RESUMEN

We previously reported that leukotriene B4 (LTB4) contained in Trichomonas vaginalis-derived secretory products (TvSP) play an essential role in interleukin-8 (IL-8) production in human mast cell line (HMC-1 cells) via LTB4 receptor (BLT)-mediated Nuclear Factor-kappa B (NF-кB) activation. Dynamin, a GTPase, has been known to be involved in endocytosis of receptors for signaling of production of cytokine or chemokines. In the present study, we investigated the role of dynamin-mediated BLT1 endocytosis in TvSP-induced IL-8 production. When HMC-1 cells were transfected with BLT1 or BLT2 siRNA, TvSP-induced IL-8 production was significantly inhibited compared with that in cells transfected with control siRNA. In addition, pretreatment of HMC-1 cells with a dynamin inhibitor (Dynasore) reduced IL-8 production induced by TvSP or LTB4. TvSP- or LTB4- induced phosphorylation of NF-кB was also attenuated by pretreatment with Dynasore. After exposing HMC-1 cells to TvSP or LTB4, BLT1 was translocated from the intracellular compartments to the plasma membrane within 30 min. At 60 min after stimulation with TvSP or LTB4, BLT1 remigrated from the cell surface to intracellular areas. Pretreatment of HMC-1 cells with dynamin-2 siRNA blocked internalization of BLT1 induced by TvSP or LTB4. Co-immunoprecipitation experiments revealed that dynamin-2 strongly interacted with BLT1 60 min after stimulation with TvSP or LTB4. These results suggest that T. vaginalis-secreted LTB4 induces IL-8 production in HMC-1 cells via dynamin 2-mediated endocytosis of BLT1 and phosphorylation of NF-кB.

Asunto(s)

Dinamina II , Endocitosis , Interleucina-8 , Receptores de Leucotrieno B4 , Trichomonas vaginalis , Humanos , Interleucina-8/metabolismo , Interleucina-8/genética , Receptores de Leucotrieno B4/metabolismo , Receptores de Leucotrieno B4/genética , Endocitosis/efectos de los fármacos , Dinamina II/metabolismo , Dinamina II/genética , Línea Celular , Trichomonas vaginalis/metabolismo , Leucotrieno B4/metabolismo , Mastocitos/metabolismo , Mastocitos/inmunología , FN-kappa B/metabolismo , ARN Interferente Pequeño/metabolismo , ARN Interferente Pequeño/genética

RESUMEN

IgA antibodies play an important role in mucosal immunity. However, there is still no effective way to consistently boost mucosal IgA responses, and the factors influencing these responses are not fully understood. We observed that colonization with the murine intestinal symbiotic protozoan Tritrichomonas musculis (T.mu) boosted antigen-specific mucosal IgA responses in wild-type C57BL/6 mice. This enhancement was attributed to the accumulation of free arachidonic acid (ARA) in the intestinal lumen, which served as a signal to stimulate the production of antigen-specific mucosal IgA. When ARA was prevented from undergoing its downstream metabolic transformation using the 5-lipoxygenase inhibitor zileuton or by blocking its downstream biological signaling through genetic deletion of the Leukotriene B4 receptor 1 (Blt1), the T.mu-mediated enhancement of antigen-specific mucosal IgA production was suppressed. Moreover, both T.mu transfer and dietary supplementation of ARA augmented the efficacy of an oral vaccine against Salmonella infection, with this effect being dependent on Blt1. Our findings elucidate a tripartite circuit linking nutrients from the diet or intestinal microbiota, host lipid metabolism, and the mucosal humoral immune response.

Asunto(s)

Inmunidad Mucosa , Inmunoglobulina A , Metabolismo de los Lípidos , Ratones Endogámicos C57BL , Receptores de Leucotrieno B4 , Transducción de Señal , Animales , Metabolismo de los Lípidos/inmunología , Inmunoglobulina A/inmunología , Inmunoglobulina A/metabolismo , Transducción de Señal/inmunología , Ratones , Receptores de Leucotrieno B4/metabolismo , Receptores de Leucotrieno B4/inmunología , Ácido Araquidónico/metabolismo , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Femenino , Microbioma Gastrointestinal/inmunología , Ratones Noqueados

RESUMEN

Acute vascular injury provokes an inflammatory response, resulting in neointimal hyperplasia (NIH) and downstream pathologies. The resolution of inflammation is an active process in which specialized proresolving lipid mediators (SPM) and their receptors play a central role. We sought to examine the acute phase response of SPM and their receptors in both circulating blood and the arterial wall in a rat angioplasty model. We found that the ratio of proresolving to pro-inflammatory lipid mediators (LM) in plasma decreased sharply 1 day after vascular injury, then increased slightly by day 7, while that in arteries remained depressed. Granulocyte expression of SPM receptors ALX/FPR2 and DRV2/GPR18, and a leukotriene B4 receptor BLT1 increased postinjury, while ERV1/ChemR23 expression was reduced early and then recovered by day 7. Importantly, we show unique arterial expression patterns of SPM receptors in the acute setting, with generally low levels through day 7 that contrasted sharply with that of the pro-inflammatory CCR2 receptor. Overall, these data document acute, time-dependent changes of LM biosynthesis and SPM receptor expression in plasma, leukocytes, and artery walls following acute vascular injury. A biochemical imbalance between inflammation and resolution LM pathways appears persistent 7 days after angioplasty in this model. These findings may help guide therapeutic approaches to accelerate vascular healing and improve the outcomes of vascular interventions for patients with advanced atherosclerosis.

Asunto(s)

Ratas Sprague-Dawley , Animales , Masculino , Ratas , Lesiones del Sistema Vascular/metabolismo , Lesiones del Sistema Vascular/patología , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores de Leucotrieno B4/metabolismo , Mediadores de Inflamación/metabolismo

RESUMEN

Colorectal cancer (CRC) presents a landscape of intricate molecular dynamics. In this study, we focused on the role of the leukotriene B4 receptor (LTB4R) in CRC, exploring its significance in the disease's progression and potential therapeutic approaches. Using bioinformatics analysis of the GSE164191 and the Cancer Genome Atlas-colorectal adenocarcinoma (TCGA-COAD) datasets, we identified LTB4R as a hub gene influencing CRC prognosis. Subsequently, we examined the relationship between LTB4R expression, apoptosis, and the phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/AKT/mTOR) signaling pathway through cellular and mice experiments. Our findings revealed that LTB4R is highly expressed in CRC samples and is pivotal for determining prognosis. In vitro experiments demonstrated that silencing LTB4R significantly impeded CRC cell viability, migration, invasion, and colony formation. Correspondingly, in vivo tests indicated that LTB4R knockdown led to markedly slower tumor growth in mice models. Further in-depth investigation revealed that LTB4R knockdown significantly amplified the apoptosis in CRC cells and upregulated the expression of apoptosis-related proteins, such as caspase-3 and caspase-9, while diminishing p53 expression. Interestingly, silencing LTB4R also resulted in a significant downregulation of the PI3K/AKT/mTOR signaling pathway. Moreover, pretreatment with the PI3K activator 740Y-P only partially attenuated the effects of LTB4R knockdown on CRC cell behavior, emphasizing LTB4R's dominant influence in CRC cell dynamics and signaling pathways. LTB4R stands out as a critical factor in CRC progression, profoundly affecting cellular behavior, apoptotic responses, and the PI3K/AKT/mTOR signaling pathway. These findings not only shed light on LTB4R's role in CRC but also establish it as a potential diagnostic biomarker and a promising target for therapeutic intervention.

Asunto(s)

Apoptosis , Neoplasias Colorrectales , Fosfatidilinositol 3-Quinasas , Proteínas Proto-Oncogénicas c-akt , Receptores de Leucotrieno B4 , Transducción de Señal , Serina-Treonina Quinasas TOR , Animales , Femenino , Humanos , Masculino , Ratones , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Neoplasias Colorrectales/metabolismo , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Ratones Endogámicos BALB C , Ratones Desnudos , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptores de Leucotrieno B4/genética , Receptores de Leucotrieno B4/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Serina-Treonina Quinasas TOR/genética

RESUMEN

Invasive fungal infections are life-threatening, and neutrophils are vital cells of the innate immune system that defend against them. The role of LTA4H-LTB4-BLT1 axis in regulation of neutrophil responses to fungal infection remains poorly understood. Here, we demonstrated that the LTA4H-LTB4-BLT1 axis protects the host against Candida albicans and Aspergillus fumigatus, but not Cryptococcus neoformans infection, by regulating the antifungal activity of neutrophils. Our results show that deleting Lta4h or Blt1 substantially impairs the fungal-specific phagocytic capacity of neutrophils. Moreover, defective activation of the spleen tyrosine kinase (Syk) and extracellular signal-related kinase (ERK1/2) pathways in neutrophils accompanies this impairment. Mechanistically, BLT1 regulates CR3-mediated, ß-1,3-glucan-induced neutrophil phagocytosis, while a physical interaction with CR3 with slight influence on its dynamics is observed. Our findings thus demonstrate that the LTA4H-LTB4-BLT1 axis is essential for the phagocytic function of neutrophils in host antifungal immune response against Candida albicans and Aspergillus fumigatus.

Asunto(s)

Antifúngicos , Neutrófilos , Antifúngicos/farmacología , Leucotrieno B4/metabolismo , Receptores de Leucotrienos/metabolismo , Receptores de Leucotrieno B4/metabolismo , Antígeno CD11b/metabolismo

RESUMEN

Epithelial cells in the skin and other tissues rely on signals from their environment to maintain homeostasis and respond to injury, and GPCRs play a critical role in this communication. A better understanding of the GPCRs expressed in epithelial cells will contribute to understanding the relationship between cells and their niche and could lead to developing new therapies to modulate cell fate. This study used human primary keratinocytes as a model to investigate the specific GPCRs regulating epithelial cell proliferation and differentiation. We identified 3 key receptors-HCAR3, LTB4R, and GPR137-and found that knockdown of these receptors led to changes in numerous gene networks that are important for maintaining cell identity and promoting proliferation while inhibiting differentiation. Our study also revealed that the metabolite receptor HCAR3 regulates keratinocyte migration and cellular metabolism. Knockdown of HCAR3 led to reduced keratinocyte migration and respiration, which could be attributed to altered metabolite use and aberrant mitochondrial morphology caused by the absence of the receptor. This study contributes to understanding the complex interplay between GPCR signaling and epithelial cell fate decisions.

Asunto(s)

Movimiento Celular , Proliferación Celular , Respiración de la Célula , Queratinocitos , Receptores Acoplados a Proteínas G , Humanos , Queratinocitos/metabolismo , Queratinocitos/citología , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Respiración de la Célula/fisiología , Transducción de Señal , Diferenciación Celular , Células Cultivadas , Receptores de Leucotrieno B4/metabolismo , Receptores de Leucotrieno B4/genética , Células Epiteliales/metabolismo , Receptores Nicotínicos

RESUMEN

Chronic inflammation is the underlying cause of many diseases, including type 1 diabetes, obesity, and non-alcoholic fatty liver disease. Macrophages are continuously recruited to tissues during chronic inflammation where they exacerbate or resolve the pro-inflammatory environment. Although leukotriene B4 receptor 2 (BLT2) has been characterized as a low affinity receptor to several key eicosanoids and chemoattractants, its precise roles in the setting of inflammation and macrophage function remain incompletely understood. Here we used zebrafish and mouse models to probe the role of BLT2 in macrophage function during inflammation. We detected BLT2 expression in bone marrow derived and peritoneal macrophages of mouse models. Transcriptomic analysis of Ltb4r2-/- and WT macrophages suggested a role for BLT2 in macrophage migration, and studies in vitro confirmed that whereas BLT2 does not mediate macrophage polarization, it is required for chemotactic function, possibly mediated by downstream genes Ccl5 and Lgals3. Using a zebrafish model of tailfin injury, we demonstrated that antisense morpholino-mediated knockdown of blt2a or chemical inhibition of BLT2 signaling impairs macrophage migration. We further replicated these findings in zebrafish models of islet injury and liver inflammation. Moreover, we established the applicability of our zebrafish findings to mammals by showing that macrophages of Ltb4r2-/- mice have defective migration during lipopolysaccharide stimulation in vivo. Collectively, our results demonstrate that BLT2 mediates macrophage migration during inflammation, which implicates it as a potential therapeutic target for inflammatory pathologies.

Asunto(s)

Movimiento Celular , Macrófagos , Receptores de Leucotrieno B4 , Animales , Ratones , Inflamación/genética , Inflamación/metabolismo , Leucotrieno B4/genética , Leucotrieno B4/metabolismo , Macrófagos/citología , Macrófagos/metabolismo , Receptores de Leucotrieno B4/genética , Receptores de Leucotrieno B4/metabolismo , Pez Cebra/genética , Pez Cebra/metabolismo

RESUMEN

G protein-coupled receptors (GPCRs) utilize complex cellular systems to respond to diverse ligand concentrations. By taking BLT1, a GPCR for leukotriene B4 (LTB4 ), as a model, our previous work elucidated that this system functions through the modulation of phosphorylation status on two specific residues: Thr308 and Ser310 . Ser310 phosphorylation occurs at a lower LTB4 concentration than Thr308 , leading to a shift in ligand affinity from a high-to-low state. However, the implications of BLT1 phosphorylation in signal transduction processes or the underlying mechanisms have remained unclear. Here, we identify the sequential BLT1-engaged conformations of ß-arrestin and subsequent alterations in signal transduction. Stimulation of the high-affinity BLT1 with LTB4 induces phosphorylation at Ser310 via the ERK1/2-GRK pathway, resulting in a ß-arrestin-bound low-affinity state. This configuration, referred to as the "low-LTB4 -induced complex," necessitates the finger loop region and the phosphoinositide-binding motif of ß-arrestins to interact with BLT1 and deactivates the ERK1/2 signaling. Under high LTB4 concentrations, the low-affinity BLT1 again binds to the ligand and triggers the generation of the low-LTB4 -induced complex into a different form termed "high-LTB4 -induced complex." This change is propelled by The308 -phosphorylation-dependent basal phosphorylation by PKCs. Within the high-LTB4 -induced complex, ß-arrestin adapts a unique configuration that involves additional N domain interaction to the low-affinity BLT1 and stimulates the PI3K/AKT pathway. We propose that the stepwise phosphorylation of BLT1 defines the formation of complex assemblies, wherein ß-arrestins perform distinct functions.

Asunto(s)

Fosfatidilinositol 3-Quinasas , Transducción de Señal , Fosforilación , beta-Arrestinas/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Ligandos , beta-Arrestina 1/metabolismo , Receptores de Leucotrieno B4/metabolismo , Leucotrieno B4/metabolismo

RESUMEN

Leukotriene B4 (LTB4) is a potent chemoattractant that can recruit and activate immune cells such as neutrophils, eosinophils, and monocytes to sites of inflammation. Excessive production of LTB4 has been linked to acute and chronic inflammatory diseases, including asthma, rheumatoid arthritis, and psoriasis. Inhibiting the binding of LTB4 to its receptors, BLT1 and BLT2, is a potential strategy for treating these conditions. While several BLT1 antagonists have been developed for clinical trials, most have failed due to efficacy and safety issues. Therefore, discovering selective BLT2 antagonists could improve our understanding of the distinct functions of BLT1 and BLT2 receptors and their pharmacological implications. In this study, we aimed to discover novel BLT2 antagonists by synthesizing a series of biphenyl analogues based on a BLT2 selective agonist, CAY10583. Among the synthesized compounds, 15b was found to selectively inhibit the chemotaxis of CHO-BLT2 cells with an IC50 value of 224 nM without inhibiting the chemotaxis of CHO-BLT1 cells. 15b also inhibited the binding of LTB4 and BLT2 with a Ki value of 132 nM. Furthermore, 15b had good metabolic stability in liver microsomes and moderate bioavailability (F = 34%) in in vivo PK studies. 15b also showed in vivo efficacy in a mouse model of asthma, reducing airway hyperresponsiveness by 59% and decreasing Th2 cytokines by up to 46%. Our study provides a promising lead for the development of selective BLT2 antagonists as potential therapeutics for inflammatory airway diseases such as asthma and chronic obstructive pulmonary disease.

Asunto(s)

Artritis Reumatoide , Asma , Ratones , Cricetinae , Animales , Leucotrieno B4 , Asma/tratamiento farmacológico , Inflamación , Células CHO , Receptores de Leucotrieno B4/metabolismo

RESUMEN

Specialized pro-resolving mediators (SPMs) and especially Resolvin E1 (RvE1) can actively terminate inflammation and promote healing during lung diseases such as acute respiratory distress syndrome (ARDS). Although ARDS primarily affects the lung, many ARDS patients also develop neurocognitive impairments. To investigate the connection between the lung and brain during ARDS and the therapeutic potential of SPMs and its derivatives, fat-1 mice were crossbred with RvE1 receptor knockout mice. ARDS was induced in these mice by intratracheal application of lipopolysaccharide (LPS, 10 µg). Mice were sacrificed at 0 h, 4 h, 24 h, 72 h, and 120 h post inflammation, and effects on the lung, liver, and brain were assessed by RT-PCR, multiplex, immunohistochemistry, Western blot, and LC-MS/MS. Protein and mRNA analyses of the lung, liver, and hypothalamus revealed LPS-induced lung inflammation increased inflammatory signaling in the hypothalamus despite low signaling in the periphery. Neutrophil recruitment in different brain structures was determined by immunohistochemical staining. Overall, we showed that immune cell trafficking to the brain contributed to immune-to-brain communication during ARDS rather than cytokines. Deficiency in RvE1 receptors and enhanced omega-3 polyunsaturated fatty acid levels (fat-1 mice) affect lung-brain interaction during ARDS by altering profiles of several inflammatory and lipid mediators and glial activity markers.

Asunto(s)

Ácidos Grasos Omega-3 , Síndrome de Dificultad Respiratoria , Animales , Ratones , Encéfalo , Cromatografía Liquida , Inflamación , Lipopolisacáridos/toxicidad , Pulmón , Ratones Noqueados , Receptores de Leucotrieno B4 , Síndrome de Dificultad Respiratoria/inducido químicamente , Síndrome de Dificultad Respiratoria/genética , Espectrometría de Masas en Tándem

RESUMEN

Leukotriene B4 (LTB4) is a potent lipid chemoattractant driving inflammatory responses during host defense, allergy, autoimmune and metabolic diseases. Gradients of LTB4 orchestrate leukocyte recruitment and swarming to sites of tissue damage and infection. How LTB4 gradients form and spread in live tissues to regulate these processes remains largely elusive due to the lack of suitable tools for monitoring LTB4 levels in vivo. Here, we develop GEM-LTB4, a genetically encoded green fluorescent LTB4 biosensor based on the human G-protein-coupled receptor BLT1. GEM-LTB4 shows high sensitivity, specificity and a robust fluorescence increase in response to LTB4 without affecting downstream signaling pathways. We use GEM-LTB4 to measure ex vivo LTB4 production of murine neutrophils. Transgenic expression of GEM-LTB4 in zebrafish allows the real-time visualization of both exogenously applied and endogenously produced LTB4 gradients. GEM-LTB4 thus serves as a broadly applicable tool for analyzing LTB4 dynamics in various experimental systems and model organisms.

Asunto(s)

Leucotrieno B4 , Pez Cebra , Humanos , Ratones , Animales , Leucotrieno B4/metabolismo , Pez Cebra/genética , Pez Cebra/metabolismo , Receptores de Leucotrieno B4/genética , Receptores de Leucotrieno B4/metabolismo , Neutrófilos , Transducción de Señal

RESUMEN

Leukotriene B4 receptor type 1 (BLT1), a high-affinity receptor for leukotriene B4 (LTB4), plays an important role in inflammatory responses, including allergic airway inflammation. In this study, we examined the effect of genetic BLT1 deletion (BLT1KO) on ovalbumin (OVA)-induced allergic enteritis in mice to determine the pathogenic role of LTB4/BLT1 in allergic enteritis, a gastrointestinal form of food allergy. Repeated oral OVA challenges after sensitization with OVA and aluminium potassium sulphate induced allergic enteritis, characterized by systemic allergic symptoms (scratching, immobility and swelling), diarrhoea, colonic oedema and colonic goblet cell hyperplasia, accompanied by increased colonic peroxidase activity, colonic inflammatory cytokine expression and increased serum OVA-specific IgE levels. The severity of enteritis was significantly attenuated in BLT1KO mice compared with wild-type (WT) mice, without an increase in serum OVA-specific IgE levels. The accumulation of neutrophils, eosinophils, M2-macrophages, dendritic cells, CD4+ T cells and mast cells was observed in the colonic mucosa of allergic enteritis, and such accumulation was significantly lower in BLT1KO mice than in WT mice. BLT1 expression was upregulated and colocalized mostly in neutrophils and partly in eosinophils and dendritic cells in the colonic mucosa of allergic enteritis. These findings indicate that BLT1 deficiency ameliorates OVA-induced allergic enteritis in mice and that LTB4/BLT1 contributes to neutrophil and eosinophil accumulation in the allergic colonic mucosa. Therefore, BLT1 is a promising drug target for treating food allergies.

Asunto(s)

Leucotrieno B4 , Receptores de Leucotrieno B4 , Ratones , Animales , Ovalbúmina , Receptores de Leucotrieno B4/genética , Receptores de Leucotrieno B4/metabolismo , Leucotrieno B4/metabolismo , Ratones Noqueados , Inflamación , Inmunoglobulina E

RESUMEN

Leukotriene B4 (LTB4) is a lipid mediator rapidly generated from arachidonic acid in response to various stimuli. This lipid mediator exerts its biological activities by binding to cognate receptors. Two LTB4 receptors have been cloned; BLT1 and BLT2 as a high- and a low-affinity receptors, respectively. In numerous analyses, physiological and pathophysiological importance of LTB4 and cognate receptors in various diseases has been clarified. For example, disruption of the BLT1 gene or treatment with blockers for this receptor reduced various diseases such as rheumatoid arthritis and bronchial asthma in mice, in contrast BLT2 deficiency facilitated several diseases in the small intestine and the skin. These data support the idea that BLT1 blockers and BLT2 agonists could be useful for the cure of these diseases. Thus, various drugs targeting each receptor are being developed by many pharmaceutical companies. In this review, we focus on our current knowledge of the biosynthesis and physiological roles of LTB4 through cognate receptors. We further describe the effects of these receptor deficiencies on several pathophysiological conditions, including the potential of LTB4 receptors as therapeutic targets for the cure of the diseases. Moreover, current information on the structure and post-translational modification of BLT1 and BLT2 is discussed.

Asunto(s)

Artritis Reumatoide , Leucotrieno B4 , Ratones , Animales , Leucotrieno B4/genética , Leucotrieno B4/metabolismo , Leucotrieno B4/farmacología , Piel/metabolismo , Receptores de Leucotrieno B4/genética , Receptores de Leucotrieno B4/metabolismo

RESUMEN

Hepatocellular carcinoma (HCC) is the 3rd most deadly malignancy. Activated hepatic stellate cells (aHSC) give rise to cancer-associated fibroblasts in HCC and are considered a potential therapeutic target. Here we report that selective ablation of stearoyl CoA desaturase-2 (Scd2) in aHSC globally suppresses nuclear CTNNB1 and YAP1 in tumors and tumor microenvironment and prevents liver tumorigenesis in male mice. Tumor suppression is associated with reduced leukotriene B4 receptor 2 (LTB4R2) and its high affinity oxylipin ligand, 12-hydroxyheptadecatrienoic acid (12-HHTrE). Genetic or pharmacological inhibition of LTB4R2 recapitulates CTNNB1 and YAP1 inactivation and tumor suppression in culture and in vivo. Single cell RNA sequencing identifies a subset of tumor-associated aHSC expressing Cyp1b1 but no other 12-HHTrE biosynthetic genes. aHSC release 12-HHTrE in a manner dependent on SCD and CYP1B1 and their conditioned medium reproduces the LTB4R2-mediated tumor-promoting effects of 12-HHTrE in HCC cells. CYP1B1-expressing aHSC are detected in proximity of LTB4R2-positive HCC cells and the growth of patient HCC organoids is blunted by LTB4R2 antagonism or knockdown. Collectively, our findings suggest aHSC-initiated 12-HHTrE-LTB4R2-CTNNB1-YAP1 pathway as a potential HCC therapeutic target.

Asunto(s)

Carcinoma Hepatocelular , Neoplasias Hepáticas , Animales , Masculino , Ratones , beta Catenina/metabolismo , Carcinogénesis/genética , Carcinogénesis/metabolismo , Carcinoma Hepatocelular/metabolismo , Ácido Graso Desaturasas , Células Estrelladas Hepáticas/metabolismo , Neoplasias Hepáticas/metabolismo , Receptores de Leucotrieno B4/genética , Receptores de Leucotrieno B4/metabolismo , Microambiente Tumoral

RESUMEN

New leukotriene B4 (LTB4) antagonists have been synthesized that can be considered as potential anti-inflammatory drugs. Structures containing the dioxygenated nucleus of 1,4-benzodioxine constitute a potential group of leukotriene B4 (LTB4) antagonists. The objective of this study was to access efficient and selective LTB4 antagonists as a way to elucidate the role of LTB4 in inflammatory processes and therefore allow the development of new types of structures based on 1,4-benzodioxine. Forty-one new 1,4-benzodioxine molecules substituted at different positions of the heterocyclic nucleus were synthesized to determine the minimum structural requirements by studying structure-activity relationships. Eighteen of them were tested in vitro and in vivo for their anti-inflammatory activity related to the antagonist character of LTB4. Pharmacological tests have shown satisfactory in vitro activity for compounds 24b, 24c and 24e with IC50's of 288, 439, 477 nM respectively. The results of the in vivo tests, carried out with the compound that presented greater activity in the in vitro tests 24b, have shown significant anti-inflammatory properties.

Asunto(s)

Leucotrieno B4 , Neutrófilos , Antagonistas de Leucotrieno , Receptores de Leucotrieno B4 , Antiinflamatorios/farmacología

RESUMEN

Leukotriene B4 (LTB4 ) was recognized as an arachidonate-derived chemotactic factor for inflammatory cells and an important drug target even before the molecular identification of its receptors. We cloned the high- and low-affinity LTB4 receptors, BLT1 and BLT2, respectively, and examined their functions by generating and studying gene-targeted mice. BLT1 is involved in the pathogenesis of various inflammatory and immune diseases, including asthma, psoriasis, contact dermatitis, allergic conjunctivitis, age-related macular degeneration, and immune complex-mediated glomerulonephritis. Meanwhile, BLT2 is a high-affinity receptor for 12-hydroxyheptadecatrienoic acid, which is involved in the maintenance of dermal and intestinal barrier function, and the acceleration of skin and corneal wound healing. Thus, BLT1 antagonists and BLT2 agonists are promising candidates in the treatment of inflammatory diseases.

Asunto(s)

Asma , Leucotrieno B4 , Ratones , Humanos , Animales , Piel , Cicatrización de Heridas , Receptores de Leucotrieno B4/genética

RESUMEN

Crescent formation is the most important pathological finding that defines the prognosis of nephritis. Although neutrophils are known to play an important role in the progression of crescentic glomerulonephritis, such as anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis, the key chemoattractant for neutrophils in ANCA-associated glomerulonephritis has not been identified. Here, we demonstrate that a lipid chemoattractant, leukotriene B4 (LTB4 ), and its receptor BLT1 are primarily involved in disease pathogenesis in a mouse model of immune complex-mediated crescentic glomerulonephritis. Circulating neutrophils accumulated into glomeruli within 1 h after disease onset, which was accompanied by LTB4 accumulation in the kidney cortex, leading to kidney injury. LTB4 was produced by cross-linking of Fc gamma receptors on neutrophils. Mice deficient in BLT1 or LTB4 biosynthesis exhibited suppressed initial neutrophil infiltration and subsequent thrombotic glomerulonephritis and renal fibrosis. Depletion of neutrophils before, but not after, disease onset prevented proteinuria and kidney injury, indicating the essential role of neutrophils in the early phase of glomerulonephritis. Administration of a BLT1 antagonist before and after disease onset almost completely suppressed induction of glomerulonephritis. Finally, we found that the glomeruli from patients with ANCA-associated glomerulonephritis contained more BLT1-positive cells than glomeruli from patients with other etiologies. Taken together, the LTB4 -BLT1 axis is the key driver of neutrophilic glomerular inflammation, and will be a novel therapeutic target for the crescentic glomerulonephritis.

Asunto(s)

Glomerulonefritis , Leucotrieno B4 , Receptores de Leucotrieno B4 , Animales , Ratones , Anticuerpos Anticitoplasma de Neutrófilos , Complejo Antígeno-Anticuerpo , Factores Quimiotácticos , Glomerulonefritis/patología , Neutrófilos/patología , Receptores de Leucotrieno B4/metabolismo

RESUMEN

12(S)-hydroxyheptadecatrienoic acid (12-HHT) is a bioactive fatty acid synthesized from arachidonic acid via the cyclooxygenase pathway and serves as an endogenous ligand for the low-affinity leukotriene B4 receptor 2 (BLT2). Although the 12-HHT/BLT2 axis contributes to the maintenance of epithelial homeostasis, 12-HHT metabolism under physiological conditions is unclear. In this study, 12-keto-heptadecatrienoic acid (12-KHT) and 10,11-dihydro-12-KHT (10,11dh-12-KHT) were detected as 12-HHT metabolites in the human megakaryocytic cell line MEG01s. We found that 12-KHT and 10,11dh-12-KHT are produced from 12-HHT by 15-hydroxyprostaglandin dehydrogenase (15-PGDH) and prostaglandin reductase 1 (PTGR1), key enzymes in the degradation of prostaglandins, respectively. The 15-PGDH inhibitor SW033291 completely suppressed the production of 12-KHT and 10,11dh-12-KHT in MEG01s cells, resulting in a 9-fold accumulation of 12-HHT. 12-KHT and 10,11dh-12-KHT were produced in mouse skin wounds, and the levels were significantly suppressed by SW033291. Surprisingly, the agonistic activities of 12-KHT and 10,11dh-12-KHT on BLT2 were comparable to that of 12-HHT. Taken together, 12-HHT is metabolized into 12-KHT by 15-PGDH, and then 10,11dh-12-KHT by PTGR1 without losing the agonistic activity.

Asunto(s)

Ácidos Grasos Insaturados , Receptores de Leucotrieno B4 , Ratones , Humanos , Animales , Receptores de Leucotrieno B4/metabolismo , Ligandos , Ácidos Grasos Insaturados/metabolismo , Leucotrieno B4/metabolismo

RESUMEN

BACKGROUND AND PURPOSE: Epoxyeicosatrienoic acids (EETs) and other epoxy fatty acids (EpFA) are lipid mediators that are rapidly inactivated by soluble epoxide hydrolase (sEH). Uncontrolled and chronic inflammatory disorders fail to sufficiently activate endogenous regulatory pathways, including the production of specialized pro-resolving mediators (SPMs). Here, we addressed the relationship between SPMs and the EET/sEH axis and explored the effects of sEH inhibition on resolving macrophage phenotype. EXPERIMENTAL APPROACH: Mice were treated with a sEH inhibitor, EETs, or sEH inhibitor + EETs (combination) before ligature placement to induce experimental periodontitis. Using RT-qPCR, gingival samples were used to examine SPM receptors and osteolytic and inflammatory biomarkers. Maxillary alveolar bone loss was quantified by micro-CT and methylene blue staining. SPM levels were analysed by salivary metabolo-lipidomics. Gingival macrophage phenotype plasticity was determined by RT-qPCR and flow cytometry. Effects of sEH inhibition on macrophage polarization and SPM production were assessed with bone marrow-derived macrophages (BMDMs). KEY RESULTS: Pharmacological inhibition of sEH suppressed bone resorption and the inflammatory cytokine storm in experimental periodontitis. Lipidomic analysis revealed that sEH inhibition augmented levels of LXA4, RvE1, RvE2, and 4-HDoHE, concomitant with up-regulation of LTB4R1, CMKLR1/ChemR23, and ALX/FPR2 SPM receptors. Notably, there is an impact on gingival macrophage plasticity was affected suggesting an inflammation resolving phenotype with sEH inhibition. In BMDMs, sEH inhibition reduced inflammatory macrophage activation, and resolving macrophages were triggered to produce SPMs. CONCLUSION AND IMPLICATIONS: Pharmacological sEH inhibition increased SPM synthesis associated with resolving macrophages, suggesting a potential target to control osteolytic inflammatory disorders.

Asunto(s)

Epóxido Hidrolasas , Periodontitis , Animales , Ratones , Epóxido Hidrolasas/metabolismo , Macrófagos/metabolismo , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Eicosanoides/metabolismo , Periodontitis/tratamiento farmacológico , Periodontitis/metabolismo , Receptores de Leucotrieno B4/metabolismo

RESUMEN

BACKGROUND AND AIMS: NAFLD is the most prevalent chronic liver disease worldwide and has emerged as a serious public health issue with no approved treatment. The development of NAFLD is strongly associated with hepatic lipid content, and patients with NAFLD have significantly higher rates of hepatic de novo lipogenesis (DNL) than lean individuals. Leukotriene B4 (LTB4), a metabolite of arachidonic acid, is dramatically increased in obesity and plays important role in proinflammatory cytokine production and insulin resistance. But the role of liver LTB4/LTB4 receptor 1 (Ltb4r1) in lipid metabolism is unclear. APPROACH AND RESULTS: Hepatocyte-specific knockout (HKO) of Ltb4r1 improved hepatic steatosis and systemic insulin resistance in both diet-induced and genetically induced obese mice. The mRNA level of key enzymes involved in DNL and fatty acid esterification decreased in Ltb4r1 HKO obese mice. LTB4/Ltb4r1 directly promoted lipogenesis in HepG2 cells and primary hepatocytes. Mechanically, LTB4/Ltb4r1 promoted lipogenesis by activating the cAMP-protein kinase A (PKA)-inositol-requiring enzyme 1α (IRE1α)-spliced X-box-binding protein 1 (XBP1s) axis in hepatocytes, which in turn promoted the expression of lipogenesis genes regulated by XBP1s. In addition, Ltb4r1 suppression through the Ltb4r1 inhibitor or lentivirus-short hairpin RNA delivery alleviated the fatty liver phenotype in obese mice. CONCLUSIONS: LTB4/Ltb4r1 promotes hepatocyte lipogenesis directly by activating PKA-IRE1α-XBP1s to promote lipogenic gene expression. Inhibition of hepatocyte Ltb4r1 improved hepatic steatosis and insulin resistance. Ltb4r1 is a potential therapeutic target for NAFLD.

Asunto(s)

Resistencia a la Insulina , Enfermedad del Hígado Graso no Alcohólico , Ratones , Animales , Enfermedad del Hígado Graso no Alcohólico/etiología , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Receptores de Leucotrieno B4/metabolismo , Leucotrieno B4/efectos adversos , Leucotrieno B4/metabolismo , Ratones Obesos , Endorribonucleasas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Hepatocitos/metabolismo , Hígado/metabolismo , Obesidad/complicaciones , Obesidad/genética , Lipogénesis/fisiología , Dieta Alta en Grasa