RESUMO
Interaction of mast cells (MCs) with fibroblasts is essential for MC maturation within tissue microenvironments, although the underlying mechanism is incompletely understood. Through a phenotypic screening of >30 mouse lines deficient in lipid-related genes, we found that deletion of the lysophosphatidic acid (LPA) receptor LPA1, like that of the phospholipase PLA2G3, the prostaglandin D2 (PGD2) synthase L-PGDS, or the PGD2 receptor DP1, impairs MC maturation and thereby anaphylaxis. Mechanistically, MC-secreted PLA2G3 acts on extracellular vesicles (EVs) to supply lysophospholipids, which are converted by fibroblast-derived autotaxin (ATX) to LPA. Fibroblast LPA1 then integrates multiple pathways required for MC maturation by facilitating integrin-mediated MC-fibroblast adhesion, IL-33-ST2 signaling, L-PGDS-driven PGD2 generation, and feedforward ATX-LPA1 amplification. Defective MC maturation resulting from PLA2G3 deficiency is restored by supplementation with LPA1 agonists or PLA2G3-modified EVs. Thus, the lipid-orchestrated paracrine circuit involving PLA2G3-driven lysophospholipid, eicosanoid, integrin, and cytokine signaling fine-tunes MC-fibroblast communication, ensuring MC maturation.
Assuntos
Anafilaxia , Fibroblastos , Lisofosfolipídeos , Mastócitos , Camundongos Knockout , Comunicação Parácrina , Diester Fosfórico Hidrolases , Receptores de Ácidos Lisofosfatídicos , Transdução de Sinais , Animais , Mastócitos/imunologia , Mastócitos/metabolismo , Anafilaxia/imunologia , Anafilaxia/metabolismo , Camundongos , Fibroblastos/metabolismo , Lisofosfolipídeos/metabolismo , Receptores de Ácidos Lisofosfatídicos/metabolismo , Receptores de Ácidos Lisofosfatídicos/genética , Diester Fosfórico Hidrolases/metabolismo , Diester Fosfórico Hidrolases/genética , Prostaglandina D2/metabolismo , Vesículas Extracelulares/metabolismo , Interleucina-33/metabolismo , Oxirredutases Intramoleculares/metabolismo , Oxirredutases Intramoleculares/genética , Receptores de Prostaglandina/metabolismo , Receptores de Prostaglandina/genética , Diferenciação Celular , Camundongos Endogâmicos C57BL , Proteína 1 Semelhante a Receptor de Interleucina-1 , LipocalinasRESUMO
Among the phospholipase A2 (PLA2 ) superfamily, which typically catalyzes the sn-2 hydrolysis of phospholipids to yield fatty acids and lysophospholipids, the secreted PLA2 (sPLA2 ) family contains 11 isoforms in mammals. Individual sPLA2 s have unique enzymatic specificity toward fatty acids and polar heads of phospholipid substrates and display distinct tissue/cellular distributions, suggesting their distinct physiological functions. Recent studies using knockout and/or transgenic mice for a full set of sPLA2 s have revealed their roles in modulation of immunity and related disorders. Application of mass spectrometric lipidomics to these mice has enabled to identify target substrates and products of individual sPLA2 s in given tissue microenvironments. sPLA2 s hydrolyze not only phospholipids in the plasma membrane of activated, damaged or dying mammalian cells, but also extracellular phospholipids such as those in extracellular vesicles, microbe membranes, lipoproteins, surfactants, and dietary phospholipids, thereby exacerbating or ameliorating various diseases. The actions of sPLA2 s are dependent on, or independent of, the generation of fatty acid- or lysophospholipid-derived lipid mediators according to the pathophysiological contexts. In this review, we make an overview of our current understanding of the roles of individual sPLA2 s in various immune responses and associated diseases.
Assuntos
Fosfolipases A2 Secretórias , Animais , Humanos , Camundongos , Fosfolipases A2 Secretórias/metabolismo , Ácidos Graxos , Camundongos Transgênicos , Membrana Celular/metabolismo , Mamíferos/metabolismoRESUMO
Microenvironment-based alterations in phenotypes of mast cells influence the susceptibility to anaphylaxis, yet the mechanisms underlying proper maturation of mast cells toward an anaphylaxis-sensitive phenotype are incompletely understood. Here we report that PLA2G3, a mammalian homolog of anaphylactic bee venom phospholipase A2, regulates this process. PLA2G3 secreted from mast cells is coupled with fibroblastic lipocalin-type PGD2 synthase (L-PGDS) to provide PGD2, which facilitates mast-cell maturation via PGD2 receptor DP1. Mice lacking PLA2G3, L-PGDS or DP1, mast cell-deficient mice reconstituted with PLA2G3-null or DP1-null mast cells, or mast cells cultured with L-PGDS-ablated fibroblasts exhibited impaired maturation and anaphylaxis of mast cells. Thus, we describe a lipid-driven PLA2G3-L-PGDS-DP1 loop that drives mast cell maturation.
Assuntos
Fosfolipases A2 do Grupo III/imunologia , Mastócitos/imunologia , Comunicação Parácrina/imunologia , Prostaglandina D2/imunologia , Receptores de Prostaglandina/imunologia , Animais , Western Blotting , Células da Medula Óssea/imunologia , Células da Medula Óssea/metabolismo , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Células Cultivadas , Fibroblastos/citologia , Fibroblastos/imunologia , Fibroblastos/metabolismo , Perfilação da Expressão Gênica , Fosfolipases A2 do Grupo III/genética , Fosfolipases A2 do Grupo III/metabolismo , Humanos , Oxirredutases Intramoleculares/genética , Oxirredutases Intramoleculares/imunologia , Oxirredutases Intramoleculares/metabolismo , Lipocalinas/genética , Lipocalinas/imunologia , Lipocalinas/metabolismo , Mastócitos/metabolismo , Mastócitos/ultraestrutura , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Eletrônica de Transmissão , Análise de Sequência com Séries de Oligonucleotídeos , Comunicação Parácrina/genética , Prostaglandina D2/metabolismo , Receptores de Prostaglandina/genética , Receptores de Prostaglandina/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Asthma is a chronic inflammatory disease of the airways characterized by recurrent episodes of airway obstruction, hyperresponsiveness, remodeling, and eosinophilia. Phospholipase A2 s (PLA2 s), which release fatty acids and lysophospholipids from membrane phospholipids, have been implicated in exacerbating asthma by generating pro-asthmatic lipid mediators, but an understanding of the association between individual PLA2 subtypes and asthma is still incomplete. Here, we show that group III-secreted PLA2 (sPLA2 -III) plays an ameliorating, rather than aggravating, role in asthma pathology. In both mouse and human lungs, sPLA2 -III was expressed in bronchial epithelial cells and decreased during the asthmatic response. In an ovalbumin (OVA)-induced asthma model, Pla2g3-/- mice exhibited enhanced airway hyperresponsiveness, eosinophilia, OVA-specific IgE production, and type 2 cytokine expression as compared to Pla2g3+/+ mice. Lipidomics analysis showed that the pulmonary levels of several lysophospholipids, including lysophosphatidylcholine, lysophosphatidylethanolamine, and lysophosphatidic acid (LPA), were decreased in OVA-challenged Pla2g3-/- mice relative to Pla2g3+/+ mice. LPA receptor 2 (LPA2 ) agonists suppressed thymic stromal lymphopoietin (TSLP) expression in bronchial epithelial cells and reversed airway hyperresponsiveness and eosinophilia in Pla2g3-/- mice, suggesting that sPLA2 -III negatively regulates allergen-induced asthma at least by producing LPA. Thus, the activation of the sPLA2 -III-LPA pathway may be a new therapeutic target for allergic asthma.
Assuntos
Asma , Eosinofilia , Fosfolipases A2 Secretórias , Hipersensibilidade Respiratória , Humanos , Animais , Camundongos , Lisofosfolipídeos , Fosfolipases A2 Secretórias/genética , CitocinasRESUMO
BACKGROUND: Although hypercholesterolemia reportedly counteracts lymphocyte trafficking across lymphatic vessels, the roles of lymphatic endothelial cells (LECs) in the lymphocyte regulations remain unclear. Previous studies showed that calpain-an intracellular modulatory protease-interferes with leukocyte dynamics in the blood microcirculation and is associated with hypercholesterolemic dysfunction in vascular endothelial cells. METHODS: This study investigated whether the calpain systems in LECs associate with the LEC-lymphocyte interaction under hypercholesterolemia using gene-targeted mice. RESULTS: Lipidomic analysis in hypercholesterolemic mice showed that several lysophospholipids, including lysophosphatidic acid, accumulated in the lymphatic environment. Lysophosphatidic acid enables the potentiation of calpain systems in cultured LECs, which limits their ability to stabilize regulatory T cells (Treg) without altering Th1/Th2 (T helper type1/2) subsets. This occurs via the proteolytic degradation of MEKK1 (mitogen-activated protein kinase kinase kinase 1) and the subsequent inhibition of TGF (transforming growth factor)-ß1 production in LECs. Targeting calpain systems in LECs expanded Tregs in the blood circulation and reduced aortic atherosclerosis in hypercholesterolemic mice, concomitant with the reduction of proinflammatory macrophages in the lesions. Treg expansion in the blood circulation and atheroprotection in calpain-targeted mice was prevented by the administration of TGF-ß type-I receptor inhibitor. Moreover, lysophosphatidic acid-induced calpain overactivation potentiated the IL (interleukin)-18/NF-κB (nuclear factor κB)/VCAM1 (vascular cell adhesion molecule 1) axis in LECs, thereby inhibiting lymphocyte mobility on the cells. Indeed, VCAM1 in LECs was upregulated in hypercholesterolemic mice and human cases of coronary artery disease. Neutralization of VCAM1 or targeting LEC calpain systems recovered afferent Treg transportation via lymphatic vessels in mice. CONCLUSIONS: Calpain systems in LECs have a key role in controlling Treg stability and trafficking under hypercholesterolemia.
Assuntos
Hipercolesterolemia , Vasos Linfáticos , Camundongos , Humanos , Animais , Células Endoteliais/metabolismo , Linfócitos T Reguladores/metabolismo , Calpaína/metabolismo , Hipercolesterolemia/complicações , Hipercolesterolemia/genética , Hipercolesterolemia/metabolismo , Vasos Linfáticos/metabolismo , NF-kappa B/metabolismoRESUMO
Psoriasis is an inflammatory disorder characterized by keratinocyte hyper-proliferation and Th17-type immune responses. However, the roles of bioactive lipids and the regulation of their biosynthesis in this chronic skin disease are not fully understood. Herein, we show that group IVE cytosolic phospholipase A2 (cPLA2 ε/PLA2G4E) plays a counterregulatory role against psoriatic inflammation by producing the anti-inflammatory lipid N-acylethanolamine (NAE). Lipidomics analysis of mouse skin revealed that NAE species and their precursors (N-acyl-phosphatidylethanolamine and glycerophospho-N-acylethanolamine) were robustly increased in parallel with the ongoing process of imiquimod (IMQ)-induced psoriasis, accompanied by a marked upregulation of cPLA2 ε in epidermal keratinocytes. Genetic deletion of cPLA2 ε exacerbated IMQ-induced ear swelling and psoriatic marker expression, with a dramatic reduction of NAE-related lipids in IMQ-treated, and even normal, skin. Stimulation of cultured human keratinocytes with psoriatic cytokines concomitantly increased PLA2G4E expression and NAE production, and supplementation with NAEs significantly attenuated the cytokine-induced upregulation of the psoriatic marker S100A9. Increased expression of cPLA2 ε was also evident in the epidermis of psoriatic patients. These findings reveal for the first time the in vivo role of cPLA2 ε, which is highly induced in the keratinocytes of the psoriatic skin, promotes the biosynthesis of NAE-related lipids, and contributes to limiting psoriatic inflammation.
Assuntos
Psoríase , Animais , Anti-Inflamatórios/uso terapêutico , Anticorpos , Citocinas/metabolismo , Etanolaminas , Humanos , Imiquimode , Inflamação , Lipídeos/efeitos adversos , Camundongos , Fosfolipases/uso terapêutico , Psoríase/induzido quimicamente , Psoríase/tratamento farmacológicoRESUMO
Milk lipids are an important energy source for infants, but the composition of milk lipids has not yet been clarified in detail. In this study, we analyzed free fatty acids and their metabolites in milk from humans and cows. In comparison to cow milk, human milk showed a higher content of free fatty acids including polyunsaturated fatty acids, especially ω-3 fatty acids and their metabolites. Polyunsaturated fatty acids were enriched at an early period of lactation, while saturated fatty acids did not change significantly over the period. Moreover, human milk contained high levels of ω-3 fatty acid metabolites, particularly 18-hydroxyeicosapentaenoic acid, an eicosapentaenoic acid-derived metabolite with anti-inflammatory activity. In comparison with human normal milk, thromboxane B2 and protectin D1 levels were significantly elevated in milk from individuals with mastitis, suggesting that these lipid mediators could be potential biomarkers of obstructive mastitis. Overall, the unique lipid profile of human milk supports the efficacy of breast-feeding for supply of more nutritional and bioactive lipids in comparison to artificial or cow milk to infants, in whom digestive and absorptive functions are still immature.
Assuntos
Ácidos Graxos Ômega-3 , Mastite , Animais , Biomarcadores/metabolismo , Bovinos , Eicosanoides/metabolismo , Ácido Eicosapentaenoico , Ácidos Graxos/metabolismo , Ácidos Graxos não Esterificados/metabolismo , Ácidos Graxos Ômega-3/metabolismo , Ácidos Graxos Insaturados/metabolismo , Feminino , Humanos , Lactente , Lactação/metabolismo , Mastite/metabolismo , Leite/metabolismo , Leite Humano/metabolismo , Tromboxanos/metabolismoRESUMO
Aortic dissection is a life-threatening aortopathy involving separation of the aortic wall, whose underlying mechanisms are still incompletely understood. Epidemiological evidence suggests that unsaturated fatty acids improve cardiovascular health. Here, using quantitative RT-PCR, histological analyses, magnetic cell sorting and flow cytometry assays, and MS-based lipidomics, we show that the activity of a lipid-metabolizing enzyme, secreted phospholipase A2 group V (sPLA2-V), protects against aortic dissection by endogenously mobilizing vasoprotective lipids. Global and endothelial cell-specific sPLA2-V-deficient mice frequently developed aortic dissection shortly after infusion of angiotensin II (AT-II). We observed that in the AT-II-treated aorta, endothelial sPLA2-V mobilized oleic and linoleic acids, which attenuated endoplasmic reticulum stress, increased the expression of lysyl oxidase, and thereby stabilized the extracellular matrix in the aorta. Of note, dietary supplementation with oleic or linoleic acid reversed the increased susceptibility of sPLA2-V-deficient mice to aortic dissection. These findings reveal an unexplored functional link between sPLA2-driven phospholipid metabolism and aortic stability, possibly contributing to the development of improved diagnostic and/or therapeutic strategies for preventing aortic dissection.
Assuntos
Aorta/metabolismo , Dissecção Aórtica/metabolismo , Estresse do Retículo Endoplasmático , Fosfolipases A2 do Grupo V/metabolismo , Fosfolipídeos/metabolismo , Dissecção Aórtica/induzido quimicamente , Dissecção Aórtica/genética , Angiotensina II/efeitos adversos , Angiotensina II/farmacologia , Animais , Aorta/patologia , Modelos Animais de Doenças , Fosfolipases A2 do Grupo V/genética , Ácido Linoleico/genética , Ácido Linoleico/metabolismo , Camundongos , Camundongos Knockout , Ácido Oleico/genética , Ácido Oleico/metabolismo , Fosfolipídeos/genéticaRESUMO
During pregnancy, up-regulation of heparin-binding (HB-) EGF and cyclooxygenase-2 (COX-2) in the uterine epithelium contributes to decidualization, a series of uterine morphological changes required for placental formation and fetal development. Here, we report a key role for the lipid mediator lysophosphatidic acid (LPA) in decidualization, acting through its G-protein-coupled receptor LPA3 in the uterine epithelium. Knockout of Lpar3 or inhibition of the LPA-producing enzyme autotaxin (ATX) in pregnant mice leads to HB-EGF and COX-2 down-regulation near embryos and attenuates decidual reactions. Conversely, selective pharmacological activation of LPA3 induces decidualization via up-regulation of HB-EGF and COX-2. ATX and its substrate lysophosphatidylcholine can be detected in the uterine epithelium and in pre-implantation-stage embryos, respectively. Our results indicate that ATX-LPA-LPA3 signaling at the embryo-epithelial boundary induces decidualization via the canonical HB-EGF and COX-2 pathways.
Assuntos
Decídua/crescimento & desenvolvimento , Embrião de Mamíferos/fisiologia , Lisofosfolipídeos/metabolismo , Diester Fosfórico Hidrolases/metabolismo , Receptores de Ácidos Lisofosfatídicos/metabolismo , Transdução de Sinais , Útero/fisiologia , Animais , Ciclo-Oxigenase 2/metabolismo , Desenvolvimento Embrionário , Feminino , Técnicas de Inativação de Genes , Fator de Crescimento Semelhante a EGF de Ligação à Heparina/metabolismo , Camundongos , Camundongos Knockout , Receptores de Ácidos Lisofosfatídicos/deficiênciaRESUMO
Phospholipase A2 enzymes have long been implicated in the promotion of inflammation by mobilizing pro-inflammatory lipid mediators, yet recent evidence suggests that they also contribute to anti-inflammatory or pro-resolving programs. Group IID-secreted phospholipase A2 (sPLA2-IID) is abundantly expressed in dendritic cells in lymphoid tissues and resolves the Th1 immune response by controlling the steady-state levels of anti-inflammatory lipids such as docosahexaenoic acid and its metabolites. Here, we show that psoriasis and contact dermatitis were exacerbated in Pla2g2d-null mice, whereas they were ameliorated in Pla2g2d-overexpressing transgenic mice, relative to littermate wild-type mice. These phenotypes were associated with concomitant alterations in the tissue levels of ω3 polyunsaturated fatty acid (PUFA) metabolites, which had the capacity to reduce the expression of pro-inflammatory and Th1/Th17-type cytokines in dendritic cells or lymph node cells. In the context of cancer, however, Pla2g2d deficiency resulted in marked attenuation of skin carcinogenesis, likely because of the augmented anti-tumor immunity. Altogether, these results underscore a general role of sPLA2-IID as an immunosuppressive sPLA2 that allows the microenvironmental lipid balance toward an anti-inflammatory state, exerting beneficial or detrimental impact depending upon distinct pathophysiological contexts in inflammation and cancer.
Assuntos
Fosfolipases A2 do Grupo II/imunologia , Imunidade Celular , Proteínas de Neoplasias/imunologia , Neoplasias Cutâneas/imunologia , Células Th1/imunologia , Células Th17/imunologia , Animais , Ácidos Graxos Ômega-3/genética , Ácidos Graxos Ômega-3/imunologia , Fosfolipases A2 do Grupo II/genética , Inflamação/genética , Inflamação/imunologia , Inflamação/patologia , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/genética , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Células Th1/patologia , Células Th17/patologiaRESUMO
Recent studies using knock-out mice for various secreted phospholipase A2 (sPLA2) isoforms have revealed their non-redundant roles in diverse biological events. In the skin, group IIF sPLA2 (sPLA2-IIF), an "epidermal sPLA2" expressed in the suprabasal keratinocytes, plays a fundamental role in epidermal-hyperplasic diseases such as psoriasis and skin cancer. In this study, we found that group IIE sPLA2 (sPLA2-IIE) was expressed abundantly in hair follicles and to a lesser extent in basal epidermal keratinocytes in mouse skin. Mice lacking sPLA2-IIE exhibited skin abnormalities distinct from those in mice lacking sPLA2-IIF, with perturbation of hair follicle ultrastructure, modest changes in the steady-state expression of a subset of skin genes, and no changes in the features of psoriasis or contact dermatitis. Lipidomics analysis revealed that sPLA2-IIE and -IIF were coupled with distinct lipid pathways in the skin. Overall, two skin sPLA2s, hair follicular sPLA2-IIE and epidermal sPLA2-IIF, play non-redundant roles in distinct compartments of mouse skin, underscoring the functional diversity of multiple sPLA2s in the coordinated regulation of skin homeostasis and diseases.
Assuntos
Dermatite de Contato/enzimologia , Regulação Enzimológica da Expressão Gênica , Fosfolipases A2 do Grupo II/biossíntese , Folículo Piloso/enzimologia , Psoríase/enzimologia , Animais , Dermatite de Contato/genética , Dermatite de Contato/patologia , Epiderme/enzimologia , Epiderme/patologia , Fosfolipases A2 do Grupo II/genética , Folículo Piloso/patologia , Camundongos , Camundongos Knockout , Psoríase/genética , Psoríase/patologiaRESUMO
Within the secreted phospholipase A2(sPLA2) family, group X sPLA2(sPLA2-X) has the highest capacity to hydrolyze cellular membranes and has long been thought to promote inflammation by releasing arachidonic acid, a precursor of pro-inflammatory eicosanoids. Unexpectedly, we found that transgenic mice globally overexpressing human sPLA2-X (PLA2G10-Tg) displayed striking immunosuppressive and lean phenotypes with lymphopenia and increased M2-like macrophages, accompanied by marked elevation of free ω3 polyunsaturated fatty acids (PUFAs) and their metabolites. Studies usingPla2g10-deficient mice revealed that endogenous sPLA2-X, which is highly expressed in the colon epithelium and spermatozoa, mobilized ω3 PUFAs or their metabolites to protect against dextran sulfate-induced colitis and to promote fertilization, respectively. In colitis, sPLA2-X deficiency increased colorectal expression of Th17 cytokines, and ω3 PUFAs attenuated their production by lamina propria cells partly through the fatty acid receptor GPR120. In comparison, cytosolic phospholipase A2(cPLA2α) protects from colitis by mobilizing ω6 arachidonic acid metabolites, including prostaglandin E2 Thus, our results underscore a previously unrecognized role of sPLA2-X as an ω3 PUFA mobilizerin vivo, segregated mobilization of ω3 and ω6 PUFA metabolites by sPLA2-X and cPLA2α, respectively, in protection against colitis, and the novel role of a particular sPLA2-X-driven PUFA in fertilization.
Assuntos
Colite/genética , Colo/enzimologia , Ácidos Graxos Ômega-3/biossíntese , Fertilidade/genética , Fosfolipases A2 do Grupo X/genética , Espermatozoides/enzimologia , Animais , Ácido Araquidônico/antagonistas & inibidores , Ácido Araquidônico/biossíntese , Colite/induzido quimicamente , Colite/enzimologia , Colite/terapia , Colo/patologia , Sulfato de Dextrana , Ácidos Graxos Ômega-3/metabolismo , Ácidos Graxos Ômega-6/biossíntese , Ácidos Graxos Ômega-6/metabolismo , Expressão Gênica , Perfilação da Expressão Gênica , Fosfolipases A2 do Grupo X/metabolismo , Humanos , Interleucina-17/biossíntese , Masculino , Camundongos , Camundongos Transgênicos , Fosfolipases A2/genética , Fosfolipases A2/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Contagem de Espermatozoides , Motilidade dos Espermatozoides , Espermatozoides/patologia , Células Th17/metabolismo , Células Th17/patologia , TransgenesRESUMO
RATIONALE: Janus kinase/signal transducer and activator of transcription (JAK/STAT) signals and their endogenous inhibitor, suppressor of cytokine signaling 3 (SOCS3), in vascular endothelial cells (ECs) reportedly dominate the pathological angiogenesis. However, how these inflammatory signals are potentiated during pathological angiogenesis has not been fully elucidated. We suspected that an intracellular protease calpain, which composes the multifunctional proteolytic systems together with its endogenous inhibitor calpastatin (CAST), contributes to the JAK/STAT regulations. OBJECTIVE: To specify the effect of EC calpain/CAST systems on JAK/STAT signals and their relationship with pathological angiogenesis. METHODS AND RESULTS: The loss of CAST, which is ensured by several growth factor classes, was detectable in neovessels in murine allograft tumors, some human malignant tissues, and oxygen-induced retinopathy lesions in mice. EC-specific transgenic introduction of CAST caused downregulation of JAK/STAT signals, upregulation of SOCS3 expression, and depletion of vascular endothelial growth factor (VEGF)-C, thereby counteracting unstable pathological neovessels and disease progression in tumors and oxygen-induced retinopathy lesions in mice. Neutralizing antibody against VEGF-C ameliorated pathological angiogenesis in oxygen-induced retinopathy lesions. Small interfering RNA-based silencing of endogenous CAST in cultured ECs facilitated µ-calpain-induced proteolytic degradation of SOCS3, leading to VEGF-C production through amplified interleukin-6-driven STAT3 signals. Interleukin-6-induced angiogenic tube formation in cultured ECs was accelerated by CAST silencing, which is suppressible by pharmacological inhibition of JAK/STAT signals, antibody-based blockage of VEGF-C, and transfection of calpain-resistant SOCS3, whereas transfection of wild-type SOCS3 exhibited modest angiostatic effects. CONCLUSIONS: Loss of CAST in angiogenic ECs facilitates µ-calpain-induced SOCS3 degradation, which amplifies pathological angiogenesis through interleukin-6/STAT3/VEGF-C axis.
Assuntos
Proteínas de Ligação ao Cálcio/fisiologia , Calpaína/metabolismo , Células Endoteliais/metabolismo , Neoplasias/irrigação sanguínea , Proteínas Supressoras da Sinalização de Citocina/antagonistas & inibidores , Adenocarcinoma/irrigação sanguínea , Sequência de Aminoácidos , Animais , Aorta , Proteínas de Ligação ao Cálcio/genética , Carcinoma Pulmonar de Lewis/irrigação sanguínea , Células Cultivadas , Citocinas/fisiologia , Feminino , Glioblastoma/irrigação sanguínea , Humanos , Janus Quinases/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Neovascularização Patológica/fisiopatologia , Proteínas Recombinantes de Fusão/metabolismo , Retinopatia da Prematuridade/fisiopatologia , Fatores de Transcrição STAT/fisiologia , Transdução de Sinais/fisiologia , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina/genética , Proteínas Supressoras da Sinalização de Citocina/fisiologia , Fator C de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator C de Crescimento do Endotélio Vascular/fisiologiaRESUMO
PGE2 has long been known as a potentiator of acute inflammation, but its mechanisms of action still remain to be defined. In this study, we employed inflammatory swelling induced in mice by arachidonate and PGE2 as models and dissected the role and mechanisms of action of each EP receptor at the molecular level. Arachidonate- or PGE2-induced vascular permeability was significantly reduced in EP3-deficient mice. Intriguingly, the PGE2-induced response was suppressed by histamine H1 antagonist treatment, histidine decarboxylase deficiency, and mast cell deficiency. The impaired PGE2-induced response in mast cell-deficient mice was rescued upon reconstitution with wild-type mast cells but not with EP3-deficient mast cells. Although the number of mast cells, protease activity, and histamine contents in ear tissues in EP3-deficient mice were comparable to those in wild-type mice, the histamine contents in ear tissues were attenuated upon PGE2 treatment in wild-type but not in EP3-deficient mice. Consistently, PGE2-EP3 signaling elicited histamine release in mouse peritoneal and bone marrow-derived mast cells, and it exerted degranulation and IL-6 production in a manner sensitive to pertussis toxin and a PI3K inhibitor and dependent on extracellular Ca(2+) ions. These results demonstrate that PGE2 triggers mast cell activation via an EP3-Gi/o-Ca(2+) influx/PI3K pathway, and this mechanism underlies PGE2-induced vascular permeability and consequent edema formation.
Assuntos
Dinoprostona/fisiologia , Edema/fisiopatologia , Inflamação/fisiopatologia , Mastócitos/fisiologia , Receptores de Prostaglandina E Subtipo EP3/fisiologia , Animais , Ácido Araquidônico/toxicidade , Cálcio/fisiologia , Permeabilidade Capilar/efeitos dos fármacos , Permeabilidade Capilar/fisiologia , Degranulação Celular , Edema/etiologia , Liberação de Histamina/efeitos dos fármacos , Inflamação/complicações , Interleucina-6/fisiologia , Mastócitos/efeitos dos fármacos , Mastócitos/enzimologia , Camundongos , Camundongos Endogâmicos C57BL , Ativação de Neutrófilo , Peroxidase/análise , Fosfatidilinositol 3-Quinases/fisiologia , Inibidores de Fosfoinositídeo-3 Quinase , Receptores de Prostaglandina E Subtipo EP3/deficiência , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Organismos Livres de Patógenos EspecíficosRESUMO
Among more than 30 members of the phospholipase A2 (PLA2) superfamily, secreted PLA2 (sPLA2) enzymes represent the largest family, being Ca(2+)-dependent low-molecular-weight enzymes with a His-Asp catalytic dyad. Individual sPLA2s exhibit unique tissue and cellular distributions and enzymatic properties, suggesting their distinct biological roles. Recent studies using transgenic and knockout mice for nearly a full set of sPLA2 subtypes, in combination with sophisticated lipidomics as well as biochemical and cell biological studies, have revealed distinct contributions of individual sPLA2s to various pathophysiological events, including production of pro- and anti-inflammatory lipid mediators, regulation of membrane remodeling, degradation of foreign phospholipids in microbes or food, or modification of extracellular noncellular lipid components. In this review, we highlight the current understanding of the in vivo functions of sPLA2s and the underlying lipid pathways as revealed by a series of studies over the last decade.
Assuntos
Fosfolipases A2 Secretórias , Animais , Humanos , Fosfolipases A2 Secretórias/química , Fosfolipases A2 Secretórias/classificação , Fosfolipases A2 Secretórias/metabolismo , Receptores da Fosfolipase A2/metabolismoRESUMO
BACKGROUND: Extracellular signal-regulated kinases 1/2 (ERK1/2) make important contributions to allergic responses via their regulation of degranulation, eicosanoid production, and cytokine expression by mast cells, yet the mechanisms underlying their positive effects on FcεRI-dependent signaling are not fully understood. Recently, we reported that mast cell activation and anaphylaxis are negatively regulated by AMP-activated protein kinase (AMPK). However, little is known about the relationship between ERK1/2-mediated positive and the AMPK-mediated negative regulation of FcεRI signaling in mast cells. OBJECTIVE: We investigated possible interactions between ERK1/2 and AMPK in the modulation of mast cell signaling and anaphylaxis. METHODS: Wild-type or AMPKα2(-/-) mice, or bone marrow-derived mast cells obtained from these mice, were treated with either chemical agents or small interfering RNAs that modulated the activity or expression of ERK1/2 or AMPK to evaluate the functional interplay between ERK1/2 and AMPK in FcεRI-dependent signaling. RESULTS: The ERK1/2 pathway inhibitor U0126 and the AMPK activator 5-aminoimidazole-4-carboxamide-1-ß-4-ribofuranoside similarly inhibited FcεRI-mediated mast cell signals in vitro and anaphylaxis in vivo. ERK1/2-specific small interfering RNA also mimicked this effect on FcεRI signals. Moreover, AMPKα2 knockdown or deficiency led to increased FcεRI-mediated mast cell activation and anaphylaxis that were insensitive to U0126 or activator 5-aminoimidazole-4-carboxamide-1-ß-4-ribofuranoside, suggesting that the suppression of FcεRI signals by the inhibition of the ERK1/2 pathway relies largely on AMPK activation. ERK1/2 controlled AMPK activity by regulating its subcellular translocation. CONCLUSIONS: ERK1/2 ablated the AMPK-dependent negative regulatory axis, thereby activating FcεRI signals in mast cells.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Anafilaxia/imunologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Hipersensibilidade/imunologia , Mastócitos/imunologia , Proteínas Quinases Ativadas por AMP/antagonistas & inibidores , Proteínas Quinases Ativadas por AMP/genética , Aminoimidazol Carboxamida/análogos & derivados , Aminoimidazol Carboxamida/farmacologia , Anafilaxia/etiologia , Animais , Butadienos/farmacologia , Degranulação Celular/efeitos dos fármacos , Células Cultivadas , MAP Quinases Reguladas por Sinal Extracelular/antagonistas & inibidores , Hipersensibilidade/complicações , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Nitrilas/farmacologia , Receptores de IgG/metabolismo , Ribonucleosídeos/farmacologia , Transdução de Sinais/efeitos dos fármacosRESUMO
Phospholipase A2s (PLA2s) are a group of enzymes that hydrolyze the sn-2 position of phospholipids to release (typically unsaturated) fatty acids and lysophospholipids, which serve as precursors for a variety of bioactive lipid mediators. Among the PLA2 superfamily, secreted PLA2 (sPLA2) enzymes comprise the largest subfamily that includes 11 isoforms with a conserved His-Asp catalytic dyad. Individual sPLA2 enzymes exhibit unique tissue and cellular localizations and specific enzymatic properties, suggesting their distinct biological roles. Recent studies using transgenic and knockout mice for individual sPLA2 isofoms have revealed their involvement in various pathophysiological events. Here, we overview the current state of knowledge about sPLA2s, specifically their roles in mast cells (MCs) in the context of allergology. In particular, we highlight group III sPLA2 (PLA2G3) as an "anaphylactic sPLA2" that promotes MC maturation and thereby anaphylaxis through a previously unrecognized lipid-orchestrated circuit.
Assuntos
Mastócitos/imunologia , Mastócitos/metabolismo , Fosfolipases A2 Secretórias/metabolismo , Animais , Diferenciação Celular , Eicosanoides/biossíntese , Fosfolipases A2 do Grupo III/metabolismo , Humanos , Mastócitos/citologia , Fosfolipases A2 Citosólicas/metabolismoRESUMO
BACKGROUND: Aggregation of FcεRI activates a cascade of signaling events leading to mast cell activation, followed by inhibitory signals that turn off the activating signals. However, the overall view of negative signals in mast cells is still incomplete. Although AMP-activated protein kinase (AMPK), which is generally known as a regulator of energy metabolism, is also associated with anti-inflammation, little is known about the role of AMPK in mast cells. OBJECTIVES: We investigated the role of AMPK and its regulatory mechanism in mast cells. METHOD: The roles of AMPK in FcεRI-dependent activation of bone marrow-derived mast cells (BMMCs) were evaluated by using chemical agents, small interfering RNAs (siRNAs), or adenovirus that modulated the activity or expression of AMPK signaling components. In addition, AMPKα2(-/-) mice were used to verify the role of AMPK in anaphylactic models. RESULTS: FcεRI signaling and associated effector functions in BMMCs were suppressed by the AMPK activator 5-aminoimidazole-4-carboxamide-1-ß-4-ribofuranoside (AICAR) and were conversely augmented by siRNA knockdown of AMPKα2 or liver kinase B1 (LKB1), an upstream kinase of AMPK. Furthermore, AMPKα2 deficiency led to increased FcεRI-mediated BMMC activation and anaphylaxis that were insensitive to AICAR, whereas enforced expression of AMPKα2 in AMPKα2(-/-) BMMCs reversed the hypersensitive FcεRI signaling to normal levels. Pharmacologic inhibition or siRNA knockdown of Fyn mimicked AMPK activation, suggesting that Fyn counterregulates the LKB1-AMPK axis. Mechanistically, Fyn controlled AMPK activity by regulating LKB1 localization. CONCLUSIONS: The Fyn-regulated LKB1-AMPK axis acts as a novel inhibitory module for mast cell activation, which points to AMPK activators as therapeutic drugs for allergic diseases.
Assuntos
Proteínas Quinases Ativadas por AMP/imunologia , Anafilaxia/imunologia , Mastócitos/imunologia , Receptores de IgE/imunologia , Proteínas Quinases Ativadas por AMP/genética , Animais , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/imunologia , Proteínas Proto-Oncogênicas c-fyn/genética , Proteínas Proto-Oncogênicas c-fyn/imunologiaRESUMO
Current treatments of eosinophilic chronic rhinosinusitis (ECRS) involve corticosteroids with various adverse effects and costly therapies such as dupilumab, highlighting the need for improved treatments. However, because of the lack of a proper mouse ECRS model that recapitulates human ECRS, molecular mechanisms underlying this disease are incompletely understood. ECRS is often associated with aspirin-induced asthma, suggesting that dysregulation of lipid mediators in the nasal mucosa may underlie ECRS pathology. We herein found that the expression of microsomal PGE synthase-1 (encoded by PTGES) was significantly lower in the nasal mucosa of ECRS patients than that of non-ECRS subjects. Histological, transcriptional, and lipidomics analyses of Ptges-deficient mice revealed that defective PGE2 biosynthesis facilitated eosinophil recruitment into the nasal mucosa, elevated expression of type-2 cytokines and chemokines, and increased pro-allergic and decreased anti-allergic lipid mediators following challenges with Aspergillus protease and ovalbumin. A nasal spray containing agonists for the PGE2 receptor EP2 or EP4, including omidenepag isopropyl that has been clinically used for treatment of glaucoma, markedly reduced intranasal eosinophil infiltration in Ptges-deficient mice. These results suggest that the present model using Ptges-deficient mice is more relevant to human ECRS than are previously reported models and that eosinophilic inflammation in the nasal mucosa can be efficiently blocked by activation of the PGE2-EP2 pathway. Furthermore, our findings suggest that drug repositioning of omidenepag isopropyl may be useful for treatment of patients with ECRS.
Assuntos
Dinoprostona , Eosinofilia , Camundongos Knockout , Mucosa Nasal , Receptores de Prostaglandina E Subtipo EP2 , Rinite , Sinusite , Animais , Sinusite/tratamento farmacológico , Sinusite/metabolismo , Sinusite/imunologia , Humanos , Camundongos , Rinite/tratamento farmacológico , Rinite/metabolismo , Rinite/imunologia , Dinoprostona/metabolismo , Mucosa Nasal/metabolismo , Mucosa Nasal/imunologia , Mucosa Nasal/efeitos dos fármacos , Eosinofilia/tratamento farmacológico , Eosinofilia/metabolismo , Receptores de Prostaglandina E Subtipo EP2/metabolismo , Modelos Animais de Doenças , Masculino , Transdução de Sinais/efeitos dos fármacos , Prostaglandina-E Sintases/genética , Prostaglandina-E Sintases/metabolismo , Eosinófilos/imunologia , Eosinófilos/metabolismo , Eosinófilos/efeitos dos fármacos , Feminino , Doença Crônica , Camundongos Endogâmicos C57BL , RinossinusiteRESUMO
[This corrects the article DOI: 10.3389/fimmu.2023.1251784.].