amino acids 32 and 36, and therefore resistant to phosphorylation-induced degradation (a NF-κB super-repressor

designated NF-κB/SR), was obtained commercially (Vector Biolabs, Philadelphia, PA, USA). In this study the two major subtypes of HTLV-2 Tax, Tax2A (expressed as recombinant protein) and Tax2B (recombinant adenovirus) were assessed to determine whether both Tax2 subtypes were able to induce the production of CC-chemokines in peripheral blood mononuclear cells. PBMCs (1 × 106/ml) in complete RPMI medium were treated with extracellular Tax (recombinant Tax1 and Tax2A) proteins at 100 pM for 1, 2, 3, 6, 12 and 24 h to determine CC-chemokine production, and for 1 and 2 h for the determination of canonical NF-κB pathway activation. Mock-treated and untreated controls were used in all experiments.

Thus, it is important to keep in mind that a certain level of DC maturity may be important for the generation of Tregs capable of inhibiting autoimmune disease [25]. The development of conventional lymphoid organ DCs in mice has been clarified recently [26]. The macrophage and DC precursor gives rise to the common DC precursor (the source of both conventional and plasmacytoid DCs). The next developmental stage for the conventional lymphoid organ DC is the pre-DC. The pre-DCs expand in the bone marrow and differentiate to conventional DCs within the spleen and mTOR inhibitor lymph nodes, where they proliferate in response to Flt3L [27]. A number of DC subsets have been

described phenotypically in both mice and humans [28]. Some of these are known to be functionally specialized [29]. For example, in mice, the DC subset expressing CD8 and DEC-205 is specialized for capture of dying cells [30] and cross-presentation of antigens on class I major histocompatibility

complex (MHC) molecules [31–33], while CD8-DCIR2+ DCs are proficient at presentation of peptides on class II MHC [32]. In addition to their well-established role in central tolerance [34], DCs employ a variety of diverse strategies and pathways to maintain T cell tolerance in the periphery (Fig. 1). Apart from induction of deletional tolerance of peripheral T cells [20,35], DCs in the steady state can also render them anergized [20] as a result of antigen recognition without sufficient co-stimulation [36]. T cell co-inhibitory molecules that transduce Z-VAD-FMK research buy negative signals, such as cytotoxic T lymphocyte antigen-4 (CTLA-4) [37] or programmed death-1 (PD-1) [38,39],

also participate in these processes. For example, steady-state DCs utilize both the PD-1 and CTLA-4 Tyrosine-protein kinase BLK pathways to induce peripheral tolerance of CD8+ T cells [40]. In addition to induction of deletion or anergy, DCs can induce increased expression of CD5 on activated T cells that leads to hyporesponsiveness, at least in the setting of the induced autoimmune disease, experimental acute encephalomyelitis [41]. Expression of Fas on antigen-presenting cells is also important for the maintenance of peripheral tolerance and the avoidance of autoimmunity [42], while the production of indoleamine 2,3-dioxygenase (IDO) by DCs is involved in peripheral tolerance in certain specialized settings [43,44]. Finally, DCs are involved in the in vivo expansion of thymic-derived natural CD4+CD25+ Tregs[45] as well as the induction of adaptive forkhead box P3 (FoxP3+) Tregs[45–48] and CD8+ Tregs[49], and interleukin (IL)-7 produced by immature DCs appears to function as a CD4+CD25+ Treg survival factor [50]. Multiple lines of investigation indicate that priming of pathogenic beta cell-specific T cells occurs in the pancreatic lymph nodes. For example, adoptive transfer of 5,6-carboxy-succinimidyl-fluorescein-ester (CFSE)-labelled transgenic CD4+ BDC2.

The molecular mechanisms through which IRF4 can influence the development of Tc9 and Th9 cells seem to be very similar. Thus,

like in Th9 cells, IRF4 is essential for IL-9 expression in Tc9 cells and binds to the Il9 promoter (author’s unpublished data). Moreover, in Irf4–/– Tc9 cells, the expression of FOXP3 was found to be elevated and retroviral overexpression of FOXP3 suppressed IL-9 production in WT Tc9 cells [63]. Inhibition of IL-9 production by FOXP3 has also been shown in Th9 cells [29]. These data suggest that IRF4 regulates IL-9 production in Tc9 cells both directly via binding to the Il9 promoter and indirectly via affecting selleck products FOXP3 expression (Fig. 2) [63]. Tc17 cells are characterized by the production of IL-17 and expression of the Tc17-specific transcriptional program including mRNAs for ROR-γt, RORα, IL-21, and IL-23 receptor (IL-23R) [64, 66, 73]. Tc17 cells have been identified in MS lesions [74]. Likewise, upon immunization with a truncated peptide from myelin oligodendrocyte glycoprotein (MOG37–50), WT mice suffer from EAE accompanied by increased numbers of IL-17-producing CD8+ T cells in the LNs and CNS [66]. By contrast,

Irf4–/– mice have been shown to be resistant to the induction of EAE and failed to develop IL-17-producing CD8+ T cells, illustrating the need for IRF4 not only for Th17-, but also for Tc17-cell differentiation in vivo. Also in vitro, IRF4 was required for the acquisition of the Tc17 phenotype: Irf4–/– CD8+ T cells failed to Selleck Navitoclax produce IL-17 upon culturing with TGF-β and IL-6 and expressed greatly diminished levels

of mRNAs characteristic of a Tc17-specific transcriptional program. Instead, under Tc17-inducing next conditions, Irf4–/– cells displayed enhanced expression of EOMES and FOXP3, which are master regulators of CTL and CD8+ Treg-cell differentiation, respectively. Forced expression of EOMES and FOXP3 additively inhibited IL-17 production by WT CD8+ T cells, illustrating that the high amounts of these transcription factors contribute to the altered phenotype of Irf4–/– CD8+ T cells. Thus, on the one hand, IRF4 acts as molecular activator of Tc17-cell differentiation by promoting expression of the master regulators ROR-γt and RORα, and on the other hand, IRF4 acts as suppressor of alternative CD8+ T-cell fates by downregulating the expression of EOMES and FOXP3 (Fig. 2) [24]. In addition, our data revealed that MOG37–50-induced EAE is mediated by reciprocal cooperation between IL-17A-producing Tc17 cells and CCR6-expressing Th17 cells [24]. Although WT CD8+ T cells that were transferred into Irf4–/– mice prior to EAE induction developed a Tc17-like phenotype, these cells failed to migrate into the CNS and to induce autoimmune inflammation. Help by CCR6-expressing Th17 cells was required to enable WT Tc17-cell-mediated CNS inflammation.

5D). The accumulation of Treg became more obvious at 14 days, when 15–20% of the cells expressed Foxp3 (Fig. 5C and D). It was accompanied by a contraction of the OT-II repertoire, greater than the one observed in mice injected only with PBS or with isotype-matched control mAb (Fig. 5A). We conclude

that antigen targeting to DNGR-1 in non-inflammatory conditions leads to a strong contraction of the antigen-specific T-cell compartment and allows the peripheral conversion of some remaining naïve T cells into PARP inhibitor Foxp3+ Treg. Antigen targeting to DC in vivo is emerging as an attractive strategy for immunomodulation 3, 4. Ab-mediated delivery of antigenic epitopes to DC has variably been shown to allow priming of CD4+ and CD8+ T-cell immunity or to induce tolerance through deletion or conversion of antigen-specific T cell into Treg 3, 4. An ideal target should be a surface receptor that delivers the targeting Ab to endocytic and cytosolic compartments for processing of the linked antigenic moiety and subsequent (cross)presentation by MHC class I and/or class II molecules. In Selleckchem HIF inhibitor addition, it might be desirable to target a “neutral” receptor, i.e. one that does not activate DC upon Ab binding, in order to be able to induce tolerance or to tune immunity by co-administering specific

immunomodulators. Finally, the target receptor should be restricted to DC, in particular to DC subsets with proved capacity for antigen presentation to T cells. In this study, we show that DNGR-1 fits all of these criteria. DNGR-1-targeted antigens are presented to CD4+ T cells selectively by CD8α+ DC without promoting strong Th-cell priming. Adjuvants can be co-administered to selectively induce Th1 or Th17 responses. In addition, small amounts of DNGR-1-targeted antigen in the absence of adjuvant can be used to delete antigen-specific T cells and promote Treg conversion. Although CD8α+ DC have been suggested to be less efficient in MHC class II antigen presentation cAMP than other DC subtypes 21, this study and many others demonstrate that they are able to present antigens to CD4+ T cells in vivo8, 26. They also excel in antigen

crosspresentation to CD8+ T cells 21, 26, 27 and, therefore, can concomitantly present antigen to both CD4+ and CD8+ T lymphocytes, allowing optimal delivery of CD4+ T-cell help for CTL priming. In addition, as shown here, CD8α+ DC can drive the differentiation of Th1 or Th17 cells depending on the adjuvant. Although the ability of CD8α+ DC to trigger a Th1 response is well documented, this is the first instance when these cells have been shown to induce Th17 differentiation. These data therefore indicate that CD8α+ DC are not ontogenetically pre-programmed to induce Th1 responses and highlight the previously noted importance of innate signals in regulating DC subset function and instruction of adaptive immune responses 28, 29.

The cerebral cortex was relatively maintained, and cortical atrophy and

neuronal loss and atrophy were mild. Metachromatic substance deposition was not seen. Although slight small vessel proliferation was confirmed, there were no globoid cells in the vicinity. In the thalamus, loosening was confirmed. The hypothalamus was maintained, and in the hippocampus, marked neuronal loss was seen. Although the cerebellum was relatively maintained, mild degeneration was seen in the white Doxorubicin cell line matter. In the brain, no membranocystic lesion was observed. The patient had a conversion of nucleotide at position 116 resulting in serine 38 to asparagine substitution. In the 1961 Convention of the Japanese Orthopedic Association, Terayama named selleck chemicals the characteristic osseous lesions of the disease multiple membranocystoses.6 After performing an autopsy, Nasu and colleagues

tentatively named the disease membrano-cystic lipodystrophy and presented it at the 1970 Tokyo Pathology Conference. Then, in the 1971 Convention of the Japanese Society of Pathology and the 1972 Convention of the Japanese Society of Neuropathology, the disease was referred to as membranous lipodystrophy.7 The disease was also reported in Finland and Sweden, and in 1964, Järvi and colleagues coined the term “cystic capillary-necrotic osteodysplasia”.8 In 1970, Hakola and colleagues documented “osteodysplasia polycystica hereditaria combined

with sclerosing leucoencephalopathy”, and Hakola reported thorough studies in 1972.2,9 In 1973, when Yakumaru reported an autopsy case of membranous lipodystrophy, he pointed out similarities with Finnish and Swedish cases.10 In Japan, the disease was named Nasu disease, and in 1981, Hanawa and colleagues proposed that since Nasu disease, membranous lipodystrophy, and the Finnish cases were the same, the diseases should be referred to as NHD.11 In Finland, the disease was named “polycystic lipomembranous osteodysplasia with sclerosing leucoencephalopathy” (PLOSL) in 1980.12 So far, there have been approximately 200 cases, and while most patients have been Japanese over and Finnish, cases have been reported in Sweden,13,14 the USA,15,16 Norway,17 Italy,18 South Africa,19 Austria,20 Turkey,21 Belgium,22 France,23 Brazil,4 Germany,24 Spain25 and Bolivia.26 The prevalence in Finland has been reported as 2/1000 000.27 A typical patient often has bone symptoms occurring during adolescence and slowly progressive dementia. Most patients first experience bone symptoms, and beginning with hand, foot and knee pains and with increased susceptibility, the patient suffers repeated pathological fractures. Dementia is mostly characterized by personality changes, and patients experience euphoria, indifference, slouchy lifestyle, apathy, disinhibition, and lack of insight into their disease.

Perren, I.

Bravi, L. Jennen, A. Feuchtinger, J. Drouin, F. Roncaroli and N. S. Pellegata (2013) Neuropathology and Applied Neurobiology39, 256–269 Characterization of MENX-associated pituitary tumours Aims: The aim of this study is to evaluate the pathological features, serum hormone levels and ex vivo cultures of pituitary adenomas that occur in rats affected by MENX syndrome. MENX is multiple endocrine neoplasia syndrome caused by a germline p38 MAPK activation mutation in the cell cycle inhibitor p27. Characterization of MENX adenomas is a prerequisite to exploit this animal model for molecular and translational studies of pituitary adenomas. Methods: We investigated MENX pituitary adenomas with immunohistochemistry, double immunofluorescence, electron microscopy, reverse transcription

polymerase chain reaction (RT-PCR), measurement of serum hormone levels and ex vivo cultures. Results: Adenomas Cobimetinib in MENX rats belong to the gonadotroph lineage. They start from 4 months of age as multiple neoplastic nodules and progress to become large lesions that efface the gland. Adenomas are composed of chromophobic cells predominantly expressing the glycoprotein alpha-subunit (αGSU). They show mitotic activity and high Ki67 labelling. A few neoplastic cells co-express gonadotropins and the transcription factor steroidogenic factor 1, together with growth hormone or prolactin and Pit-1, suggesting that they are not fully committed to one Nabilone cell lineage. Ex vivo cultures show features similar to the primary tumour. Conclusions: Our results suggest that p27 function is critical to regulate gonadotroph cells growth. The MENX syndrome represents a unique model to elucidate the physiological and molecular mechanisms mediating the pathogenesis of gonadotroph adenomas. “
“Intracranial malignant solitary fibrous tumor (SFT) is very rare. It was identified in a 39-year-old female patient who underwent malignant transformation over 6 months. MRI revealed an 8 × 5 × 6 cm mass with heterogenous strong enhancement in the left occipital lobe. Histologic findings and immunophenotype (positive for CD34, bcl-2 and vimentin, and negative for epithelial membrane

antigen or S100 protein) of the primary tumor were typical of SFT. However, there was a focal area (<10% of tumor volume) showing hypercellularity, nuclear pleomorphism and increased Ki-67 labeling index (LI) of 10% (average, 1%). At the second operation, the recurrent tumor revealed full-blown histologic features of malignant SFT, such as infiltrative brain invasion, marked nuclear pleomorphism, frequent mitotic figures (15/10 high power fields), and high Ki-67 LI (25%). The presence of atypical histologic finding or increased Ki-67 LI in the typical SFT, although it is focal, needs to be mentioned in the diagnosis and also may require more aggressive surgical management. "
“Circumventricular organs (CVOs) are specialized ventricular structures around the third and fourth ventricles of the brain.

Some Treg cells also infiltrate germinal centers to negatively regulate TFH cells and this https://www.selleckchem.com/products/PD-0332991.html process would lead to higher affinity B-cell responses [[20, 21]]. Finally, mast cells also directly activate B cells to induce IgA production via CD40L, IL-6, and IL-10 [[121]]. This activation

may contribute to TI IgA responses in the intestinal lamina propria. Basophils, an innate cell type closely related to mast cells, also deliver helper signals to B cells via both direct and indirect mechanisms (Fig. 4). Firstly, under certain circumstances, basophils can migrate to draining lymph nodes where they release IL-4 to induce the formation of TH2 cells, an IL-4-producing T-cell subset critically involved in the induction of protective IgG1 and IgE responses against various www.selleckchem.com/products/LBH-589.html allergens and pathogens, including helminths [[122-125]]. Secondly, after secondary immunization, basophils recognize antigen through prebound antigen-specific IgE generated during a primary immune response [[126]]. Antigen recognition via IgE causes upregulation of CD40L and release of IL-4 and IL-6, which provide antibody-inducing

signals to B cells not only directly, but also indirectly via enhancement of IL-4, IL-6, IL-10, and IL-13 production by TH2 cells [[112, 126]]. Presumably, the antigen-IgE interaction does not trigger pathological release of preformed highly inflammatory compounds, such as histamine, from basophils owing to the low affinity

of IgE for antigen. It must be also noted that IgE can also bind DCs, which raises the possibility that DCs could account for at least part of the Th2-inducing activity ascribed to basophils [[127]]. Basophils may deliver similar B-cell helper signals by interacting with IgD (Fig. 4), an enigmatic antibody isotype released by IgD class-switched plasmablasts originating in the human upper respiratory mucosa [[52, 128]]. In spite of being heavily hypermutated, these IgD antibodies are largely polyreactive and may afford mucosal protection by binding not only to commensals and pathogens but also to their virulence factors [[52, 83, 129, 130]]. In addition to crossing the epithelial barrier to reach the surface of upper respiratory mucosal surfaces, IgD binds to circulating basophils, monocytes, and neutrophils, as well Interleukin-2 receptor as mucosal mast cells, via an unknown receptor [[52]]. Crosslinking of prebound IgD induces basophil release of BAFF, IL-4, and IL-13, which in turn stimulate B cells to undergo IgM production, as well as CSR to IgG and IgA, in a TI manner [[52]]. CD40L and APRIL further help the activation of B cells by IgD-activated basophils [[52]]. Thus, basophils may utilize both prebound IgE and IgD as immune amplifiers of both systemic and mucosal B-cell responses. TFH cells, TFR cells, NKTFH cells, and Treg cells play a pivotal role in TD antibody responses against microbial proteins.

[4] It has been demonstrated that allergens in the presence of

endotoxins trigger a substantially stronger allergic inflammation, compared with that evoked in the absence of endotoxins.[5-7] After inhalation, endotoxins, such as lipopolysaccharide (LPS), encounter and activate alveolar macrophages, leading to the production and release of pro-inflammatory cytokines, chemokines, adhesion molecules and other mediators.[8] Nasal and lung lavage samples of allergic subjects show increased levels of interleukin-1β (IL-1β),[9] primarily produced by activated macrophages.[10] Production Crizotinib molecular weight of mature IL-1β requires distinct signals, some of which induce gene expression in the so called ‘priming step’, whereas other signals trigger the maturation of pro-IL-1β to IL-1β by a multiprotein complex called inflammasome. The NLRP3 inflammasome complex consists of NLRP3 (NOD-like receptor family pyrin domain-containing 3) sensor, caspase-1 and ASC (apoptosis-associated speck-like protein containing a caspase recruitment domain) adaptor.[11, 12] NLRP3 inflammasomes play a crucial role in the detection and sensing of exogenous danger signals like pathogen-associated molecular patterns and toxins of microbes, asbestos or silica, as well as endogenous danger signals like monosodium urate and amyloid.[13, 14] Most NLRP3 activators have been shown to induce ROS find more generation,[15]

and click here inhibitors of ROS production or ROS scavengers attenuate NLRP3 inflammasome activation[16] implying an essential role for ROS in NLRP3 function. As pollen NADPH oxidases are able to generate ROS, and ROS have been implicated in the NLRP3 inflammasome-mediated IL-1β production, we hypothesized that exposure to pollen extract may influence inflammatory responses and IL-1β production of macrophages via NLRP3 inflammasome. Here we report for the first time that ragweed

pollen extract (RWE), typically used as a model for pollen action,[3] significantly elevates LPS-induced IL-1β production of THP-1 or primary macrophages and dendritic cells in an NADPH-dependent manner. We also demonstrate that a caspase-1 inhibitor or NLRP3 silencing abolish this enhancing effect together with the original LPS-triggered inductions. We also show that RWE in the presence of NADPH enhances LPS-induced p38 and Jun N-terminal kinase (JNK) signalling pathways resulting in the activation of AP-1 transcription factors and the subsequent gene transcription/expression of pro-IL-1β and key components of the inflammasome. This effect is mediated by a ROS-dependent mechanism. The THP-1 cell line (ATCC TIB-202) was a generous gift from Professor Laszlo Nagy. THP-1 monocytes were cultured in RPMI-1640 (Gibco BRL Inc., Grand Island, NY) containing 10% heat-inactivated fetal calf serum, penicillin-streptomycin and glutamine, and maintained at 37° under 5% CO2.