Development of Monoclonal Antibody-Based EIA for Tetranor-PGDM which Reflects PGD2 Production in the Body.

Tetranor-PGDM is a metabolite of PGD2. Urinary tetranor-PGDM levels were reported to be increased in some diseases, including food allergy, Duchenne muscular dystrophy, and aspirin-intolerant asthma. In this study, we developed a monoclonal antibody (MAb) and a competitive enzyme immunoassay (EIA) for measuring tetranor-PGDM. Spleen cells isolated from mice immunized with tetranor-PGDM were utilized to generate Ab-producing hybridomas. We chose hybridomas and purified MAb against tetranor-PGDM to develop competitive EIA. The assay evaluated the optimal ionic strength, pH, precision, and reliability. Specificity was determined by cross-reactivity to tetranor-PGEM, tetranor-PGFM, and tetranor-PGAM. Recovery was determined by spiking experiments on artificial urine. Optimal ionic strength was 150 mM NaCl, and optimal pH was pH 7.5. Metabolites other than tetranor-PGDM did not show any significant cross-reactivity in the EIA. The assay exhibited a half-maximal inhibition concentration (IC50) of 1.79 ng/mL, limit of detection (LOD) of 0.0498 ng/mL, and range of quantitation (ROQ) value of 0.252 to 20.2 ng/mL. The intra- and inter-assay variation for tetranor-PGDM was 3.9-6.0% and 5.7-10.4%, respectively. The linearity-dilution effect showed excellent linearity under dilution when artificial urine samples were applied to solid-phase extraction (SPE). After SPE, recovery of tetranor-PGDM in artificial urine averaged from 82.3% to 113.5% and was within acceptable limits (80%-120%). We successfully generated one monoclonal antibody and developed a sensitive competitive EIA. The established EIA would be useful for routine detection and monitoring of tetranor-PGDM in research or diagnostic body fluids.

Eosinophils increase macrophage ability to control intracellular Leishmania amazonensis infection via PGD2 paracrine activity in vitro.

Clinical and experimental studies have described eosinophil infiltration in Leishmania amazonensis infection sites, positioning eosinophils strategically adjacent to the protozoan-infected macrophages in cutaneous leishmaniasis. Here, by co-culturing mouse eosinophils with L. amazonensis-infected macrophages, we studied the impact of eosinophils on macrophage ability to regulate intracellular L. amazonensis infection. Eosinophils prevented the increase in amastigote numbers within macrophages by a mechanism dependent on a paracrine activity mediated by eosinophil-derived prostaglandin (PG) D2 acting on DP2 receptors. Exogenous PGD2 mimicked eosinophil-mediated effect on managing L. amazonensis intracellular infection by macrophages and therefore may function as a complementary tool for therapeutic intervention in L. amazonensis-driven cutaneous leishmaniasis.

The Prostaglandin D2 Receptor CRTH2 Promotes IL-33-Induced ILC2 Accumulation in the Lung.

Group 2 innate lymphoid cells (ILC2s) are rare innate immune cells that accumulate in tissues during allergy and helminth infection, performing critical effector functions that drive type 2 inflammation. ILC2s express ST2, the receptor for the cytokine IL-33, and chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2), a receptor for the bioactive lipid prostaglandin D2 (PGD2). The IL-33-ST2 and the PGD2-CRTH2 pathways have both been implicated in promoting ILC2 accumulation during type 2 inflammation. However, whether these two pathways coordinate to regulate ILC2 population size in the tissue in vivo remains undefined. In this study, we show that ILC2 accumulation in the murine lung in response to systemic IL-33 treatment was partially dependent on CRTH2. This effect was not a result of reduced ILC2 proliferation, increased apoptosis or cell death, or differences in expression of the ST2 receptor in the absence of CRTH2. Rather, data from adoptive transfer studies suggested that defective accumulation of CRTH2-deficient ILC2s in response to IL-33 was due to altered ILC2 migration patterns. Whereas donor wild-type ILC2s preferentially accumulated in the lungs compared with CRTH2-deficient ILC2s following transfer into IL-33-treated recipients, wild-type and CRTH2-deficient ILC2s accumulated equally in the recipient mediastinal lymph node. These data suggest that CRTH2-dependent effects lie downstream of IL-33, directly affecting the migration of ILC2s into inflamed lung tissues. A better understanding of the complex interactions between the IL-33 and PGD2-CRTH2 pathways that regulate ILC2 population size will be useful in understanding how these pathways could be targeted to treat diseases associated with type 2 inflammation.

The relationship between inflammatory markers and spirometric parameters in ACOS, Asthma, and COPD.

Objective: The inflammatory mechanisms underpinning asthma-chronic obstructive pulmonary disease (COPD) overlap syndrome (ACOS) have not been fully elucidated. Here, we examined the levels of cysteinyl leukotrienes (cys-LTs), prostaglandin D2 (PG-D2), prostaglandin E2 (PG-E2), interleukin 5 (IL-5), and a disintegrin and metalloprotease domain (ADAM 33) in ACOS patients to determine the relationship between levels of these inflammatory markers and pulmonary functions.Methods: Blood samples were obtained from asthma, COPD, and ACOS patients who received combined therapy and were stable for the last month to measure cys-LTs, PG-D2, PG-E2, IL-5, and ADAM33 levels. Differences between groups and their correlations with pulmonary function tests were evaluated.Results: In total, 24 ACOS, 27 asthma, and 35 COPD patients were included. . PG-D2 levels were higher in ACOS (120.9 ± 117.2 ng/L) and asthma (119.6 ± 111.7 ng/L) patients than in COPD (82.6 ± 46.7 ng/L) patients (p = 0.036 and p = 0.038, respectively). In ACOS patients, PG-D2, cys-LTs, and ADAM33 levels were negatively correlated with FEV1/FVC% values (p = 0.021, p = 0.008, and p = 0.028, respectively). In COPD patients, a negative correlation was detected between PG-E2 and FEV1/FVC% (p = 0.007), whereas positive correlations were detected between IL-5 and pulmonary function tests, including FVC, FVC%, FEV1, FEV1%, FEF25-75, and FEF25-75% (p = 0.047, p = 0.005, p = 0.002, p = 0.002, p = 0.010, and p = 0.005, respectively). In asthma patients, cys-LTs levels were negatively correlated with FEV1 and FEF25-75 values (p = 0.045 and p = 0.037, respectively).Conclusions: PG-D2 levels may be a valuable biomarker to differentiate COPD in asthma and ACOS patients.

Untargeted metabolomics reveals alterations in metabolites of lipid metabolism and immune pathways in the serum of rats after long-term oral administration of Amalaki rasayana.

Amalaki rasayana, a traditional preparation, is widely used by Ayurvedic physicians for the treatment of inflammatory conditions, cardiovascular diseases, and cancer. Metabolic alterations induced by Amalaki rasayana intervention are unknown. We investigated the modulations in serum metabolomic profiles in Wistar rats following long-term oral administration of Amalaki rasayana. Global metabolic profiling was performed of the serum of rats administered with either Amalaki rasayana (AR) or ghee + honey (GH) for 18 months and control animals which were left untreated. Amalaki rasayana components were confirmed from AR extract using HR-LCMS analysis. Significant reductions in prostaglandin J2, 11-dehydrothromboxane B2, and higher levels of reduced glutathione and glycitein metabolites were observed in the serum of AR administered rats compared to the control groups. Eleven different metabolites classified as phospholipids, glycerophospholipids, glucoside derivatives, organic acids, and glycosphingolipid were exclusively observed in the AR administered rats. Pathway analysis suggests that altered metabolites in AR administered rats are those associated with different biochemical pathways of arachidonic acid metabolism, fatty acid metabolism, leukotriene metabolism, G-protein mediated events, phospholipid metabolism, and the immune system. Targeted metabolomics confirmed the presence of gallic acid, ellagic acid, and arachidonic acid components in the AR extract. The known activities of these components can be correlated with the altered metabolic profile following long-term AR administration. AR also activates IGF1R-Akt-Foxo3 signaling axis in heart tissues of rats administered with AR. Our study identifies AR components that induce alterations in lipid metabolism and immune pathways in animals which consume AR for an extended period.

New treatments for asthma: From the pathogenic role of prostaglandin D2 to the therapeutic effects of fevipiprant.

Prostaglandin D2 (PGD2) is a pleiotropic mediator, significantly involved in the pathogenesis of type 2 (T2) asthma because of its biologic actions exerted on both immune/inflammatory and airway structural cells. In particular, the pro-inflammatory and pro-remodelling effects of PGD2 are mainly mediated by stimulation of chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2). This receptor is the target of the oral competitive antagonist fevipiprant, which on the basis of recent phase II studies is emerging as a potential very promising anti-asthma drug. Indeed, fevipiprant appears to be safe and effective, especially in consideration of its ability to inhibit eosinophilic bronchial inflammation and improve forced expiratory volume in one second (FEV1). Further ongoing phase III trials will definitely clarify if fevipiprant can prospectively become a valid option for an efficacious add-on treatment of moderate-to-severe T2-high asthma.

Mast Cell Activation Syndrome and Mastocytosis: Initial Treatment Options and Long-Term Management.

Patients with clonal mast cell activation syndromes (MCAS) including cutaneous and systemic mastocytosis (SM) may present with symptoms of mast cell activation, but in addition can have organ damage from the local effects of tissue infiltration by clonal mast cells. Patients with nonclonal MCAS may have chronic or episodic mast cell activation symptoms with an increase in serum tryptase and/or urinary metabolites of histamine, prostaglandin D2, and leukotrienes. Symptoms of MCAS and SM can be managed by blockade of mediator receptors (H1 and H2 antihistamines, leukotriene receptor blockade), inhibition of mediator synthesis (aspirin, zileuton), mediator release (sodium cromolyn), anti-IgE therapy, or a combination of these approaches. Acute episodes of mast cell activation require epinephrine, and prolonged episodes may be addressed with corticosteroids. Patients with clonal mast cell syndromes may need a reduction in the number of mast cells to prevent severe symptoms including anaphylaxis and/or progression to aggressive diseases.

Prostaglandin D2 Levels Regulate CD103+ Conventional Dendritic Cell Activation in Neonates During Respiratory Viral Infection.

During respiratory viral infection, conventional dendritic cells (cDCs) take up antigen and migrate to the draining lymph nodes to present viral antigen and activate cytotoxic T lymphocytes; however, regulation of cDC activation and migration may be age dependent. In this study, we used a mouse model of paramyxoviral infection (Sendai virus) and demonstrated that cDCs, which have migrated from lungs to the draining lymph nodes, are delayed in expressing activation markers in neonatal mice compared with adults. Neonatal lung cDCs expressed reduced levels of MHC Class II (major histocompatibility complex II) and CCR7 (chemokine receptor type 7) on postinfection days 3 and 5, respectively. The level of the CCR7 ligand CCL19 was significantly reduced in neonatal lungs during the course of viral infection. Interestingly, the arachidonic acid metabolite prostaglandin D2 (PGD2) was present at significantly higher levels in neonatal bronchoalveolar lavage fluid compared with adults. This was associated with increased expression of lipocalin PGD2 synthase mRNA levels in neonatal lungs and in isolated neonatal tracheal epithelial cells. Although thymic stromal lymphopoietin (TSLP) expression has been associated with increased PGD2 production, we found that TSLP levels were reduced in neonatal lungs. Importantly, blocking PGD2 function using a prostaglandin D2 receptor 1 (DP1) antagonist restored cDC activation in neonates. Together, these data suggest that cDC activation in neonates is delayed by a PGD2 mechanism and associated decreased chemokine signals.

Synergistic activation of pro-inflammatory type-2 CD8+ T lymphocytes by lipid mediators in severe eosinophilic asthma.

Human type-2 CD8+ T cells are a cell population with potentially important roles in allergic disease. We investigated this in the context of severe asthma with persistent airway eosinophilia-a phenotype associated with high exacerbation risk and responsiveness to type-2 cytokine-targeted therapies. In two independent cohorts we show that, in contrast to Th2 cells, type-2 cytokine-secreting CD8+CRTH2+ (Tc2) cells are enriched in blood and airways in severe eosinophilic asthma. Concentrations of prostaglandin D2 (PGD2) and cysteinyl leukotriene E4 (LTE4) are also increased in the airways of the same group of patients. In vitro PGD2 and LTE4 function synergistically to trigger Tc2 cell recruitment and activation in a TCR-independent manner. These lipids regulate diverse genes in Tc2 cells inducing type-2 cytokines and many other pro-inflammatory cytokines and chemokines, which could contribute to eosinophilia. These findings are consistent with an important innate-like role for human Tc2 cells in severe eosinophilic asthma and suggest a potential target for therapeutic intervention in this and other diseases.

Interleukin-6 potentiates FcεRI-induced PGD2 biosynthesis and induces VEGF from human in situ-matured skin mast cells.

BACKGROUND: Interleukin-6 is a gp130 utilizing cytokine that is consistently associated with allergic diseases like asthma and urticaria in humans where mast cells are known to play a critical role. However, the role of IL-6 in allergic disease in not known. IL-6 was reported to enhance degranulation of in vitro-derived mast cells, but the effect of IL-6 on mediator release from human in situ-matured tissue-isolated mast cells had not been reported. METHODS: Human mature mast cells were isolated and purified from normal skin tissue from different donors. The expression of surface-expressed IL-6 receptors was demonstrated by flow cytometry. The effect of IL-6 on FcεRI-induced degranulation, PGD2 biosynthesis, and cytokine production was determined with ß­hexosaminidase release assay, Western blotting, quantitative real-time PCR, and ELISA. The small molecule inhibitor of STAT-3, C188-9, was used to demonstrate STAT3 dependency. RESULTS: IL-6 significantly potentiated FcεRI-induced PGD2 biosynthesis, but had no effect on degranulation. IL-6 also induced VEGF gene expression and protein secretion, and enhanced FcεRI-induced IL-8 production. Mechanistically, IL-6 enhanced FcεRI-induced COX­2 expression, PGD2 biosynthesis, and VEGF production in a STAT3 dependent manner. CONCLUSION: Here, we demonstrate that IL-6 is a potentiator of FcεRI-induced PGD2 biosynthesis, and can induce or enhance production of pro-angiogenesis factors VEGF and IL-8 from human in situ-matured skin mast cells. GENERAL SIGNIFICANCE: These findings from this study indicate that IL-6 contributes to human allergic disease by enhancing the production of inflammatory PGD2 from tissue-resident mast cells. Moreover, the data suggest a novel role for IL-6 in mast cell-mediated angiogenesis.

Prostaglandin D2 amplifies lupus disease through basophil accumulation in lymphoid organs.

In systemic lupus erythematosus (SLE), autoantibody production can lead to kidney damage and failure, known as lupus nephritis. Basophils amplify the synthesis of autoantibodies by accumulating in secondary lymphoid organs. Here, we show a role for prostaglandin D2 (PGD2) in the pathophysiology of SLE. Patients with SLE have increased expression of PGD2 receptors (PTGDR) on blood basophils and increased concentration of PGD2 metabolites in plasma. Through an autocrine mechanism dependent on both PTGDRs, PGD2 induces the externalization of CXCR4 on basophils, both in humans and mice, driving accumulation in secondary lymphoid organs. Although PGD2 can accelerate basophil-dependent disease, antagonizing PTGDRs in mice reduces lupus-like disease in spontaneous and induced mouse models. Our study identifies the PGD2/PTGDR axis as a ready-to-use therapeutic modality in SLE.

Tumour-derived PGD2 and NKp30-B7H6 engagement drives an immunosuppressive ILC2-MDSC axis.

Group 2 innate lymphoid cells (ILC2s) are involved in human diseases, such as allergy, atopic dermatitis and nasal polyposis, but their function in human cancer remains unclear. Here we show that, in acute promyelocytic leukaemia (APL), ILC2s are increased and hyper-activated through the interaction of CRTH2 and NKp30 with elevated tumour-derived PGD2 and B7H6, respectively. ILC2s, in turn, activate monocytic myeloid-derived suppressor cells (M-MDSCs) via IL-13 secretion. Upon treating APL with all-trans retinoic acid and achieving complete remission, the levels of PGD2, NKp30, ILC2s, IL-13 and M-MDSCs are restored. Similarly, disruption of this tumour immunosuppressive axis by specifically blocking PGD2, IL-13 and NKp30 partially restores ILC2 and M-MDSC levels and results in increased survival. Thus, using APL as a model, we uncover a tolerogenic pathway that may represent a relevant immunosuppressive, therapeutic targetable, mechanism operating in various human tumour types, as supported by our observations in prostate cancer.Group 2 innate lymphoid cells (ILC2s) modulate inflammatory and allergic responses, but their function in cancer immunity is still unclear. Here the authors show that, in acute promyelocytic leukaemia, tumour-activated ILC2s secrete IL-13 to induce myeloid-derived suppressor cells and support tumour growth.

miR-27b-3p, miR-181a-1-3p, and miR-326-5p are involved in the inhibition of macrophage activation in chronic liver injury.

Macrophages are central players in inflammation, which leads to liver injury. It has been reported that continuous macrophage activation initiates this process. Our previous data show that the anti-inflammatory factor, 15-deoxy-Δ12, 14-prostaglandin J2 (15d-PGJ2), inhibits bone marrow (BM)-derived macrophage (BMM) migration and inflammatory cytokine production. However, the underlying mechanism of 15d-PGJ2 inhibited BMM activation is still unclear. Here, we evaluate the role of 15d-PGJ2/PPARγ axis in BMM activation. 15d-PGJ2 reduced activated BMM population in injured livers. Inflammatory cytokine expressions (MIP-1ß, TNF-α, NOS2) were depressed by 15d-PGJ2 in macrophages isolated from treated livers. In vitro, 15d-PGJ2 inhibited BMM activation via PPARγ. Moreover, miR-27b-3p, miR-181a-1-3p, and miR-326-5p target MIP-1ß, TNF-α, and NOS2 mRNA, respectively. The miRNA expressions were decreased in damaged livers, macrophages isolated from injured livers, and activated BMMs, which were renewed by 15d-PGJ2/PPARγ axis. In activated BMMs, the miRNA inhibitors attenuated inhibitory effect of PPARγ agonist (troglitazone or ciglitazone), while replenishing the lack of miRNAs induced by PPARγ deficiency using miRNA mimics caused a decline of inflammatory cytokines. In conclusion, these data suggest that 15d-PGJ2/PPARγ axis regulates BMM activation via promoting miR-27b-3p, miR-181a-1-3p, and miR-326-5p expressions. KEY MESSAGES: 15d-PGJ2 inhibits BMM activation via PPARγ activation. 15d-PGJ2/PPARγ axis promotes expression of miR-27b-3p, miR-181a-1-3p, and miR-326-5p. miR-27b-3p, miR-181a-1-3p, and miR-326-5p have an inhibitory effect on BMM activation via 15d-PGJ2/PPARγ axis.

Niacin ameliorates ulcerative colitis via prostaglandin D2-mediated D prostanoid receptor 1 activation.

Niacin, as an antidyslipidemic drug, elicits a strong flushing response by release of prostaglandin (PG) D2 However, whether niacin is beneficial for inflammatory bowel disease (IBD) remains unclear. Here, we observed niacin administration-enhanced PGD2 production in colon tissues in dextran sulfate sodium (DSS)-challenged mice, and protected mice against DSS or 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis in D prostanoid receptor 1 (DP1)-dependent manner. Specific ablation of DP1 receptor in vascular endothelial cells, colonic epithelium, and myeloid cells augmented DSS/TNBS-induced colitis in mice through increasing vascular permeability, promoting apoptosis of epithelial cells, and stimulating pro-inflammatory cytokine secretion of macrophages, respectively. Niacin treatment improved vascular permeability, reduced apoptotic epithelial cells, promoted epithelial cell update, and suppressed pro-inflammatory gene expression of macrophages. Moreover, treatment with niacin-containing retention enema effectively promoted UC clinical remission and mucosal healing in patients with moderately active disease. Therefore, niacin displayed multiple beneficial effects on DSS/TNBS-induced colitis in mice by activation of PGD2/DP1 axis. The potential efficacy of niacin in management of IBD warrants further investigation.

Eosinophil production of prostaglandin D2 in patients with aspirin-exacerbated respiratory disease.

BACKGROUND: Aspirin-exacerbated respiratory disease (AERD) differs from aspirin-tolerant disease in part because of eosinophilic tissue infiltration and overexpression of arachidonic acid metabolic pathway components that lead to enhanced secretion of cysteinyl leukotrienes and prostaglandin (PG) D2 observed constitutively and paradoxically in response to aspirin and other COX inhibitors. We have previously demonstrated the capacity of IFN-γ to drive cysteinyl leukotriene expression and response. OBJECTIVE: We investigated eosinophils as a source of PGD2 production in patients with AERD. METHODS: Eosinophils were enriched from tissue and peripheral blood obtained from control subjects, patients with aspirin-tolerant disease, and patients with AERD. mRNA was extracted and evaluated for expression of hematopoietic prostaglandin D synthase (hPGDS). Expression of hPGDS protein was confirmed with Western hybridization and immunofluorescence staining. Cells were stimulated with aspirin, and secretion of PGD2 was quantified. CD34+ progenitor cells were isolated and matured into eosinophils in the presence or absence of IFN-γ and hPGDS mRNA, and PGD2 release was measured. RESULTS: Gene expression analysis revealed that eosinophils from tissue and blood of patients with AERD display increased levels of hPGDS compared with asthmatic and control samples. Western hybridization confirmed the increase in hPGDS mRNA translated to increased protein expression. Immunofluorescence confirmed mast cells and eosinophils from tissue of patients with AERD and asthma demonstrated hPGDS expression, with higher levels in eosinophils from patients with AERD. Incubation of eosinophils from blood and tissue with aspirin stimulated PGD2 release. IFN-γ-matured eosinophil progenitors showed enhanced hPGDS expression and increased levels of PGD2 release at baseline and after aspirin stimulation. CONCLUSIONS: In addition to mast cells, eosinophils represent an important source of PGD2 in patients with AERD and identify a new target for therapeutic intervention.

Btk Inhibitor RN983 Delivered by Dry Powder Nose-only Aerosol Inhalation Inhibits Bronchoconstriction and Pulmonary Inflammation in the Ovalbumin Allergic Mouse Model of Asthma.

BACKGROUND: In allergen-induced asthma, activated mast cells start the lung inflammatory process with degranulation, cytokine synthesis, and mediator release. Bruton's tyrosine kinase (Btk) activity is required for the mast cell activation during IgE-mediated secretion. METHODS: This study characterized a novel inhaled Btk inhibitor RN983 in vitro and in ovalbumin allergic mouse models of the early (EAR) and late (LAR) asthmatic response. RESULTS: RN983 potently, selectively, and reversibly inhibited the Btk enzyme. RN983 displayed functional activities in human cell-based assays in multiple cell types, inhibiting IgG production in B-cells with an IC50 of 2.5 ± 0.7 nM and PGD2 production from mast cells with an IC50 of 8.3 ± 1.1 nM. RN983 displayed similar functional activities in the allergic mouse model of asthma when delivered as a dry powder aerosol by nose-only inhalation. RN983 was less potent at inhibiting bronchoconstriction (IC50(RN983) = 59 µg/kg) than the ß-agonist salbutamol (IC50(salbutamol) = 15 µg/kg) in the mouse model of the EAR. RN983 was more potent at inhibiting the antigen induced increase in pulmonary inflammation (IC50(RN983) = <3 µg/kg) than the inhaled corticosteroid budesonide (IC50(budesonide) = 27 µg/kg) in the mouse model of the LAR. CONCLUSIONS: Inhalation of aerosolized RN983 may be effective as a stand-alone asthma therapy or used in combination with inhaled steroids and ß-agonists in severe asthmatics due to its potent inhibition of mast cell activation.

The low molecular weight fraction of human serum albumin upregulates production of 15d-PGJ2 in Peripheral Blood Mononuclear Cells.

Activation of the innate immune system involves a series of events designed to counteract the initial insult followed by the clearance of debris and promotion of healing. Aberrant regulation can lead to systemic inflammatory response syndrome, multiple organ failure, and chronic inflammation. A better understanding of the innate immune response may help manage complications while allowing for proper immune progression. In this study, the ability of several classes of anti-inflammatory drugs to affect LPS-induced cytokine and prostaglandin release from peripheral blood mononuclear cells (PBMC) was evaluated. PBMC were cultured in the presence of dexamethasone (DEX), ibuprofen (IBU), and the low molecular weight fraction of 5% albumin (LMWF5A) followed by stimulation with LPS. After 24 h, TNFα, PGE2, and 15d-PGJ2 release was determined by ELISA. Distinct immunomodulation patterns emerged following LPS stimulation of PBMC in the presence of said compounds. DEX, a steroid with strong immunosuppressive properties, reduced TNFα, PGE2, and 15d-PGJ2 release. IBU caused significant reduction in prostaglandin release while TNFα release was unchanged. An emerging biologic with known anti-inflammatory properties, LMWF5A, significantly reduced TNFα release while enhancing PGE2 and 15d-PGJ2 release. Incubating LMWF5A together with IBU negated this observed increased prostaglandin release without affecting the suppression of TNFα release. Additionally, LMWF5A caused an increase in COX-2 transcription and translation. LMWF5A exhibited a unique immune modulation pattern in PBMC, disparate from steroid or NSAID administration. This enhancement of prostaglandin release (specifically 15d-PGJ2), in conjunction with a decrease in TNFα release, suggests a switch that favors resolution and decreased inflammation.

Resveratrol preferentially inhibits IgE-dependent PGD2 biosynthesis but enhances TNF production from human skin mast cells.

BACKGROUND: Resveratrol, a natural polyphenol found in the skin of red grapes, is reported to have anti-inflammatory properties including protective effects against aging. Consequently, Resveratrol is a common nutritional supplement and additive in non-prescription lotions and creams marketed as anti-aging products. Studies in mice and with mouse bone marrow-derived mast cells (BMMCs) have indicated anti-allergic effects of Resveratrol. However, the effects of Resveratrol on human primary mast cells have not been reported. METHODS: Human mast cells were isolated and purified from normal skin tissue of different donors. The effect of Resveratrol on IgE-dependent release of allergic inflammatory mediators was determined using various immunoassays, Western blotting, and quantitative real-time PCR. RESULTS: Resveratrol at low concentrations (≤10 µM) inhibited PGD2 biosynthesis but not degranulation. Accordingly, COX-2 expression was inhibited but phosphorylation of Syk, Akt, p38, and p42/44 (ERKs) remained intact. Surprisingly, TNF production was significantly enhanced with Resveratrol. At a high concentration (100 µM), Resveratrol significantly inhibited all parameters analyzed except Syk phosphorylation. CONCLUSIONS: Here, we show that Resveratrol at low concentrations exerts its anti-inflammatory properties by preferentially targeting the arachidonic acid pathway. We also demonstrate a previously unrecognized pro-inflammatory effect of Resveratrol--the enhancement of TNF production from human mature mast cells following IgE-dependent activation. GENERAL SIGNIFICANCE: These findings suggest that Resveratrol as a therapeutic agent could inhibit PGD2-mediated inflammation but would be ineffective against histamine-mediated allergic reactions. However, Resveratrol could potentially exacerbate or promote allergic inflammation by enhancing IgE-dependent TNF production from mast cells in human skin.

Thymic stromal lymphopoietin controls prostaglandin D2 generation in patients with aspirin-exacerbated respiratory disease.

BACKGROUND: Prostaglandin (PG) D2 is the dominant COX product of mast cells and is an effector of aspirin-induced respiratory reactions in patients with aspirin-exacerbated respiratory disease (AERD). OBJECTIVE: We evaluated the role of the innate cytokine thymic stromal lymphopoietin (TSLP) acting on mast cells to generate PGD2 and facilitate tissue eosinophilia and nasal polyposis in patients with AERD. METHODS: Urinary eicosanoid levels were measured in aspirin-tolerant control subjects and patients with AERD. Nasal polyp specimens from patients with AERD and chronic rhinosinusitis were analyzed by using quantitative PCR, Western blotting, and immunohistochemistry. Human cord blood-and peripheral blood-derived mast cells were stimulated with TSLP in vitro to assess PGD2 generation. RESULTS: Urinary levels of a stable PGD2 metabolite (uPGD-M) were 2-fold higher in patients with AERD relative to those in control subjects and increased further during aspirin-induced reactions. Peak uPGD-M levels during aspirin reactions correlated with reductions in blood eosinophil counts and lung function and increases in nasal congestion. Mast cells sorted from nasal polyps expressed PGD2 synthase (hematopoietic PGD2 synthase) mRNA at higher levels than did eosinophils from the same tissue. Whole nasal polyp TSLP mRNA expression correlated strongly with mRNA encoding hematopoietic PGD2 synthase (r = .75), the mast cell-specific marker carboxypeptidase A3 (r = .74), and uPGD-M (r = 0.74). Levels of the cleaved active form of TSLP were increased in nasal polyps from patients with AERD relative to those in aspirin-tolerant control subjects. Recombinant TSLP induced PGD2 generation by cultured human mast cells. CONCLUSIONS: Our study demonstrates that mast cell-derived PGD2 is a major effector of type 2 immune responses driven by TSLP and suggests that dysregulation of this innate system contributes significantly to the pathophysiology of AERD.

BDNF and NGF Signalling in Early Phases of Psychosis: Relationship With Inflammation and Response to Antipsychotics After 1 Year.

Previous studies have indicated systemic deregulation of the proinflammatory or anti-inflammatory balance in individuals with first-episode psychosis (FEP) that persists 12 months later. To identify potential risk/protective factors and associations with symptom severity, we assessed possible changes in plasma levels of neurotrophins (brain-derived neurotrophic factor [BDNF] and nerve growth factor [NGF]) and their receptors in peripheral blood mononuclear cells (PBMCs). Expression of the 2 forms of BDNF receptors (active TrkB-FL and inactiveTrkB-T1) in PBMCs of FEP patients changed over time, TrkB-FL expression increasing by 1 year after diagnosis, while TrkB-T1 expression decreased. The TrkB-FL/TrkB-T1 ratio (hereafter FL/T1 ratio) increased during follow-up in the nonaffective psychosis group only, suggesting different underlying pathophysiological mechanisms in subgroups of FEP patients. Further, the expression of the main NGF receptor, TrkA, generally increased in patients at follow-up. After adjusting for potential confounders, baseline levels of inducible isoforms of nitric oxide synthase, cyclooxygenase, and nuclear transcription factor were significantly associated with the FL/T1 ratio, suggesting that more inflammation is associated with higher values of this ratio. Interestingly, the FL/T1 ratio might have a role as a predictor of functioning, a regression model of functioning at 1 year suggesting that the effect of the FL/T1 ratio at baseline on functioning at 1 year depended on whether patients were treated with antipsychotics. These findings may have translational relevance; specifically, it might be useful to assess the expression of TrkB receptor isoforms before initiating antipsychotic treatment in FEPs.