The role of human 5-Lipoxygenase (5-LO) in carcinogenesis - a question of canonical and non-canonical functions.

5-Lipoxygenase (5-LO), a fatty acid oxygenase, is the central enzyme in leukotriene (LT) biosynthesis, potent arachidonic acid-derived lipid mediators released by innate immune cells, that control inflammatory and allergic responses. In addition, through interaction with 12- and 15-lipoxgenases, the enzyme is involved in the formation of omega-3 fatty acid-based oxylipins, which are thought to be involved in the resolution of inflammation. The expression of 5-LO is frequently deregulated in solid and liquid tumors, and there is strong evidence that the enzyme plays an important role in carcinogenesis. However, global inhibition of LT formation and signaling has not yet shown the desired success in clinical trials. Curiously, the release of 5-LO-derived lipid mediators from tumor cells is often low, and the exact mechanism by which 5-LO influences tumor cell function is poorly understood. Recent data now show that in addition to releasing oxylipins, 5-LO can also influence gene expression in a lipid mediator-independent manner. These non-canonical functions, including modulation of miRNA processing and transcription factor shuttling, most likely influence cancer cell function and the tumor microenvironment and might explain the low clinical efficacy of pharmacological strategies that previously only targeted oxylipin formation and signaling by 5-LO. This review summarizes the canonical and non-canonical functions of 5-LO with a particular focus on tumorigenesis, highlights unresolved issues, and suggests future research directions.

Qijiao Shengbai Capsule alleviated leukopenia by interfering leukotriene pathway: Integrated network study of multi-omics.

BACKGROUND: Leukopenia could be induced by chemotherapy, which leads to bone marrow suppression and even affects the therapeutic progression of cancer. Qijiao Shengbai Capsule (QSC) has been used for the treatment of leukopenia in clinic, but its bioactive components and mechanisms have not yet been elucidated clearly. PURPOSE: This study aimed to elucidate the molecular mechanisms of QSC in treating leukopenia. STUDY DESIGN: Serum pharmacochemistry, multi-omics, network pharmacology, and validation experiment were combined to study the effect of QSC in murine leukopenia model. METHODS: First, UPLC-QTOF-MS was used to clarify the absorbed components of QSC. Then, cyclophosphamide (CTX) was used to induce mice model with leukopenia, and the therapeutic efficacy of QSC was assessed by an integrative approach of multi-omics and network pharmacology strategy. Finally, molecular mechanisms and potential therapeutic targets were identified by validated experiments. RESULTS: 121 compounds absorbed in vivo were identified. QSC significantly increase the count of white blood cells (WBCs) in peripheral blood of leukopenia mice with 15 days treatment. Multi-omics and network pharmacology revealed that leukotriene pathway and MAPK signaling pathway played crucial roles during the treatment of leukopenia with QSC. Six targets (ALOX5, LTB4R, CYSLTR1, FOS, JUN, IL-1ß) and 13 prototype compounds were supposed to be the key targets and potential active components, respectively. The validation experiment further confirmed that QSC could effectively inhibit the inflammatory response induced by leukopenia. The inhibitors of ALOX5 activity can significantly increase the number of WBCs in leukopenia mice. Molecular docking of ALOX5 suggested that calycosin, daidzein, and medicarpin were the potentially active compounds of QSC. CONCLUSION: Leukotriene pathway was found for the first time to be a key role in the development of leukopenia, and ALOX5 was conformed as the potential target. QSC may inhibit the inflammatory response and interfere the leukotriene pathway, it is able to improve hematopoiesis and achieve therapeutic effects in the mice with leukopenia.

A critical role for Macrophage-derived Cysteinyl-Leukotrienes in HIV-1 induced neuronal injury.

Macrophages (MΦ) infected with human immunodeficiency virus (HIV)-1 or activated by its envelope protein gp120 exert neurotoxicity. We found previously that signaling via p38 mitogen-activated protein kinase (p38 MAPK) is essential to the neurotoxicity of HIVgp120-stimulated MΦ. However, the associated downstream pathways remained elusive. Here we show that cysteinyl-leukotrienes (CysLT) released by HIV-infected or HIVgp120 stimulated MΦ downstream of p38 MAPK critically contribute to neurotoxicity. SiRNA-mediated or pharmacological inhibition of p38 MAPK deprives MΦ of CysLT synthase (LTC4S) and, pharmacological inhibition of the cysteinyl-leukotriene receptor 1 (CYSLTR1) protects cerebrocortical neurons against toxicity of both gp120-stimulated and HIV-infected MΦ. Components of the CysLT pathway are differentially regulated in brains of HIV-infected individuals and a transgenic mouse model of NeuroHIV (HIVgp120tg). Moreover, genetic ablation of LTC4S or CysLTR1 prevents neuronal damage and impairment of spatial memory in HIVgp120tg mice. Altogether, our findings suggest a novel critical role for cysteinyl-leukotrienes in HIV-associated brain injury.

Hyphal-associated protein expression is crucial for Candida albicans-induced eicosanoid biosynthesis in immune cells.

Candida albicans causes opportunistic infections ranging from mucosal mycoses to life-threatening systemic infections in immunocompromised patients. During C. albicans infection, leukotrienes and prostaglandins are formed from arachidonic acid by 5-lipoxygenase (5-LOX) and cyclooxygenases, respectively to amplify inflammatory conditions, but also to initiate macrophage infiltration to achieve tissue homeostasis. Since less is known about the cellular mechanisms triggering such lipid mediator biosynthesis, we investigated the eicosanoid formation in monocyte-derived M1 and M2 macrophages, neutrophils and HEK293 cells transfected with 5-LOX and 5-LOX-activating protein (FLAP) in response to C. albicans yeast or hyphae. Leukotriene biosynthesis was exclusively induced by hyphae in neutrophils and macrophages, whereas prostaglandin E2 was also formed in response to yeast cells by M1 macrophages. Eicosanoid biosynthesis was significantly higher in M1 compared to M2 macrophages. In HEK_5-LOX/FLAP cells only hyphae activated the essential 5-LOX translocation to the nuclear membrane. Using yeast-locked C. albicans mutants, we demonstrated that hyphal-associated protein expression is critical in eicosanoid formation. For neutrophils and HEK_5-LOX/FLAP cells, hyphal wall protein 1 was identified as the essential surface protein that stimulates leukotriene biosynthesis. In summary, our data suggest that hyphal-associated proteins of C. albicans are central triggers of eicosanoid biosynthesis in human phagocytes.

The role of captopril in leukotriene deficient type 1 diabetic mice.

T1D can be associated with metabolic disorders and several impaired pathways, including insulin signaling, and development of insulin resistance through the renin-angiotensin system (RAS). The main precursor of RAS is angiotensinogen (Agt) and this system is often linked to autophagy dysregulation. Dysregulated autophagy has been described in T1D and linked to impairments in both glucose metabolism, and leukotrienes (LTs) production. Here, we have investigated the role of RAS and LTs in both muscle and liver from T1D mice, and its effects on insulin and autophagy pathways. We have chemically induced T1D in 129sve and 129sve 5LO-/- mice (lacking LTs) with streptozotocin (STZ). To further inhibit ACE activity, mice were treated with captopril (Cap). In muscle of T1D mice, treatment with Cap increased the expression of RAS (angiotensinogen and angiotensin II receptor), insulin signaling, and autophagy markers, regardless of the genotype. In the liver of T1D mice, the treatment with Cap increased the expression of RAS and insulin signaling markers, mostly when LTs were absent. 5LO-/- T1D mice showed increased insulin sensitivity, and decreased NEFA, after the Cap treatment. Cap treatment impacted both insulin signaling and autophagy pathways at the mRNA levels in muscle and liver, indicating the potential role of ACE inhibition on insulin sensitivity and autophagy in T1D.

Eosinophils induces glioblastoma cell suppression and apoptosis - Roles of GM-CSF and cysteinyl-leukotrienes.

BACKGROUND: Glioblastoma is the most common and lethal primary brain tumor in adults. Despite the available cancer treatments, the recurrence of the tumor is high, and the survival rate is low. New approaches to antitumor therapies are needed. Eosinophils are prominent in allergic diseases and accumulate in several human brain tumors. Recently, the antitumor role of eosinophils has been targeted as eosinophils release several cytotoxic factors that induce cell impairment and death. OBJECTIVE: Here we aim to evaluate the interaction of the eosinophil and glioblastoma cells, the mechanism involved in the potential killing of the glioblastoma cells by the eosinophils, and how allergy/asthma could confer a better glioblastoma prognosis. METHODS: Eosinophils and serum from asthmatic and non-asthmatic donors were cultivated with different glioblastoma cell lines. RESULTS: Glioblastoma cells recruit eosinophils via GM-CSF signaling, activating and increasing eosinophil survivability and function on a GM-CSF-dependent manner. Eosinophils reduce glioblastoma cells metabolism, proliferation, and migration, via Fas/FasL. Cysteinyl-leukotrienes are accounted for the asthmatic serum enhancement of the glioblastoma cell migration and proliferation. Cysteinyl-leukotrienes enhance glioblastoma cell proliferation and migration, albeit activate eosinophils that suppress glioblastoma cells. CONCLUSION: Eosinophils have the potential to be key cells on glioblastoma therapeutics, as allergy and eosinophilia are correlated with a better glioblastoma prognosis. Eosinophils are elicited and attach to glioblastoma cells, where, by its cytotoxic function, via Fas/FasL, hind glioblastoma cell metabolism, proliferation, migration, and induce cell death.

The Multifarious Functions of Leukotrienes in Bone Metabolism.

Leukotrienes (LTs) are derived from arachidonic acid metabolism by the 5-lipoxygenase (5-LO) enzyme. The production of LTs is stimulated in the pathogenesis of rheumatoid arthritis (RA), osteoarthritis, and periodontitis, with a relevant contribution to bone resorption. However, its role in bone turnover, particularly the suppression of bone formation by modulating the function of osteoclasts and osteoblasts, remains unclear. We investigated the effects of LTs on bone metabolism and their impact on osteogenic differentiation and osteoclastogenesis using a 5-LO knockout (KO) mouse model. Results from micro-computed tomography (µCT) analysis of femur from 8-week-old 5-LO-deficient mice showed increased cortical bone and medullary region in females and males and decreased trabecular bone in females. In the vertebra, we observed increased marrow area in both females and males 5-LO KO and decreased trabecular bone only in females 5-LO KO. Immunohistochemistry (IHC) analysis showed higher levels of osteogenic markers tissue-nonspecific alkaline phosphatase (TNAP) and osteopontin (OPN) and lower expression of osteoclastogenic marker tartrate-resistant acid phosphatase (TRAP) in the femurs of 5-LO KO mice versus wild-type (WT). Alkaline phosphatase activity and mineralization assay results showed that the 5-LO absence enhances osteoblasts differentiation and mineralization but decreases the proliferation. Alkaline phosphatase (ALP), Bglap, and Sp7 gene expression were higher in 5-LO KO osteoblasts compared to WT cells. Eicosanoids production was higher in 5-LO KO osteoblasts except for thromboxane 2, which was lower in 5-LO-deficient mice. Proteomic analysis identified the downregulation of proteins related to adenosine triphosphate (ATP) metabolism in 5-LO KO osteoblasts, and the upregulation of transcription factors such as the adaptor-related protein complex 1 (AP-1 complex) in long bones from 5-LO KO mice leading to an increased bone formation pattern in 5-LO-deficient mice. We observed enormous differences in the morphology and function of osteoclasts with reduced bone resorption markers and impaired osteoclasts in 5-LO KO compared to WT osteoclasts. Altogether, these results demonstrate that the absence of 5-LO is related to the greater osteogenic profile. © 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

The role of oxylipins in NSAID-exacerbated respiratory disease (N-ERD).

Nonsteroidal anti-inflammatory drug (NSAID)-exacerbated respiratory disease (N-ERD) is characterized by nasal polyp formation, adult-onset asthma, and hypersensitivity to all cyclooxygenase-1 (COX-1) inhibitors. Oxygenated lipids are collectively known as oxylipins and are polyunsaturated fatty acids (PUFA) oxidation products. The most extensively researched oxylipins being the eicosanoids formed from arachidonic acid (AA). There are four major classes of eicosanoids including leukotrienes, prostaglandins, thromboxanes, and lipoxins. In N-ERD, the underlying inflammatory process of the upper and lower respiratory systems begins and occurs independently of NSAID consumption and is due to the overproduction of cysteinyl leukotrienes. Leukotriene mediators all induce edema, bronchoconstriction, and airway mucous secretion. Thromboxane A2 is a potent bronchoconstrictor and induces endothelial adhesion molecule expression. Elevated Prostaglandin D2 metabolites lead to vasoconstriction, additionally impaired up-regulation of prostaglandin E2 leads to symptoms seen in N-ERD as it is essential for maintaining homeostasis of inflammatory responses in the airway and has bronchoprotective and anti-inflammatory effects. A characteristic feature of N-ERD is diminished lipoxin levels, this decreased capacity to form endogenous mediators with anti-inflammatory properties could facilitate local inflammatory response and expose bronchial smooth muscle to relatively unopposed actions of broncho-constricting substances. Treatment options, such as leukotriene modifying agents, aspirin desensitization, biologic agents and ESS, appear to influence eicosanoid pathways, however more studies need to be done to further understand the role of oxylipins. Besides AA-derived eicosanoids, other oxylipins may also pay a role but have not been sufficiently studied. Identifying pathogenic N-ERD mechanism is likely to define more effective treatment targets.

Combinatorial targeting of G-protein-coupled bile acid receptor 1 and cysteinyl leukotriene receptor 1 reveals a mechanistic role for bile acids and leukotrienes in drug-induced liver injury.

BACKGROUND AND AIM: Drug-induced liver injury (DILI) is a common disorder that involves both direct liver cell toxicity and immune activation. The bile acid receptor, G-protein-coupled bile acid receptor 1 (GPBAR1; Takeda G-protein-coupled receptor 5 [TGR5]), and cysteinyl leukotriene receptor (CYSLTR) 1 are G-protein-coupled receptors activated by bile acids and leukotrienes, exerting opposite effects on cell-to-cell adhesion, inflammation, and immune cell activation. To investigate whether GPBAR1 and CYSLTR1 mutually interact in the development of DILI, we developed an orally active small molecule, CHIN117, that functions as a GPBAR1 agonist and CYSLTR1 antagonist. APPROACH AND RESULTS: RNA-sequencing analysis of liver explants showed that acetaminophen (APAP) intoxication positively modulates the leukotriene pathway, CYSLTR1, 5-lipoxygenase, and 5-lipoxygenase activating protein, whereas GPBAR1 gene expression was unchanged. In mice, acute liver injury induced by orally dosing APAP (500 mg/kg) was severely exacerbated by Gpbar1 gene ablation and attenuated by anti-Cysltr1 small interfering RNA pretreatment. Therapeutic dosing of wild-type mice with CHIN117 reversed the liver damage caused by APAP and modulated up to 1300 genes, including 38 chemokines and receptors, that were not shared by dosing mice with a selective GPBAR1 agonist or CYSLTR1 antagonist. Coexpression of the two receptors was detected in liver sinusoidal endothelial cells (LSECs), monocytes, and Kupffer cells, whereas combinatorial modulation of CYSLTR1 and GPBAR1 potently reversed LSEC/monocyte interactions. CHIN117 reversed liver damage and liver fibrosis in mice administered CCl 4 . CONCLUSIONS: By genetic and pharmacological approaches, we demonstrated that GPBAR1 and CYSLTR1 mutually interact in the development of DILI. A combinatorial approach designed to activate GPBAR1 while inhibiting CYSLTR1 reverses liver injury in models of DILI.

Structures of Leukotriene Biosynthetic Enzymes and Development of New Therapeutics.

Leukotrienes are potent immune-regulating lipid mediators with patho-genic roles in inflammatory and allergic diseases, particularly asthma. These autacoids also contribute to low-grade inflammation, a hallmark of cardiovascular, neurodegenerative, metabolic, and tumor diseases. Biosynthesis of leukotrienes involves release and oxidative metabolism of arachidonic acid and proceeds via a set of cytosolic and integral membrane enzymes that are typically expressed by cells of the innate immune system. In activated cells, these enzymes traffic and assemble at the endoplasmic and perinuclear membrane, together comprising a biosynthetic complex. Here we describe recent advances in our molecular understanding of the protein components of the leukotriene-synthesizing enzyme machinery and also briefly touch upon the leukotriene receptors. Moreover, we discuss emerging opportunities for pharmacological intervention and development of new therapeutics.

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.

Integrated genomic, transcriptomic, and epigenetic analyses identify a leukotriene synthesis-related M2 macrophage gene signature that predicts prognosis and treatment vulnerability in gliomas.

The pathological implications of tumor-associated macrophages in the glioma microenvironment have been highlighted, while there lacks a gene signature to characterize the functional status and clinical implications of these cells. Comprehensive bioinformatics approaches were employed to develop an M2 macrophage-associated gene signature at bulk-tumor and single-cell levels and explore immunological and metabolic features. Consequently, the PI3K pathway and fatty acid metabolism were correlated with the M2 fraction. Further distilling the pathway members resulted in a leukotriene synthesis-related gene signature (Macro index), including PIK3R5, PIK3R6, ALOX5, ALOX5AP, and ALOX15B, that was primarily expressed by monocytes/macrophages. Increased Macro index predicted IL13-induced macrophages, and was associated with T-cell dysfunction at both transcriptional and epigenetic levels and predicted an unfavorable outcome. Besides, the Macro index was proportional with PAI1 at the protein level, with high levels of the latter suggesting a decreased progression-free interval of glioblastoma. Notably, the monocytes/macrophages in the glioma environment contribute to the expression of immune checkpoints and the Macro index predicts glioma responsiveness to anti-PD1 treatment. Together, our study proposed a leukotriene synthesis-related M2 macrophage gene signature, which may provide insights into the role of these cells in the glioma microenvironment and facilitate individually tailored therapeutic strategies for the disease.

Loss of full-length dystrophin expression results in major cell-autonomous abnormalities in proliferating myoblasts.

Duchenne muscular dystrophy (DMD) affects myofibers and muscle stem cells, causing progressive muscle degeneration and repair defects. It was unknown whether dystrophic myoblasts-the effector cells of muscle growth and regeneration-are affected. Using transcriptomic, genome-scale metabolic modelling and functional analyses, we demonstrate, for the first time, convergent abnormalities in primary mouse and human dystrophic myoblasts. In Dmdmdx myoblasts lacking full-length dystrophin, the expression of 170 genes was significantly altered. Myod1 and key genes controlled by MyoD (Myog, Mymk, Mymx, epigenetic regulators, ECM interactors, calcium signalling and fibrosis genes) were significantly downregulated. Gene ontology analysis indicated enrichment in genes involved in muscle development and function. Functionally, we found increased myoblast proliferation, reduced chemotaxis and accelerated differentiation, which are all essential for myoregeneration. The defects were caused by the loss of expression of full-length dystrophin, as similar and not exacerbated alterations were observed in dystrophin-null Dmdmdx-ßgeo myoblasts. Corresponding abnormalities were identified in human DMD primary myoblasts and a dystrophic mouse muscle cell line, confirming the cross-species and cell-autonomous nature of these defects. The genome-scale metabolic analysis in human DMD myoblasts showed alterations in the rate of glycolysis/gluconeogenesis, leukotriene metabolism, and mitochondrial beta-oxidation of various fatty acids. These results reveal the disease continuum: DMD defects in satellite cells, the myoblast dysfunction affecting muscle regeneration, which is insufficient to counteract muscle loss due to myofiber instability. Contrary to the established belief, our data demonstrate that DMD abnormalities occur in myoblasts, making these cells a novel therapeutic target for the treatment of this lethal disease.

Brain-Specific Increase in Leukotriene Signaling Accompanies Chronic Neuroinflammation and Cognitive Impairment in a Model of Gulf War Illness.

Persistent cognitive impairment is a primary central nervous system-related symptom in veterans afflicted with chronic Gulf War Illness (GWI). Previous studies in a rat model have revealed that cognitive dysfunction in chronic GWI is associated with neuroinflammation, typified by astrocyte hypertrophy, activated microglia, and enhanced proinflammatory cytokine levels. Studies in a mouse model of GWI have also shown upregulation of several phospholipids that serve as reservoirs of arachidonic acid, a precursor of leukotrienes (LTs). However, it is unknown whether altered LT signaling is a component of chronic neuroinflammatory conditions in GWI. Therefore, this study investigated changes in LT signaling in the brain of rats displaying significant cognitive impairments six months after exposure to GWI-related chemicals and moderate stress. The concentration of cysteinyl LTs (CysLTs), LTB4, and 5-Lipoxygenase (5-LOX), the synthesizing enzyme of LTs, were evaluated. CysLT and LTB4 concentrations were elevated in the hippocampus and the cerebral cortex, along with enhanced 5-LOX expression in neurons and microglia. Such changes were also associated with increased proinflammatory cytokine levels in the hippocampus and the cerebral cortex. Enhanced CysLT and LTB4 levels in the brain could also be gleaned from their concentrations in brain-derived extracellular vesicles in the circulating blood. The circulating blood in GWI rats displayed elevated proinflammatory cytokines with no alterations in CysLT and LTB4 concentrations. The results provide new evidence that a brain-specific increase in LT signaling is another adverse alteration that potentially contributes to the maintenance of chronic neuroinflammation in GWI. Therefore, drugs capable of modulating LT signaling may reduce neuroinflammation and improve cognitive function in GWI. Additional findings demonstrate that altered LT levels in the brain could be tracked efficiently by analyzing brain-derived EVs in the circulating blood.

Novel salivary antihemostatic activities of long-form D7 proteins from the malaria vector Anopheles gambiae facilitate hematophagy.

To successfully feed on blood, hematophagous arthropods must combat the host's natural hemostatic and inflammatory responses. Salivary proteins of blood-feeding insects such as mosquitoes contain compounds that inhibit these common host defenses against blood loss, including vasoconstriction, platelet aggregation, blood clotting, pain, and itching. The D7 proteins are some of the most abundantly expressed proteins in female mosquito salivary glands and have been implicated in inhibiting host hemostatic and inflammatory responses. Anopheles gambiae, the primary vector of malaria, expresses three D7 long-form and five D7 short-form proteins. Previous studies have characterized the AngaD7 short-forms, but the D7 long-form proteins have not yet been characterized in detail. Here, we characterized the A. gambiae D7 long-forms by first determining their binding kinetics to hemostatic agonists such as leukotrienes and serotonin, which are potent activators of vasoconstriction, edema formation, and postcapillary venule leakage, followed by ex vivo functional assays. We found that AngaD7L1 binds leukotriene C4 and thromboxane A2 analog U-46619; AngaD7L2 weakly binds leukotrienes B4 and D4; and AngaD7L3 binds serotonin. Subsequent functional assays confirmed AngaD7L1 inhibits U-46619-induced platelet aggregation and vasoconstriction, and AngaD7L3 inhibits serotonin-induced platelet aggregation and vasoconstriction. It is therefore possible that AngaD7L proteins counteract host hemostasis by scavenging these mediators. Finally, we demonstrate that AngaD7L2 had a dose-dependent anticoagulant effect via the intrinsic coagulation pathway by interacting with factors XII, XIIa, and XI. The uncovering of these interactions in the present study will be essential for comprehensive understanding of the vector-host biochemical interface.

P. aeruginosa augments irradiation injury via 15-lipoxygenase-catalyzed generation of 15-HpETE-PE and induction of theft-ferroptosis.

Total body irradiation (TBI) targets sensitive bone marrow hematopoietic cells and gut epithelial cells, causing their death and inducing a state of immunodeficiency combined with intestinal dysbiosis and nonproductive immune responses. We found enhanced Pseudomonas aeruginosa (PAO1) colonization of the gut leading to host cell death and strikingly decreased survival of irradiated mice. The PAO1-driven pathogenic mechanism includes theft-ferroptosis realized via (a) curbing of the host antiferroptotic system, GSH/GPx4, and (b) employing bacterial 15-lipoxygenase to generate proferroptotic signal - 15-hydroperoxy-arachidonoyl-PE (15-HpETE-PE) - in the intestines of irradiated and PAO1-infected mice. Global redox phospholipidomics of the ileum revealed that lysophospholipids and oxidized phospholipids, particularly oxidized phosphatidylethanolamine (PEox), represented the major factors that contributed to the pathogenic changes induced by total body irradiation and infection by PAO1. A lipoxygenase inhibitor, baicalein, significantly attenuated animal lethality, PAO1 colonization, intestinal epithelial cell death, and generation of ferroptotic PEox signals. Opportunistic PAO1 mechanisms included stimulation of the antiinflammatory lipoxin A4, production and suppression of the proinflammatory hepoxilin A3, and leukotriene B4. Unearthing complex PAO1 pathogenic/virulence mechanisms, including effects on the host anti/proinflammatory responses, lipid metabolism, and ferroptotic cell death, points toward potentially new therapeutic and radiomitigative targets.

The Oxidative Potential of Fine Particulate Matter and Biological Perturbations in Human Plasma and Saliva Metabolome.

Particulate oxidative potential may comprise a key health-relevant parameter of particulate matter (PM) toxicity. To identify biological perturbations associated with particulate oxidative potential and examine the underlying molecular mechanisms, we recruited 54 participants from two dormitories near and far from a congested highway in Atlanta, GA. Fine particulate matter oxidative potential ("FPMOP") levels at the dormitories were measured using dithiothreitol assay. Plasma and saliva samples were collected from participants four times for longitudinal high-resolution metabolic profiling. We conducted metabolome-wide association studies to identify metabolic signals with FPMOP. Leukotriene metabolism and galactose metabolism were top pathways associated with ≥5 FPMOP-related indicators in plasma, while vitamin E metabolism and leukotriene metabolism were found associated with most FPMOP indicators in saliva. We observed different patterns of perturbed pathways significantly associated with water-soluble and -insoluble FPMOPs, respectively. We confirmed five metabolites directly associated with FPMOP, including hypoxanthine, histidine, pyruvate, lactate/glyceraldehyde, and azelaic acid, which were implications of perturbations in acute inflammation, nucleic acid damage and repair, and energy perturbation. The unique metabolic signals were specific to FPMOP, but not PM mass, providing initial indication that FPMOP might constitute a more sensitive, health-relevant measure for elucidating etiologies related to PM2.5 exposures.

Emerging role of cysteinyl LTs in cancer.

Cysteinyl leukotrienes (CysLTs) are inflammatory lipid mediators that play a central role in the pathophysiology of several inflammatory diseases. Recently, there has been an increased interest in determining how these lipid mediators orchestrate tumour development and metastasis through promoting a pro-tumour micro-environment. Up-regulation of CysLTs receptors and CysLTs production is found in a number of cancers and has been associated with increased tumorigenesis. Understanding the molecular mechanisms underlying the role of CysLTs and their receptors in cancer progression will help investigate the potential of targeting CysLTs signalling for anti-cancer therapy. This review gives an overview of the biological effects of CysLTs and their receptors, along with current knowledge of their regulation and expression. It also provides a recent update on the molecular mechanisms that have been postulated to explain their role in tumorigenesis and on the potential of anti-CysLTs in the treatment of cancer.

Natural chalcones elicit formation of specialized pro-resolving mediators and related 15-lipoxygenase products in human macrophages.

Specialized pro-resolving mediators (SPMs) comprise lipid mediators (LMs) produced from polyunsaturated fatty acids (PUFAs) via stereoselective oxygenation particularly involving 12/15-lipoxygenases (LOXs). In contrast to pro-inflammatory LMs such as leukotrienes formed by 5-LOX and prostaglandins formed by cyclooxygenases, the SPMs have anti-inflammatory and inflammation-resolving properties. Although glucocorticoids and non-steroidal anti-inflammatory drugs (NSAIDs) that block prostaglandin production are still prime therapeutics for inflammation-related diseases despite severe side effects, novel concepts focus on SPMs as immunoresolvents for anti-inflammatory pharmacotherapy. Here, we studied the natural chalcone MF-14 and the corresponding dihydrochalcone MF-15 from Melodorum fruticosum, for modulating the biosynthesis of LM including leukotrienes, prostaglandins, SPM and their 12/15-LOX-derived precursors in human monocyte-derived macrophage (MDM) M1- and M2-like phenotypes. In MDM challenged with Staphylococcus aureus-derived exotoxins both compounds (10 µM) significantly suppressed 5-LOX product formation but increased the biosynthesis of 12/15-LOX products, especially in M2-MDM. Intriguingly, in resting M2-MDM, MF-14 and MF-15 strikingly evoked generation of 12/15-LOX products and of SPMs from liberated PUFAs, along with translocation of 15-LOX-1 to membranous compartments. Enhanced 12/15-LOX product formation by the chalcones was evident also when exogenous PUFAs were supplied, excluding increased substrate supply as sole underlying mechanism. Rather, MF-14 and MF-15 stimulate the activity of 15-LOX-1, supported by experiments with HEK293 cells transfected with either 5-LOX, 15-LOX-1 or 15-LOX-2. Together, the natural chalcone MF-14 and the dihydrochalcone MF-15 favorably modulate LM biosynthesis in human macrophages by suppressing pro-inflammatory leukotrienes but stimulating formation of SPMs by differential interference with 5-LOX and 15-LOX-1.

Ethoxy acetalated dextran-based nanocarriers accomplish efficient inhibition of leukotriene formation by a novel FLAP antagonist in human leukocytes and blood.

Leukotrienes are pro-inflammatory lipid mediators generated by 5-lipoxygenase aided by the 5-lipoxygenase-activating protein (FLAP). BRP-201, a novel benzimidazole-based FLAP antagonist, inhibits leukotriene biosynthesis in isolated leukocytes. However, like other FLAP antagonists, BRP-201 fails to effectively suppress leukotriene formation in blood, which limits its therapeutic value. Here, we describe the encapsulation of BRP-201 into poly(lactide-co-glycolide) (PLGA) and ethoxy acetalated dextran (Ace-DEX) nanoparticles (NPs), aiming to overcome these detrimental pharmacokinetic limitations and to enhance the bioactivity of BRP-201. NPs loaded with BRP-201 were produced via nanoprecipitation and the physicochemical properties of the NPs were analyzed in-depth using dynamic light scattering (size, dispersity, degradation), electrophoretic light scattering (effective charge), NP tracking analysis (size, dispersity), scanning electron microscopy (size and morphology), UV-VIS spectroscopy (drug loading), an analytical ultracentrifuge (drug release, degradation kinetics), and Raman spectroscopy (chemical attributes). Biological assays were performed to study cytotoxicity, cellular uptake, and efficiency of BRP-201-loaded NPs versus free BRP-201 to suppress leukotriene formation in primary human leukocytes and whole blood. Both PLGA- and Ace-DEX-based NPs were significantly more efficient to inhibit leukotriene formation in neutrophils versus free drug. Whole blood experiments revealed that encapsulation of BRP-201 into Ace-DEX NPs strongly increases its potency, especially upon pro-longed (≥ 5 h) incubations and upon lipopolysaccharide-challenge of blood. Finally, intravenous injection of BRP-201-loaded NPs significantly suppressed leukotriene levels in blood of mice in vivo. These results reveal the feasibility of our pharmacological approach using a novel FLAP antagonist encapsulated into Ace-DEX-based NPs with improved efficiency in blood to suppress leukotriene biosynthesis.