Leukotriene D4 accelerates antigen-mediated mast cell responses via the cysteinyl leukotriene 1 receptor.

Cysteinyl leukotrienes (CysLTs), released from mast cells (MCs), are important mediators in allergy. Type 1 receptors for CysLTs (CysLT1R) are involved in accelerating IgE-mediated MC activation. In this study, we aimed to elucidate the mechanisms underlying CysLT1R-mediated MC activation. The CysLT1R agonist/antagonist was applied to two types of major MC models-RBL-2H3 cells and bone marrow-derived MCs (BMMCs). The use of CysLT1R and CysLT2R inhibitors revealed that CysLT1R plays a major role in the acceleration of MC activation. The administration of the CysLT1R agonist leukotriene D4 upregulated IgE-mediated Akt and ERK phosphorylation and subsequently enhanced TNF-α expression, suggesting that CysLT1R regulates the downstream pathway of MC activation. However, these observations were not corroborated by CysLT1R knockdown using shRNA, suggesting a differential regulatory mechanism between the temporal and constitutive inhibitions of CysLT. In conclusion, CysLT1R enhances MC activation by accelerating IgE-induced signal transduction, which enables the co-regulation of rapid degranulation and delayed synthesis of inflammatory mediators in MCs.

Anti-Allergic Drug Suppressed Pancreatic Carcinogenesis via Down-Regulation of Cellular Proliferation.

Pancreatic cancer is a fatal disease, and thus its chemoprevention is an important issue. Based on the recent report that patients with allergic diseases have a low risk for pancreatic cancer, we examined the potential chemopreventive effect of anti-allergic agents using a hamster pancreatic carcinogenesis model. Among the three anti-allergic drugs administered, montelukast showed a tendency to suppress the incidence of pancreatic cancer. Further animal study revealed a significantly decreased incidence of pancreatic cancer in the high-dose montelukast group compared with controls. The development of the pancreatic intraepithelial neoplasia lesions was also significantly suppressed. The Ki-67 labeling index was significantly lower in pancreatic carcinomas in the high-dose montelukast group than in controls. In vitro experiments revealed that montelukast suppressed proliferation of pancreatic cancer cells in a dose-dependent manner with decreased expression of phospho-ERK1/2. Montelukast induced G1 phase arrest. Conversely, leukotriene D4 (LTD4), an agonist of CYSLTR1, increased cellular proliferation of pancreatic cancer cells with an accumulation of phospho-ERK1/2. In our cohort, pancreatic ductal adenocarcinoma patients with high CYSLTR1 expression showed a significantly unfavorable clinical outcome compared with those with low expression. Our results indicate that montelukast exerts a chemopreventive effect on pancreatic cancer via the LTD4-CYSLTR1 axis and has potential for treatment of pancreatic carcinogenesis.

Leukotriene D4 role in allergic asthma pathogenesis from cellular and therapeutic perspectives.

Asthma is a chronic inflammatory and allergic disease that is mainly characterized by reversible airway obstruction and bronchial hyperresponsiveness. The incidence of asthma is increasing with more than 350 million people worldwide are affected. Up to now, there is no therapeutic option for asthma and most of the prescribed drugs aim to ameliorate the symptoms of the disease especially during the acute exacerbations after trigger exposure. Asthma is a heterogonous disease that involves interactions between inflammatory mediators and cellular components within the disease microenvironment including inflammatory and structural cells. Cysteinyl leukotrienes (cys-LTs) are inflammatory lipid mediators that have potent roles in asthma pathogenesis. CysLTs consisting of LTC4, LTD4, and LTE4 are mainly secreted by leukocytes and act through three main G-protein coupled receptors (CysLT1R, CysLT2R, and CysLT3R). LTD4 is the most potent bronchoconstrictor which gives it the priority to be discussed in detail in this review. LTD4 binds with high affinity to CysLT1R and many studies showed that using CysLT1R antagonists such as montelukast has a beneficial effect for asthmatics especially in corticosteroid refractory cases. Since asthma is a heterogeneous inflammatory disease of many cell types involved in the disease pathogenies and LTD4 has a special role in inflammation and bronchoconstriction, this review highlights the role of LTD4 on each cellular component in asthma and the benefits of using CysLT1R antagonists in ameliorating LTD4-induced effects.

Identification of cysteinyl-leukotriene-receptor 1 antagonists as ligands for the bile acid receptor GPBAR1.

The cysteinyl leukotrienes (CysLTs), i.e. LTC4, LTD4 and LTE4, are a family of proinflammatory agents synthesized from the arachidonic acid. In target cells, these lipid mediators bind to the cysteinyl leukotriene receptors (CysLTR), a family of seven transmembrane G-protein coupled receptors. The CysLT1R is a validated target for treatment of pulmonary diseases and several selective antagonists for this receptor, including montelukast, zafirlukast and pranlukast, have shown effective in the management of asthma. Nevertheless, others CysLT1R antagonists, such as the alpha-pentyl-3-[2-quinolinylmethoxy] benzyl alcohol (REV5901), have been extensively characterized without reaching sufficient priority for clinical development. Since drug reposition is an efficient approach for maximizing investment in drug discovery, we have investigated whether CysLT1R antagonists might exert off-target effects. In the report we demonstrate that REV5901 interacts with GPBAR1, a well characterized cell membrane receptor for secondary bile acids. REV5901 transactivates GPBAR1 in GPBAR1-transfected cells with an EC50 of 2.5 µM and accommodates the GPBAR1 binding site as shown by in silico analysis. Exposure of macrophages to REV5901 abrogates the inflammatory response elicited by bacterial endotoxin in a GPBAR1-dependent manner. In vivo, in contrast to montelukast, REV5901 attenuates inflammation and immune dysfunction in rodent models of colitis. The beneficial effects exerted by REV5901 in these models were abrogated by GPBAR1 gene ablation, confirming that REV5901, a shelved CysLT1R antagonist, is a GPBAR1 ligand. These data ground the basis for the development of novel hybrid ligands designed for simultaneous modulation of CysTL1R and GPBAR1.

Structural basis of ligand selectivity and disease mutations in cysteinyl leukotriene receptors.

Cysteinyl leukotriene G protein-coupled receptors CysLT1 and CysLT2 regulate pro-inflammatory responses associated with allergic disorders. While selective inhibition of CysLT1R has been used for treating asthma and associated diseases for over two decades, CysLT2R has recently started to emerge as a potential drug target against atopic asthma, brain injury and central nervous system disorders, as well as several types of cancer. Here, we describe four crystal structures of CysLT2R in complex with three dual CysLT1R/CysLT2R antagonists. The reported structures together with the results of comprehensive mutagenesis and computer modeling studies shed light on molecular determinants of CysLTR ligand selectivity and specific effects of disease-related single nucleotide variants.

Structure-guided probing of the leukotriene C4 binding site in human multidrug resistance protein 1 (MRP1; ABCC1).

The human multidrug resistance protein 1 (hMRP1) transporter is implicated in cancer multidrug resistance as well as immune responses involving its physiologic substrate, glutathione (GSH)-conjugated leukotriene C4 (LTC4). LTC4 binds a bipartite site on hMRP1, which a recent cryoelectron microscopy structure of LTC4-bound bovine Mrp1 depicts as composed of a positively charged pocket and a hydrophobic (H) pocket that binds the GSH moiety and surrounds the fatty acid moiety, respectively, of LTC4. Here, we show that single Ala and Leu substitutions of H-pocket hMRP1-Met1093 have no effect on LTC4 binding or transport. Estrone 3-sulfate transport is also unaffected, but both hMRP1-Met1093 mutations eliminate estradiol glucuronide transport, demonstrating that these steroid conjugates have binding sites distinct from each other and from LTC4. To eliminate LTC4 transport by hMRP1, mutation of 3 H-pocket residues was required (W553/M1093/W1246A), indicating that H-pocket amino acids are key to the vastly different affinities of hMRP1 for LTC4vs. GSH alone. Unlike organic anion transport, hMRP1-mediated drug resistance was more diminished by Ala than Leu substitution of Met1093. Although our findings generally support a structure in which H-pocket residues bind the lipid tail of LTC4, their critical and differential role in the transport of conjugated estrogens and anticancer drugs remains unexplained.-Conseil, G., Arama-Chayoth, M., Tsfadia, Y., Cole, S. P. C. Structure-guided probing of the leukotriene C4 binding site in human multidrug resistance protein 1 (MRP1; ABCC1).

Dysregulated fatty acid metabolism in nasal polyp-derived eosinophils from patients with chronic rhinosinusitis.

BACKGROUND: Eosinophils are multifunctional granulocytes capable of releasing various cytokines, chemokines, and lipid mediators. We previously reported dysregulated fatty acid metabolism in peripheral blood-derived eosinophils from patients with severe asthma. However, functional characteristics of eosinophils present in allergic inflammatory tissues remain largely uncharacterized. METHODS: We established a method for isolating CD69hi CCR3low CXCR4- siglec-8int eosinophils from nasal polyps of patients with eosinophilic rhinosinusitis (NP-EOS). Multi-omics analysis including lipidomics, proteomics, and transcriptomics was performed to analyze NP-EOS as compared to peripheral blood-derived eosinophils from healthy subjects (PB-EOS). RESULTS: Lipidomic analysis revealed impaired synthesis of prostaglandins and 15-lipoxygenase (15-LOX)-derived mediators, and selective upregulation of leukotriene D4 production. Furthermore, proteomics and transcriptomics revealed changes in the expression of specific enzymes (GGT5, DPEP2, and 15-LOX) responsible for dysregulated lipid metabolism. Ingenuity pathway analysis indicated the importance of type 2 cytokines and pattern recognition receptor pathways. Stimulation of PB-EOS with eosinophil activators IL-5, GM-CSF, and agonists of TLR2 and NOD2 mimicked the observed changes in lipid metabolism. CONCLUSION: Inflammatory tissue-derived eosinophils possess a specific phenotype with dysregulated fatty acid metabolism that may be targeted therapeutically to control eosinophilic inflammatory diseases.

Exosomes and cells from lung cancer pleural exudates transform LTC4 to LTD4, promoting cell migration and survival via CysLT1.

Tumor-derived exosomes can modulate the cancer microenvironment and induce metastatic spread. Exosomes may carry enzymes for leukotriene (LT) biosynthesis, but the role of exosomal LTs has not been studied in cancer. We isolated exosomes and malignant cells from pleura exudates from 14 patients with non-small cell lung cancer. Lipidomic profiles, migration and apoptosis were determined. Both exosomes and primary cancer cells contained γ-glutamyl transpeptidase 1 (GGT-1) and avidly transformed exogenous LTC4 to pro-tumorigenic LTD4, for the cells to levels 100-fold above their endogenous CysLT production. This suggests that cancer cells promote their own survival via LTD4 if supplied with LTC4, which in the exudates was produced by monocytic cells. Furthermore, exosomes promoted migration of cancer cells, which was counteracted by the CysLT1 antagonist montelukast. Montelukast also induced apoptosis of cancer cells, and this was partially inhibited by exosomes. Our results demonstrate how cancer cells and exosomes, together with monocytic cells in lung cancer tissue, can produce high amounts of LTD4, to stimulate cancer cell migration and survival. This suggests that part of the pro-metastatic effect of exosomes is mediated by the leukotriene machinery, further supporting the use of CysLT1 antagonists for lung cancer therapy.

Enhanced lipoxygenase/LTD4 signaling accounts for the exaggerated hypertensive and nephrotoxic effects of cyclosporine plus indomethacin in rats.

The combined use of cyclosporine (CSA) and nonsteroidal antiinflammatory drugs (NSAIDs) causes exaggerated rises in systolic blood pressure (SBP) and nephrotoxicity. We examined whether these influences relate to the arachidonate/5-lipoxygenase (LOX) pathway. Rats were treated with CSA (20 mg kg-1 day-1), indomethacin (5 mg kg-1 day-1), or their combination for 10 days. Changes in SBP and renal biochemical/histopathological characteristics along with leukotriene levels were determined in rats treated with or without LT receptor antagonists. CSA or indomethacin caused: (i) renal tubular atrophy and interstitial fibrosis, (ii) increases in serum creatinine, blood urea nitrogen (BUN), and renal LTD4, LTB4, TNF-α, TGF-ß1, and caspase-3, and (iii) decreases in renal PGE2 and total antioxidant capacity (TAC). SBP measured by tail-cuff plethysmography was increased by CSA but not indomethacin. These effects were mostly intensified in rats treated with CSA plus indomethacin. The co-treatment with montelukast (cysteinyl LT receptor blocker), but not ONO-4057 (LTB4 receptor blocker), ameliorated CSA/indomethacin-evoked hypertension, renal structural/biochemical deterioration, and LTD4 levels. Moreover, montelukast exhibited a greater capacity in reversing inflammatory, oxidative, apoptotic, and fibrotic abnormalities induced by CSA/indomethacin. Overall, lipoxygenase/LTD4 upregulation contributes to the exaggerated hypertension and nephrotoxicity caused by CSA/indomethacin. The therapeutic potential of cysteinyl LT receptor antagonism in rectifying CSA/NSAIDs-evoked anomalies is warranted.

Leukotriene D4 induces cellular senescence in osteoblasts.

Aging is associated with the development of osteoporosis, in which cellular senescence in osteoblasts plays a key role. Leukotriene D4 (LTD4), an important cysteinyl leukotriene (cysLT), is a powerful pro-inflammatory mediator formed from arachidonic acid. However, little information regarding the effects of LTD4 on the pathogenesis of osteoporosis has been reported before. In the present study, we defined the physiological roles of LTD4 in cellular senescence in osteoblasts. Our results indicate that LTD4 treatment decreased the expression of SIRT1 in a dose-dependent manner in MC3T3-E1 osteoblastic cells. Additionally, LTD4 significantly increased the expression of p53, p21 and plasminogen activator inhibitor-1 (PAI-1). LTD4 was also found to elevate the activity of ß-galactosidase (SA-ß-Gal) but to prevent BrdU incorporation. Our results indicate that cysteinyl leukotriene receptor 1 (cysLT1R) could be detected in MC3T3-E1 osteoblastic cells at both the mRNA and protein levels. However, cysLT2R was not expressed in these cells. Interestingly, we found that knockdown of cysLT1R or use of the selective cysLT1R antagonist montelukast abolished the LTD4-induced reduction in SIRT1 and increase in p53, p21, and PAI-1. Notably, knockdown of cysLT1R by transfection with cysLT1R siRNA or treatment with montelukast attenuated the LTD4-induced increase in SA-ß-Gal activity. Our study shows for the first time that LTD4 has a significant impact on cellular senescence in osteoblasts.

Zileuton alleviates acute cisplatin nephrotoxicity: Inhibition of lipoxygenase pathway favorably modulates the renal oxidative/inflammatory/caspase-3 axis.

OBJECTIVE: The current study investigated for the first time the possible beneficial effect of zileuton, a selective 5-lipoxygenase inhibitor (5-LOX), against cisplatin-induced acute renal failure. METHODOLOGY: Male Sprague-Dawley rats (180-200 g) were administered cisplatin once (5 mg/kg, i.p.) alone or combined with oral zileuton (10 mg/kg, given twice; 1 h before and 12 h after cisplatin). RESULTS: Compared with control rats, acute cisplatin administration caused significant increases of BUN (33.76 ±â€¯7.74 vs 61.88 ±â€¯11.35 mg/dl), serum creatinine (0.61 ±â€¯0.21 vs 1.56 ±â€¯0.28 mg/dl), renal levels of MDA (6.40 ±â€¯1.04 vs 20.52 ±â€¯2.18 nmol/g tissue), NOx (3.45 ±â€¯1.20 vs 17.70 ±â€¯2.27 nmol/g tissue), TNF-α (6.71 ±â€¯0.66 vs 23.71 ±â€¯3.41 pg/g tissue), MPO (0.87 ±â€¯0.09 vs 3.12 ±â€¯0.41 U/mg tissue protein) and renal caspase-3 activity (2.81 ±â€¯0.37 vs 12.70 ±â€¯2.94 U/mg tissue protein). Whereas, total SOD activity (1.99 ±â€¯0.53 vs 0.79 ±â€¯0.06 U/mg tissue protein) and IL-10 (110.98 ±â€¯19.70 vs 62.34 ±â€¯14.42 pg/g tissue) were significantly decreased. Cisplatin-induced nephrotoxicity was further confirmed histopathologically (tubular necrosis, cystic dilatation of renal tubules, vacuolar degeneration of renal tubular epithelium with perivascular oedema, and interstitial fibrosis). These changes were accompanied by alteration in 5-LOX pathway manifested as elevated renal levels of 5-LOX, LTD4 and LTB4. Simultaneous administration of zileuton to the cisplatin-treated rats reversed the deleterious renal insults and restored the measured parameters near to control values. CONCLUSIONS: These data establish the first experimental evidence that zileuton abrogates cisplatin nephrotoxicity in rats probably via the inhibition of detrimental actions of 5-LOX products, thus favorably affecting renal oxidative/inflammatory/caspase-3 axis. Based on current findings, the therapeutic prospect of zileuton for this purpose is recommended.

A new CysLT1 and CysLT2 receptors-mediated anaphylaxis guinea pig model.

Although the effectiveness of CysLT1 receptor antagonists on asthma has been clinically established, the effects of CysLT2 receptor antagonists are still unclear. The purpose of this study was to develop a new CysLT1 and CysLT2 receptors-mediated anaphylaxis guinea pig model using S-hexyl GSH, a γ-glutamyl transpeptidase (GTP) inhibitor, to suppress conversion of LTC4 to LTD4. Actively sensitized guinea pigs were challenged with OVA in the absence or presence of S-hexyl GSH, and survival rate following anaphylactic response was monitored. OVA-induced fatal anaphylaxis in the absence of S-hexyl GSH was almost completely inhibited by montelukast, a CysLT1 receptor antagonist, but not by the CysLT2 receptor antagonist BayCysLT2RA. However, under treatment with S-hexyl-GSH, the inhibitory effect of motelukast was dramatically diminished, whereas that of BayCysLT2RA was markedly increased. The dual CysLT1/2 receptor antagonist ONO-6950 effectively inhibited anaphylactic response in both S-hexyl GSH-treated and non-treated animals. LC/MS/MS analysis revealed that S-hexyl GSH treatment actually inhibited LTC4 metabolism in the blood and lung tissues. Using S-hexyl GSH, we developed a novel CysLT1 and CysLT2 receptors-mediated anaphylaxis guinea pig model that can be useful for not only screening both CysLT2 and CysLT1/2 receptors antagonists, but also for functional analysis of CysLT2 receptors.

M2-like macrophages induce colon cancer cell invasion via matrix metalloproteinases.

The inflammatory milieu plays an important role in colon cancer development and progression. Previously, we have shown that tumor-associated macrophages (TAMs), an important component of the tumor microenvironment, are enriched in tumors compared with normal tissue and confer a poorer prognosis. In the present study, we found that matrix metallopeptidase-9 (MMP-9), which degrades extracellular matrix proteins, was increased in biopsies from colon cancer patients and in mouse xenografts with SW480 cell-derived tumors. SW480 colon cancer cells exposed to M2-like macrophage-conditioned medium (M2-medium) exhibited increased MMP-9 mRNA, protein expression and gelatinase activity. A similar effect was obtained by the addition of tumor necrosis factor-α (TNFα) and leukotriene D4 (LTD4 ). MMP-9 expression and activity were reduced by a TNFα blocking antibody adalimumab and a cysteinyl leukotriene receptor 1 (CysLTR1, the receptor for LTD4 ) antagonist montelukast. M2-medium also induced changes in the epithelial-mesenchymal transition (EMT) markers E-cadherin, ß-catenin, vimentin, and snail in SW480 cells. We also found that both M2-medium and TNFα and LTD4 induced stabilization/nuclear translocation of ß-catenin. Furthermore, we also observed an elongated phenotype that may indicate increased invasiveness, as confirmed in a collagen I invasion assay. M2-medium increased the invasive ability, and a similar effect was also obtained by the addition of TNFα and LTD4 . The specific MMP inhibitor I or adalimumab and montelukast reduced the number of invasive cells. In conclusion, our findings show that M2-medium enriched in TNFα and LTD4 promote colon cancer cell invasion via MMP-9 expression and activation and the induction of EMT.

Dehydropeptidase 1 promotes metastasis through regulation of E-cadherin expression in colon cancer.

Dehydropeptidase 1 (DPEP1) is a zinc-dependent metalloproteinase that is expressed aberrantly in several cancers. The role of DPEP1 in cancer remain controversial. In this study, we demonstrate that DPEP1 functions as a positive regulator for colon cancer cell metastasis. The expression of DPEP1 mRNA and proteins were upregulated in colon cancer tissues compared to normal mucosa. Gain-of-function and loss-of-function approaches were used to examine the malignant phenotype of DPEP1-expressing or DPEP1-depleted cells. DPEP1 expression caused a significant increase in colon cancer cell adhesion and invasion in vitro, and metastasis in vivo. In contrast, DPEP1 depletion induced opposite effects. Furthermore, cilastatin, a DPEP1 inhibitor, suppressed the invasion and metastasis of DPEP1-expressing cells. DPEP1 inhibited the leukotriene D4 signaling pathway and increased the expression of E-cadherin. We also show that DPEP1 mediates TGF-ß-induced EMT. TGF-ß transcriptionally repressed DPEP1 expression. TGF-ß treatment decreased E-cadherin expression and promoted cell invasion in DPEP1-expressing colon cancer cell lines, whereas it did not affect these parameters in DPEP1-depleted cell lines. These results suggest that DPEP1 promotes cancer metastasis by regulating E-cadherin plasticity and that it might be a potential therapeutic target for preventing the progression of colon cancer.

[The relationship between the key nasal symptoms and the level of histamineand leukotriene D4 in serum and nasal secretions in allergic rhinitis].

Objective:To investigate the relationship between main nasal symptoms and the levels of histamine and leukotriene D4(LTD4) in serum and nasal secretions in allergic rhinitis(AR),and provide a preliminary guidance for individualized treatment in AR.Method:One hundred and eight cases of AR were divided into two groups,i.e.sneezing group and nasal congestion group,according to 2014 clinical guidelines for allergic rhinitis from January 2014 to June 2015.The levels of histamine and LTD4 in serum and nasal secretions were determined by enzyme-linked immunosorbent assay(ELISA) and the relationship was explored between the clinical main nasal symptoms score and the levels of histamine and LTD4.Result:The scores of sneezing(5.58±2.59)for AR were obviously related to the levels of histamine in serum(8.39±4.07)ng/ml and nasal secretion(5.06±2.47)ng/ml,(r=0.79,0.78,all P<0.05).The scores of nasal congestion(5.34±2.36) for AR were also related to the levels of LTD4 in serum(0.356±0.155 ng/ml) and nasal secretion(0.215±0.092)ng/ml,(r=0.74,0.72,all P<0.05).And the levels of histamine(8.39±4.07)ng/ml and LTD4(0.356±0.155) ng/ml in serum for AR patients were positively correlated with the levels in nasal secretions(r=0.99,P<0.01;r=0.98,P<0.01).Conclusion:In AR patients,the high levels of histamine and LTD4 in serum and nasal secretions are closely related to the sneezing symptoms and nasal obstruction symptoms,respectively.

Expression of CysLT2 receptors in asthma lung, and their possible role in bronchoconstriction.

BACKGROUND: The expression and functional role of CysLT2 receptors in asthma have not been clarified. In this study, we evaluated CysLT2 receptors expression, and effects of CysLT2-and CysLT1/2-receptor antagonists on antigen-induced bronchoconstriction using isolated lung tissues from both asthma and non-asthma subjects. METHODS: CysLT1 and CysLT2 receptors expression in asthma and non-asthma lung tissue preparations was examined in immunohistochemistry experiments, and their functional roles in antigen-induced bronchoconstriction were assessed using ONO-6950, a dual CysLT1/2-receptor antagonist, montelukast, a CysLT1 receptor antagonist, and BayCysLT2RA, a CysLT2 receptor-specific antagonist. RESULTS: CysLT1 receptors were expressed on the bronchial smooth muscle and epithelium, and on alveolar leukocytes in 5 in 5 non-asthma subjects and 2 in 2 asthma subjects. On the other hand, although degrees of CysLT2 receptors expression were variable among the 5 non-asthma subjects, the expression in the asthma lung was detected on bronchial smooth muscle, epithelium and alveolar leukocytes in 2 in 2 asthma subjects. In the non-asthma specimens, antagonism of CysLT2 receptors did not affect antigen-induced bronchial contractions, even after pretreatment with the CysLT1-receptor specific antagonist, montelukast. However, in the bronchus isolated from one of the 2 asthma subjects, antagonism of CysLT2 receptors suppressed contractions, and dual antagonism of CysLT1 and CysLT2 receptors resulted in additive inhibitory effect on anaphylactic contractions. CONCLUSIONS: CysLT2 receptors were expressed in lung specimens isolated from asthma subjects. Activation of CysLT2 receptors may contribute to antigen-induced bronchoconstriction in certain asthma population.

Intravenous anesthetic propofol suppresses prostaglandin E2 and cysteinyl leukotriene production and reduces edema formation in arachidonic acid-induced ear inflammation.

Propofol is an intravenous drug widely used for anesthesia and sedation. Previously, propofol was shown to inhibit cyclo-oxygenase (COX) and 5-lipoxygenase (5-LOX) activities. Because these enzyme-inhibiting effects have only been demonstrated in vitro, this study sought to ascertain whether similar effects might also be observed in vivo. In the current studies, effects of propofol were tested in a murine model of arachidonic acid-induced ear inflammation. Specifically, propofol - as a pre-treatment -- was intraperitoneally and then topical application of arachidonic acid was performed. After 1 h, tissue biopsies were collected and tested for the presence of edema and for levels of inflammatory mediators. The results indicated that the administration of propofol significantly suppressed ear edema formation, tissue myeloperoxidase activity, and tissue production of both prostaglandin E2 and cysteinyl leukotrienes. From the data, it can be concluded that propofol could exert anti-COX and anti-5-LOX activities in an in vivo model and that these activities in turn could have, at least in part, suppressed arachidonic acid-induced edema formation in the ear.

Activation of calcium- and voltage-gated potassium channels of large conductance by leukotriene B4.

Calcium/voltage-gated, large conductance potassium (BK) channels control numerous physiological processes, including myogenic tone. BK channel regulation by direct interaction between lipid and channel protein sites has received increasing attention. Leukotrienes (LTA4, LTB4, LTC4, LTD4, and LTE4) are inflammatory lipid mediators. We performed patch clamp studies in Xenopus oocytes that co-expressed BK channel-forming (cbv1) and accessory ß1 subunits cloned from rat cerebral artery myocytes. Leukotrienes were applied at 0.1 nm-10 µm to either leaflet of cell-free membranes at a wide range of [Ca(2+)]i and voltages. Only LTB4 reversibly increased BK steady-state activity (EC50 = 1 nm; Emax reached at 10 nm), with physiological [Ca(2+)]i and voltages favoring this activation. Homomeric cbv1 or cbv1-ß2 channels were LTB4-resistant. Computational modeling predicted that LTB4 docked onto the cholane steroid-sensing site in the BK ß1 transmembrane domain 2 (TM2). Co-application of LTB4 and cholane steroid did not further increase LTB4-induced activation. LTB4 failed to activate ß1 subunit-containing channels when ß1 carried T169A, A176S, or K179I within the docking site. Co-application of LTB4 with LTA4, LTC4, LTD4, or LTE4 suppressed LTB4-induced activation. Inactive leukotrienes docked onto a portion of the site, probably preventing tight docking of LTB4. In summary, we document the ability of two endogenous lipids from different chemical families to share their site of action on a channel accessory subunit. Thus, cross-talk between leukotrienes and cholane steroids might converge on regulation of smooth muscle contractility via BK ß1. Moreover, the identification of LTB4 as a highly potent ligand for BK channels is critical for the future development of ß1-specific BK channel activators.

Vitamin E, γ-tocopherol, diminishes ex vivo basophil response to dust mite allergen.

Epidemiologic studies suggest that dietary vitamin E is a candidate intervention for atopic disease. We used in vitro and ex vivo exposures to test the hypothesis that the most common dietary isoform of vitamin E, γ-tocopherol (γT), could suppress FcεRI-mediated basophil activation. Rat basophilic leukemia (RBL)-SX38 cells that express human FcεRI were treated with or without γT, followed by stimulation with α-IgE. In the ex vivo study, 20 Der f 1-allergic volunteers consumed a γT-enriched supplement for 7 days. Their basophils were challenged ex vivo with α-IgE and graded doses of Der f 1 before and after the supplementation period. γt treatment of RBL-SX38 cells significantly reduced basophil degranulation and de novo TH2 cytokine production. Daily consumption of a γT-rich supplement by dust mite-allergic volunteers reduced basophil activation after ex vivo dust mite challenge. Vitamin E supplements rich in γT may be useful adjuncts in decreasing atopic disease.

Bisphenol A at concentrations relevant to human exposure enhances histamine and cysteinyl leukotriene release from bone marrow-derived mast cells.

Bisphenol A (BPA), a monomer of polycarbonate plastics and epoxide resin, acts as an endocrine-active compound and has been shown to enhance the inflammatory response to allergen challenge. Previous reports in rodents have demonstrated that perinatal BPA exposure alters airway inflammation following sensitization and challenge to ovalbumin in juvenile and adult offspring. Additionally, a high concentration of BPA has been shown to enhance mediator release in mast cell lines. This study aimed to determine if short-term BPA exposure, at levels relevant to human exposure, enhances mast cell release of histamine and cysteinyl leukotrienes (CysLTs). Primary murine bone marrow-derived mast cells (BMMC) produced from the femurs of female C57BL/6 mice were stimulated with BPA or estradiol (E2) in vitro. It was observed that both BPA and E2 increased BMMC histamine release over a range of nanomolar concentrations (1-1000 nM). The estrogen receptor (ER) antagonist ICI 182,780 partially blocked the ability of E2, but not BPA, to elevate histamine release. BPA also increased CysLT release, which was not abrogated by ER inhibition. It was also observed that the ability of BPA to enhance histamine and CysLT release was inhibited by blocking the extracellular signal-regulated kinase (ERK) pathway with U0126 or by chelating extracellular calcium (Ca(2+)) using EGTA. In summary, these experiments are the first to demonstrate that acute BPA exposure enhances mast cell histamine and CysLT release in vitro--an effect that is not dependent on an ER-mediated mechanism. Instead, BPA-induced mast cell histamine and CysLT release may be mediated, in part, by the ERK pathway and extracellular Ca(2+) concentrations. These data suggest that exposure to BPA at levels relevant to human exposure may provoke an acute inflammatory response in atopic individuals via enhanced mast cell activation.