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We previously found that ribosomal protein L9 (RPL9) is a novel advanced glycation end product (AGE)‐binding protein that can decrease pro‐inflammatory TNF‐α expression stimulated by lipopolysaccharide (LPS) plus high‐mobility group box 1 (HMGB1), suggesting that RPL9 has a role in regulating LPS+HMGB1‐stimulated inflammatory reactions. Among the various ribosomal proteins, it was found that RPS5 reproduced the regulatory activity of RPL9 on LPS+HMGB1‐stimulated TNF‐α expression in macrophage‐like RAW264.7 cells. RPL9 and RPS5 share a common feature as cationic proteins. Polylysine, a cationic polypeptide, and a synthetic peptide of the cationic region from RPL9 also exhibited reducing activity on LPS+HMGB1‐induced TNF‐α expression. By pull‐down assay, RPL9 and RPS5 were confirmed to interact with AGEs. When AGEs coexisted with LPS, HMGB1, plus RPL9 or RPS5, the reducing effect of TNF‐α expression by these cationic ribosomal proteins was shown to be abrogated. The results suggest that cationic ribosomal proteins have a regulatory role in the pro‐inflammatory response induced by LPS+HMGB1, and in the pathophysiological condition of accumulating AGEs, this regulatory effect is abolished, which exacerbates inflammation. [ABSTRACT FROM AUTHOR]

Advanced glycation end products (AGEs) are considered to be related to the pathogenesis of some inflammatory diseases. AGEs were reported to stimulate the receptor for AGEs (RAGE), which causes inflammatory reactions. However, recently, toll‐like receptors (TLRs), in addition to RAGE, have been reported to be related to AGE‐mediated cellular responses, and it remains unclear which receptor is responsible for AGE recognition. To reveal the role of pattern‐recognition receptors, including TLRs and/or RAGE, in AGE‐mediated cellular responses, we generated macrophage‐like RAW264.7 knockout (KO) cells lacking these receptors by genome editing using the CRISPR/Cas9 system and assessed AGE‐stimulated changes in these cells. Comparison of the established clones suggested that RAGE partially affects the expression of TLRs. In the KO clone lacking TLR4 and TLR2, AGE‐stimulated tumor necrosis factor alpha (TNF‐α) expression and phosphorylation of IκBα, p38, and extracellular signal‐regulated kinase (ERK) were significantly attenuated, suggesting that AGE‐mediated responses are largely dependent on TLRs. On the other hand, on comparison of the AGE‐stimulated responses between the KO clone lacking TLR4 and TLR2, and the clone lacking TLR4, TLR2, and RAGE, RAGE played little role in AGE‐stimulated TNF‐α transcription and ERK phosphorylation. Taken together, this study suggested that AGE‐stimulated inflammatory responses occur mainly through TLRs rather than RAGE. [ABSTRACT FROM AUTHOR]

Intrathecal treatment with recombinant high‐mobility group box‐1 (rHMGB1) in naïve mice leads to a persistent and significantly decreased hind paw withdrawal threshold to mechanical stimuli, suggesting that spinal HMGB1 evokes abnormal pain processing. By contrast, repeated intrathecal treatment with anti‐HMGB1 antibody significantly reverses hind paw mechano‐hypersensitivity in mice with a partial sciatic nerve ligation (PSNL). By contrast, the cellular mechanism by which spinal HMGB1 induces neuropathic pain has yet to be fully elaborated. The current study tested the hypothesis that spinal HMGB1 could induce mechanical hypersensitivity through the activation of specific receptor in glial cells. Intrathecal pretreatment with toll‐like receptor (TLR) 4 inhibitors, but not TLR5, receptor for advanced glycation end‐products and C‐X‐C chemokine receptor type 4 inhibitors, prevented rHMGB1‐evoked mechanical hypersensitivity. Activation of spinal astrocytes appears to be crucial for the mechanism of action of rHMGB1 in naïve mice, as intrathecal pretreatment with astrocytic inhibitors prevented the rHMGB1‐induced mechanical hypersensitivity. Interleukin‐1β (IL‐1β) was up‐regulated within activated astrocytes and block of TLR4 prevented the upregulation of IL‐1β. Interleukin‐1β appears to be secreted by activated astrocytes, as IL‐1β neutralizing antibody prevented rHMGB1‐induced mechanical hypersensitivity. Furthermore, intrathecal pretreatment with either MK801 or gabapentin prevented the rHMGB1‐induced mechanical hypersensitivity, suggesting roles for spinal glutamate and the N‐methyl‐d‐aspartate receptor in the mediation of rHMGB1‐induced mechanical hypersensitivity. Thus, the current findings suggest that spinal HMGB1 upregulates IL‐1β in spinal astrocytes through a TLR4‐dependent pathway and increases glutamatergic nociceptive transduction. These spinal mechanisms could be key steps that maintain neuropathic pain. [ABSTRACT FROM AUTHOR]

Background and Purpose: Microvascular barrier breakdown is a hallmark of sepsis that is associated with sepsis‐induced multiorgan failure. Histidine‐rich glycoprotein (HRG) is a 75‐kDa plasma protein that was demonstrated to improve the survival of septic mice through regulation of cell shape, spontaneous ROS production in neutrophils, and adhesion of neutrophils to vascular endothelial cells. We investigated HRG's role in the LPS/TNF‐α‐induced barrier dysfunction of endothelial cells in vitro and in vivo and the possible mechanism, to clarify the definitive roles of HRG in sepsis. Experimental Approach EA.hy 926 endothelial cells were pretreated with HRG or human serum albumin before stimulation with LPS/TNF‐α. A variety of biochemical assays were applied to explore the underlying molecular mechanisms on how HRG protected the barrier function of vascular endothelium. Key Results: Immunostaining results showed that HRG maintains the endothelial monolayer integrity by inhibiting cytoskeleton reorganization, losses of VE‐cadherin and β‐catenin, focal adhesion kinase degradation, and cell detachment induced by LPS/TNF‐α. HRG also inhibited the cytokine secretion from endothelial cells induced by LPS/TNF‐α, which was associated with reduced NF‐κB activation. Moreover, HRG effectively prevented the LPS/TNF‐α‐induced increase in capillary permeability in vitro and in vivo. Finally, Western blot results demonstrated that HRG prevented the phosphorylation of MAPK family and RhoA activation, which are involved mainly in the regulation of cytoskeleton reorganization and barrier permeability. Conclusions and Implications: Taken together, our results demonstrate that HRG has protective effects on vascular barrier function in vitro and in vivo, which may be due to the inhibition of MAPK family and Rho activation. [ABSTRACT FROM AUTHOR]

Human pandemic H1N1 2009 influenza virus causes significant morbidity and mortality with severe acute lung injury due to the excessive inflammatory reaction, even with neuraminidase inhibitor use. The anti‐inflammatory effect of anti‐high‐mobility group box‐1 (HMGB1) monoclonal antibody (mAb) against influenza pneumonia has been reported. In this study, we evaluated the combined effect of anti‐HMGB1 mAb and peramivir against pneumonia induced by influenza A (H1N1) virus in mice. Nine‐week‐old male C57BL/6 mice were inoculated with H1N1 and treated with intramuscularly administered peramivir at 2 and 3 days post‐infection (dpi). The anti‐HMGB1 mAb or a control mAb was administered at 2, 3, and 4 dpi. Survival rates were assessed, and lung lavage and pathological analyses were conducted at 5 and 7 dpi. The combination of peramivir with the anti‐HMGB1 mAb significantly improved survival rate whereas the anti‐HMGB1 mAb alone did not affect virus proliferation in the lungs. This combination therapy also significantly ameliorated histopathological changes, neutrophil infiltration, and macrophage aggregation by inhibiting HMGB1, inflammatory cytokines, and oxidative stress. Fluorescence immunostaining showed that the anti‐HMGB1 mAb inhibited HMGB1 translocation from type I alveolar epithelial cells. In summary, combining anti‐HMGB1 with conventional anti‐influenza therapy might be useful against severe influenza virus infection. ‐Combined therapy of anti‐HMGB1 monoclonal antibody and peramivir significantly improved the survival rate of influenza‐induced pneumonia model mice.‐Combined therapy of anti‐HMGB1 monoclonal antibody and peramivir also significantly ameliorated histopathological changes, neutrophil infiltration, and macrophage aggregation by inhibiting HMGB1 translocation from type I alveolar epithelial cells, leading to the inhibition of TLR4 signaling, cytokine/chemokine responses, and oxidative stress.‐Anti‐HMGB1 therapy in combination with an anti‐influenza drug may be an option for severe influenza pneumonia. [ABSTRACT FROM AUTHOR]

Previously, we found that endogenously produced pro‐inflammatory molecules, advanced glycation end products (AGEs), interact with tumor necrosis factor‐like weak inducer of apoptosis (TWEAK), and attenuate its immunomodulatory function. In the present study, to elucidate the mechanism by which AGEs attenuate TWEAK function, we searched for regions responsible for TWEAK–AGE interaction using TWEAK deletion mutants. Pull‐down assays with the TWEAK mutants and AGEs revealed that the C‐terminal half of TWEAK, which is the region essential for receptor stimulation, was required for this interaction. On the other hand, the N‐terminal deletion mutants did not exhibit a significant decrease in AGE binding. Moreover, a moderate decrease in the AGE binding by double‐deletion in quartered C‐terminal half regions and a substantial decrease by triple‐deletion in this region were observed. In addition, full‐length TWEAK stimulated IL‐8 gene expression in endothelial EA.hy.926 cells, whereas the triple‐deletion mutant lost much of this activity, suggesting that the TWEAK–AGE interaction sites overlap with the region needed to exert normal function of TWEAK. Our present findings may help to elucidate the pathophysiological roles of the TWEAK–AGE interaction for prevention and treatment of AGE‐related inflammatory diseases. [ABSTRACT FROM AUTHOR]

Encephalopathy is a major cause of influenza‐associated child death and severe neurological sequelae in Japan, highlighting the urgent need for new therapeutic strategies. In this study, we evaluated the effects of anti‐high mobility group box‐1 monoclonal antibody (α‐HMGB1) treatment on brain edema induced by influenza A virus (IAV) and lipopolysaccharide in 4‐week‐old BALB/c female mice. The results showed that administration of 7.5 mg/kg α‐HMGB1 1 h after IAV (A/Puerto Rico/8/34) inoculation significantly alleviated brain edema at 48 h after IAV inoculation, as confirmed by the suppression of Evans Blue dye leakage and matrix metallopeptidase‐9 mRNA expression in the brain. Moreover, we also observed suppression of oxidative stress and different cytokines in IAV‐inoculated mice. The expression of plasminogen activator inhibitor‐1 was also attenuated following treatment with α‐HMGB1. Notably, α‐HMGB1 treatment had no effect on virus propagation in the lung. In summary, anti‐HMGB1 treatment may improve the prognosis in cases with influenza‐associated encephalopathy by attenuating brain edema and reducing the inflammatory responses induced by HMGB1. [ABSTRACT FROM AUTHOR]

High-mobility group box-1 (HMGB1) has been shown to be critical in the modulation of nociceptive transduction following a peripheral neuropathy. However, the precise role of peripherally expressed HMGB1 in neuropathic pain has yet to be fully elaborated. Following a partial sciatic nerve ligation (PSNL) in mice, a persistent ipsilateral up-regulation of HMGB1 was observed from 3 to 21 days after PSNL, in paralleled with a robust ipsilateral hind paw mechanical hypersensitivity. Increased HMGB1 was detected in both infiltrating macrophages and proliferating Schwann cells in the ipsilateral nerve 14 days following PSNL. Repeated perineural treatment with anti-HMGB1 antibody significantly ameliorated PSNL-induced mechanical hypersensitivity. Several pronociceptive molecules, including matrix metalloprotease-9 (MMP-9), tumor necrosis factor-α, interleukin-1β (IL-1β), and cyclooxygenase-2, were up-regulated in injured sciatic nerve 14 days following PSNL. Repeated perineural treatment with an anti-HMGB1 antibody significantly suppressed expression of MMP-9, but not other pronociceptive molecules. Perineural treatment with a selective MMP-9 inhibitor ameliorated PSNL-induced mechanical hypersensitivity. The current findings demonstrate that the maintenance of the neuropathic state following an injured nerve is dependent on the up-regulation of HMGB1 and MMP-9. Thus, blocking HMGB1 function in sciatic nerve could be a potent therapeutic strategy for the treatment of neuropathic pain. [ABSTRACT FROM AUTHOR]

Mast cells are the major sources of histamine, which is released in response to immunological stimulations. The synthesis of histamine is catalyzed by histidine decarboxylase ( HDC). Previous studies have shown that Hdc−/− mast cells exhibit aberrant granule morphology with severely decreased granule content. Here, we investigated whether the histamine synthesized in mast cells regulates the granule maturation of murine mast cells. Several genes, including those encoding granule proteases and enzymes involved in heparin biosynthesis, were downregulated in Hdc−/− peritoneal mast cells. Impaired granule maturation was also found in Hdc−/− BM-derived cultured mast cells when they were cocultured with fibroblasts in the presence of c-kit ligand. Exogenous application of histamine and several H4 receptor agonists restored the granule maturation of Hdc−/− cultured mast cells. However, the maturation of granules was largely normal in Hrh4−/− peritoneal mast cells. Depletion of cellular histamine with tetrabenazine, an inhibitor of vesicular monoamine transporter-2, did not affect granule maturation. In vivo experiments with mast cell deficient KitW/ KitW-v mice indicated that the expression of the Hdc gene in mast cells is required for granule maturation. These results suggest that histamine promotes granule maturation in mast cells and acts as an proinflammatory mediator. [ABSTRACT FROM AUTHOR]

Objective: High mobility group box-1 (HMGB1) plays an important role in triggering inflammatory responses in many types of diseases. In this study, we examined the involvement of HMGB1 in traumatic brain injury (TBI) and evaluated the ability of intravenously administered neutralizing anti-HMGB1 monoclonal antibody (mAb) to attenuate brain injury. Methods: Traumatic brain injury was induced in rats or mice by fluid percussion. Anti-HMGB1 mAb or control mAb was administered intravenously after TBI. Results: Anti-HMGB1 mAb remarkably inhibited fluid percussion-induced brain edema in rats, as detected by T2-weighted magnetic resonance imaging; this was associated with inhibition of HMGB1 translocation, protection of blood-brain barrier (BBB) integrity, suppression of inflammatory molecule expression, and improvement of motor function. In contrast, intravenous injection of recombinant HMGB1 dose-dependently produced the opposite effects. Experiments using receptor for advanced glycation end product (RAGE)−/−, toll-like receptor-4 (TLR4)−/−, and TLR2−/− mice suggested the involvement of RAGE as the predominant receptor for HMGB1. Interpretation: Anti-HMGB1 mAb may provide a novel and effective therapy for TBI by protecting against BBB disruption and reducing the inflammatory responses induced by HMGB1. ANN NEUROL 2012;72:373-384 [ABSTRACT FROM AUTHOR]

Inflammatory reactions play an important role in ischaemia/reperfusion injury in various organs. Since histamine H4 action has been shown to prevent the development of ischaemia/reperfusion liver injury, we examined the effects of dimaprit, a histamine H2/H4 receptor agonist, on ischaemia-induced cytokine release and liver damage. Male Wistar rats (300 g) were subjected to warm ischaemia for 30 min. by occlusion of the left portal vein and hepatic artery under halothane anaesthesia. Saline or dimaprit (20 mg/kg, subcutaneously) was injected immediately after reperfusion of blood flow. Transient ischaemia provoked severe liver damage 24 hr after reperfusion, and the plasma concentrations of alanine transaminase and aspartate transaminase were 4600 IU/l and 13,200 IU/l, respectively. The values in the dimaprit group were 55% and 46% of those in control animals, respectively. Dimaprit also reduced the infarct size to 50%. Liver ischaemia markedly increased interleukin-12 levels 2–24 hr after reperfusion. The dimaprit treatment depressed the values to 40–64% of those in the corresponding control group 4–24 hr after reperfusion. Since interleukin-12 facilitates cell-mediated cytotoxicity, the protective effect of dimaprit may be attributed to regulation of cytokine release during reperfusion. [ABSTRACT FROM AUTHOR]

The high mobility group box-1 (HMGB1), originally identified as an architectural nuclear protein, exhibits an inflammatory cytokine-like activity in the extracellular space. Here we show that treatment with neutralizing anti-HMGB1 monoclonal antibody (mAb; 200 µg, twice) remarkably ameliorated brain infarction induced by 2-h occlusion of the middle cerebral artery in rats, even when the mAb was administered after the start of reperfusion. Consistent with the 90% reduction in infarct size, the accompanying neurological deficits in locomotor function were significantly improved. Anti-HMGB1 mAb inhibited the increased permeability of the blood-brain barrier, the activation of microglia, the expression of TNF-α and iNOS, and suppressed the activity of MMP-9, whereas it had little effect on blood flow. Intracerebroventricular injection of HMGB1 increased the severity of infarction. Immunohistochemical study revealed that HMGB1 immunoreactivity in the cell nuclei decreased or disappeared in the affected areas, suggesting the release of HMGB1 into the extracellular space. These results indicate that HMGB1 plays a critical role in the development of brain infarction through the amplification of plural inflammatory responses in the ischemic region and could be an outstandingly suitable target for the treatment. Intravenous injection of neutralizing anti-HMGB1 mAb provides a novel therapeutic strategy for ischemic stroke. [ABSTRACT FROM AUTHOR]

Histamine not only mediates immediate allergic reactions, it also regulates cellular immune responses. H4R is the most recently identified histamine receptor (HR). In the present study, we examined the in vitro effect of histamine and H4R agonists on the responses of human T cells to purified protein derivative from Mycobacterium tuberculosis (PPD) and to Cry j1, the major allergen of Cryptomeria japonica pollen. Dimaprit, clobenpropit and clozapine, which are H4R agonists, dose-dependently blocked both PPD-induced interferon-γ and Cry j1-induced interleukin-5 production by both peripheral blood mononuclear cells (PBMCs) and antigen-specific T-cell lines. However, the addition of thioperamide, an H3R/H4R antagonist, as well as a mixture of d-chlropheniramine, famotidine and thioperamide, did not reverse the inhibition. Pretreatment of PBMCs with SQ22536 and 8-bromoadenosine-3′,5′-cyclic monophosphorothioate, Rp-isomer, had varying abilities to reverse the inhibitory effects of H4R agonists, except for clobenpropit. Moreover, the addition of H4R agonists induced annexin-V expression on PBMCs, especially in CD19+ and CD4+ cells. cDNA microarray analysis revealed that, among 16 600 genes tested, increased expression following treatment with clozapine was seen in 0·8% of the genes, whereas decreased expression was seen in 3·0% of the genes. These results suggest that H4R agonists inhibit antigen-specific human T-cell responses, although H4R does not appear to be important for this effect. In addition, the present study indicated that there may be orphan receptors or HR subtypes which can bind dimaprit, clobenpropit and clozapine, and that can exert an inhibitory effect on antigen-specific cellular responses via a cAMP/cAMP-dependent protein kinase-dependent, apoptotic pathway. [ABSTRACT FROM AUTHOR]

Prostaglandin E2 (PGE2) is a lipid mediator that displays important immunomodulatory properties, such as polarization of cytokine production by T cells. Recent investigations have revealed that the effect of PGE2 on cytokine production is greatly influenced by external stimuli; however, it is unclear whether PGE2 plays a significant role in major histocompatibility complex-mediated antigen-specific T-cell responses via binding to one of four subtypes of E prostanoid (EP) receptor alone or in combination. In the present study, we sought to determine the effect of PGE2 on antigen-specific CD4+ T-cell responses in humans, especially in terms of receptor specificity. We used purified protein derivative (PPD) and Cry j 1 as T helper type 1 (Th1) and Th2-inducing antigens, respectively. We generated several different Cry j 1- and PPD-specific T-cell lines (TCLs). PGE2 significantly and dose-dependently inhibited the proliferation and subsequent production of interleukin-4 by Cry j 1-specific TCLs and of interferon-γ by PPD-specific TCLs upon antigen stimulation. Administration of EP2 receptor agonist and EP4 receptor agonist suppressed these responses in an adenylate cyclase-dependent manner, while EP1 and EP3 receptor agonists did not. Messenger RNA for EP2, EP3 and EP4, but not EP1, receptors were detected in Cry j 1- and PPD-specific TCLs, and no differences in EP receptor expression were observed between them. Furthermore, PGE2 and EP2 receptor agonist significantly inhibited interleukin-5 and interferon-γ production by peripheral blood mononuclear cells in response to Cry j 1 and PPD stimulation, respectively. These results suggest that PGE2 suppresses both Th1- and Th2-polarized antigen-specific human T-cell responses via a cAMP-dependent EP2/EP4-mediated pathway. [ABSTRACT FROM AUTHOR]

Abstract: A trypsin-like serine proteinase was purified from the incubation medium of rat brain slices by gelatin zymography. The purification consisted of ammonium sulfate precipitation, benzamidine-Sepharose 6B affinity chromatography, and carboxymethyl-cellulose and gel filtration chromatographies. The gelatinolytic activity, identified at 22 kDa (P22) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under nonreducing conditions, was eluted as one active peak throughout the purification, and the final preparation gave a single protein peak on reverse-phase HPLC. Diisopropyl fluorophosphate, benzamidine, p-toluenesulfonyl-L-lysine chloromethyl ketone, and aprotinin completely inhibited the activity of P22, whereas phenanthroline, p-toluene-sulfonyl-L-phenylalanine chloromethyl ketone, and elastinal did not. P22 efficiently digested the extracellular matrix proteins laminin and type IV collagen. P22 produced an increase in intracellular Ca[sup 2+] concentration in A172 glioblastoma, which was desensitized through prior stimulation with protease-activated receptor-2 agonist peptide SLIGKV, indicating that P22 can stimulate protease-activated receptor-2. Rat brain penetration injury induced gelatinolytic activity in the lesioned area whose molecular size was consistent with that of P22. These results indicated that on incubation of rat brain slices, a trypsin-like serine proteinase was secreted into the medium that was capable of digesting extracellular matrix and stimulating protease-activated receptor-2. It is suggested that the gelatinolytic activity induced by brain injury might be that of P22... [ABSTRACT FROM AUTHOR]

Abstract Histamine N-methyltransferase (HMT) (EC 2.1.1.8) plays a crucial role in the inactivation of the neurotransmitter histamine in the CNS. However, the localization of HMT remains to be determined. In the present study, we investigated immunohistochemical localization of HMT in the bovine CNS using a polyclonal antibody against bovine HMT. The HMT-like immunoreactivity was observed mainly in neurons. Strongly immunoreactive neurons were present in the oculomotor nucleus and ruber nucleus in the midbrain, the facial nucleus in the pons, the dorsal vagal nucleus and hypoglossal nucleus in the medulla oblongata and in the anterior horn as well as intermediolateral zone of the spinal cord. Intermediately immunoreactive neurons were present in the piriform cortex and the inferior olivary nucleus. The grey matter of the forebrain regions was diffusely and faintly stained. In the cerebellum and the striatum, the nerve fibres in the white matter were positive. The tuberomammillary nucleus, where histaminergic neurons are present, were weakly positive. The other immunoreactive structures in the CNS were blood vessels. Almost all of the blood vessel walls, irrespective of whether they were arterial or venous, were variably stained. The glial fibrillary acidic protein- (GFAP-) immunoreactive astrocytes were not stained. These findings indicated that histamine released from histaminergic nerve terminals or varicose fibres is methylated mainly in postsynaptic or extrasynaptic neurons rather than in astrocytes. The localization of HMT in the blood vessel wall may mean that blood-borne histamine and histamine released from mast cells associated with the blood vessels are catabolized in this structure. [ABSTRACT FROM AUTHOR]

Prostaglandin E2 (PGE2) is a lipid mediator that displays important immunomodulatory properties, such as polarization of cytokine production by T cells. Recent investigations have revealed that the effect of PGE2 on cytokine production is greatly influenced by external stimuli; however, it is unclear whether PGE2 plays a significant role in major histocompatibility complex-mediated antigen-specific T-cell responses via binding to one of four subtypes of E prostanoid (EP) receptor alone or in combination. In the present study, we sought to determine the effect of PGE2 on antigen-specific CD4+ T-cell responses in humans, especially in terms of receptor specificity. We used purified protein derivative (PPD) and Cry j 1 as T helper type 1 (Th1) and Th2-inducing antigens, respectively. We generated several different Cry j 1- and PPD-specific T-cell lines (TCLs). PGE2 significantly and dose-dependently inhibited the proliferation and subsequent production of interleukin-4 by Cry j 1-specific TCLs and of interferon-γ by PPD-specific TCLs upon antigen stimulation. Administration of EP2 receptor agonist and EP4 receptor agonist suppressed these responses in an adenylate cyclase-dependent manner, while EP1 and EP3 receptor agonists did not. Messenger RNA for EP2, EP3 and EP4, but not EP1, receptors were detected in Cry j 1- and PPD-specific TCLs, and no differences in EP receptor expression were observed between them. Furthermore, PGE2 and EP2 receptor agonist significantly inhibited interleukin-5 and interferon-γ production by peripheral blood mononuclear cells in response to Cry j 1 and PPD stimulation, respectively. These results suggest that PGE2 suppresses both Th1- and Th2-polarized antigen-specific human T-cell responses via a cAMP-dependent EP2/EP4-mediated pathway. [ABSTRACT FROM AUTHOR]