RESUMEN
Polysaccharide fractions from the mycelium of the lion's mane medicinal mushroom Hericium erinaceus BP 16, cultivated on sterile grain substrates (barley, oats, wheat, rice, rye), were isolated and characterized. One percent solutions were prepared from the resulting fractions, mixed with blood, which was then subjected to cold stress at a temperature of 6°C for 3, 5 and 7 d. It has been shown that the fraction of H. erinaceus grown on rye is characterized by a high content of the protein fraction and arabinose monosaccharide and contributes to the preservation of higher phagocytic, bactericidal and antioxidant activity cells throughout the entire period of stress. Polysaccharide fractions of the fungus H. erinaceus, grown on various grain substrates, can serve as an immunomodulatory and antioxidant food additive and provide significant benefits in the daily life of people with stress and reduced immunity.
Asunto(s)
Antioxidantes , Hericium , Neutrófilos , Antioxidantes/farmacología , Hericium/química , Neutrófilos/efectos de los fármacos , Neutrófilos/inmunología , Neutrófilos/metabolismo , Factores Inmunológicos/farmacología , Polisacáridos/farmacología , Polisacáridos/química , Polisacáridos/aislamiento & purificación , Micelio/química , Humanos , Estrés Fisiológico/efectos de los fármacos , Agentes Inmunomoduladores/farmacología , Agentes Inmunomoduladores/química , Agentes Inmunomoduladores/aislamiento & purificación , Polisacáridos Fúngicos/farmacología , Polisacáridos Fúngicos/químicaRESUMEN
Neutrophil extracellular traps (NETs) are three-dimensional reticular structures that release chromatin and cellular contents extracellularly upon neutrophil activation. As a novel effector mechanism of neutrophils, NETs possess the capacity to amplify localized inflammation and have been demonstrated to contribute to the exacerbation of various inflammatory diseases, including cardiovascular diseases and tumors. It is suggested that lysophosphatidylcholine (LPC), as the primary active component of oxidized low-density lipoprotein, represents a significant risk factor for various inflammatory diseases, such as cardiovascular diseases and neurodegenerative diseases. However, the specific mechanism of NETs formation induced by LPC remains unclear. Quercetin has garnered considerable attention due to its anti-inflammatory properties, serving as a prevalent flavonoid in daily diet. However, little is currently known about the underlying mechanisms by which quercetin inhibits NETs formation and alleviates associated diseases. In our study, we utilized LPC-treated primary rat neutrophils to establish an in vitro model of NETs formation, which was subsequently subjected to treatment with a combination of quercetin or relevant inhibitors/activators. Compared to the control group, the markers of NETs and the expression of P2X7R/P38MAPK/NOX2 pathway-associated proteins were significantly increased in cells treated with LPC alone. Quercetin intervention decreased the LPC-induced upregulation of the P2X7R/P38MAPK/NOX2 pathway and effectively reduced the expression of NETs markers. The results obtained using a P2X7R antagonist/activator and P38MAPK inhibitor/activator support these findings. In summary, quercetin reversed the upregulation of the LPC-induced P2X7R/P38MAPK/NOX2 pathway, further mitigating NETs formation. Our study investigated the potential mechanism of LPC-induced NETs formation, elucidated the inhibitory effect of quercetin on NETs formation, and offered new insights into the anti-inflammatory properties of quercetin.
Asunto(s)
Trampas Extracelulares , Lisofosfatidilcolinas , NADPH Oxidasa 2 , Neutrófilos , Quercetina , Receptores Purinérgicos P2X7 , Proteínas Quinasas p38 Activadas por Mitógenos , Quercetina/farmacología , Lisofosfatidilcolinas/metabolismo , Lisofosfatidilcolinas/farmacología , Trampas Extracelulares/metabolismo , Trampas Extracelulares/efectos de los fármacos , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Animales , Ratas , Neutrófilos/metabolismo , Neutrófilos/efectos de los fármacos , Receptores Purinérgicos P2X7/metabolismo , NADPH Oxidasa 2/metabolismo , Transducción de Señal/efectos de los fármacos , MasculinoRESUMEN
Inflammatory bowel disease (IBD) is a chronic non-specific intestinal inflammatory disease that affects millions of people worldwide, and current treatment methods have certain limitations. This study aimed to explore the therapeutic potential and mechanism of action of lemairamin (Wgx-50) in inflammatory bowel disease (IBD). We used dextran sulfate sodium (DSS)-treated zebrafish as an inflammatory bowel disease model, and observed the effect of Wgx-50 on DSS-induced colitis inflammation. The results of the study showed that Wgx-50 could reduce the expression of pro-inflammatory cytokines induced by DSS and inhibit the recruitment of neutrophils to the site of intestinal injury. Further experiments revealed that Wgx-50 exerted its anti-inflammatory effect by regulating the activation of the Akt pathway. These research findings indicate that Wgx-50 possesses anti-inflammatory activity.
Asunto(s)
Antiinflamatorios , Colitis , Sulfato de Dextran , Modelos Animales de Enfermedad , Pez Cebra , Animales , Sulfato de Dextran/efectos adversos , Antiinflamatorios/farmacología , Colitis/inducido químicamente , Colitis/tratamiento farmacológico , Colitis/metabolismo , Colitis/patología , Citocinas/metabolismo , Neutrófilos/efectos de los fármacos , Neutrófilos/metabolismo , Transducción de Señal/efectos de los fármacos , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Enfermedades Inflamatorias del Intestino/inducido químicamente , Enfermedades Inflamatorias del Intestino/patología , Enfermedades Inflamatorias del Intestino/metabolismo , Intestinos/efectos de los fármacos , Intestinos/patología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Inflamación/tratamiento farmacológico , Inflamación/patología , Inflamación/inducido químicamente , Inflamación/metabolismoRESUMEN
Factor XII (FXII) is the zymogen of the plasma protease FXIIa that activates the intrinsic coagulation pathway and the kallikrein kinin-system. The role of FXII in inflammation has been obscure. Here, we report a single-domain antibody (nanobody, Nb) fused to the Fc region of a human immunoglobulin (Nb-Fc) that recognizes FXII in a conformation-dependent manner and interferes with FXIIa formation. Nb-Fc treatment inhibited arterial thrombosis in male mice without affecting hemostasis. In a mouse model of extracorporeal membrane oxygenation (ECMO), FXII inhibition or knockout reduced thrombus deposition on oxygenator membranes and systemic microvascular thrombi. ECMO increased circulating levels of D-dimer, alkaline phosphatase, creatinine and TNF-α and triggered microvascular neutrophil adherence, platelet aggregation and their interaction, which were substantially attenuated by FXII blockade. Both Nb-Fc treatment and FXII knockout markedly ameliorated immune complex-induced local vasculitis and anti-neutrophil cytoplasmic antibody-induced systemic vasculitis, consistent with selectively suppressed neutrophil migration. In human blood microfluidic analysis, Nb-Fc treatment prevented collagen-induced fibrin deposition and neutrophil adhesion/activation. Thus, FXII is an important mediator of inflammatory responses in vasculitis and ECMO, and Nb-Fc provides a promising approach to alleviate thrombo-inflammatory disorders.
Asunto(s)
Factor XII , Inflamación , Ratones Noqueados , Neutrófilos , Anticuerpos de Dominio Único , Trombosis , Animales , Humanos , Trombosis/inmunología , Trombosis/metabolismo , Anticuerpos de Dominio Único/farmacología , Anticuerpos de Dominio Único/inmunología , Masculino , Factor XII/metabolismo , Factor XII/antagonistas & inhibidores , Inflamación/metabolismo , Ratones , Neutrófilos/inmunología , Neutrófilos/metabolismo , Neutrófilos/efectos de los fármacos , Ratones Endogámicos C57BL , Modelos Animales de Enfermedad , Agregación Plaquetaria/efectos de los fármacos , Factor XIIa/metabolismo , Factor XIIa/antagonistas & inhibidores , Fibrina/metabolismo , Productos de Degradación de Fibrina-Fibrinógeno/metabolismoRESUMEN
Recurring lung injury, chronic inflammation, aberrant tissue repair and impaired tissue remodelling contribute to the pathogenesis of pulmonary fibrosis (PF). Neutrophil extracellular traps (NETs) are released by activated neutrophils to trap, immobilise and kill invading pathogen and is facilitated by peptidyl arginine deiminase-4 (PAD-4). Dysregulated NETs release and abnormal PAD-4 activation plays a crucial role in activating pro-fibrotic events in PF. Developmental endothelial locus-1 (Del-1), expressed by the endothelial cells of lungs and brain acts as an endogenous inhibitor of inflammation and fibrosis. We have hypothesised that PAD-4 inhibitor exerts anti-inflammatory and anti-fibrotic effects in mice model of PF. We have also hypothesised by PAD-4 regulated the transcription of Del-1 through co-repression and its inhibition potentiates anti-fibrotic effects of Del-1. In our study, the PAD-4 inhibitor chloro-amidine (CLA) demonstrated anti-NETotic and anti-inflammatory effects in vitro in differentiated HL-60 cells. In a bleomycin-induced PF mice model, CLA administration in two doses (3 mg/kg, I.P and 10 mg/kg, I.P) improved lung function, normalized bronchoalveolar lavage fluid parameters, and attenuated fibrotic events, including markers of extracellular matrix and epithelial-mesenchymal transition. Histological analyses confirmed the restoration of lung architecture and collagen deposition with CLA treatment. ELISA, IHC, IF, RT-PCR, and immunoblot analysis supported the anti-NETotic effects of CLA. Furthermore, BLM-induced PF reduced Del-1 and p53 expression, which was normalized by CLA treatment. These findings suggest that inhibition of PAD-4 results in amelioration of PF in animal model and may involve modulation of Del-1 and p53 pathways, warranting further investigation.
Asunto(s)
Bleomicina , Arginina Deiminasa Proteína-Tipo 4 , Fibrosis Pulmonar , Animales , Fibrosis Pulmonar/tratamiento farmacológico , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/patología , Humanos , Arginina Deiminasa Proteína-Tipo 4/antagonistas & inhibidores , Arginina Deiminasa Proteína-Tipo 4/metabolismo , Ratones , Trampas Extracelulares/efectos de los fármacos , Trampas Extracelulares/metabolismo , Células HL-60 , Ratones Endogámicos C57BL , Modelos Animales de Enfermedad , Proteína p53 Supresora de Tumor/metabolismo , Proteína p53 Supresora de Tumor/genética , Pulmón/patología , Pulmón/efectos de los fármacos , Masculino , Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico , Neutrófilos/efectos de los fármacos , Neutrófilos/inmunología , Proteínas de Unión al Calcio/metabolismo , Proteínas de Unión al Calcio/genética , Ornitina/análogos & derivadosRESUMEN
A balanced immune system is essential to maintain adequate host defense and effective self-tolerance. While an immune system that fails to generate appropriate response will permit infections to develop, uncontrolled activation may lead to autoinflammatory or autoimmune diseases. To identify drug candidates capable of modulating immune cell functions, we screened 1200 small molecules from the Prestwick Chemical Library for their property to inhibit innate or adaptive immune responses. Our studies focused specifically on drug interactions with T cells, B cells, and polymorphonuclear leukocytes (PMNs). Candidate drugs that were validated in vitro were examined in preclinical models to determine their immunomodulatory impact in chronic inflammatory diseases, here investigated in chronic inflammatory skin diseases. Using this approach, we identified several candidate drugs that were highly effective in preclinical models of chronic inflammatory disease. For example, we found that administration of pyrvinium pamoate, an FDA-approved over-the-counter anthelmintic drug, suppressed B cell activation in vitro and halted the progression of B cell-dependent experimental pemphigoid by reducing numbers of autoantigen-specific B cell responses. In addition, in studies performed in gene-deleted mouse strains provided additional insight into the mechanisms underlying these effects, for example, the receptor-dependent actions of tamoxifen that inhibit immune-complex-mediated activation of PMNs. Collectively, our methods and findings provide a vast resource that can be used to identify drugs that may be repurposed and used to promote or inhibit cellular immune responses.
Asunto(s)
Inmunidad Adaptativa , Linfocitos B , Ensayos Analíticos de Alto Rendimiento , Inmunidad Innata , Animales , Ratones , Inmunidad Innata/efectos de los fármacos , Ensayos Analíticos de Alto Rendimiento/métodos , Humanos , Inmunidad Adaptativa/efectos de los fármacos , Linfocitos B/inmunología , Linfocitos B/efectos de los fármacos , Linfocitos B/metabolismo , Reposicionamiento de Medicamentos/métodos , Linfocitos T/inmunología , Linfocitos T/efectos de los fármacos , Linfocitos T/metabolismo , Activación de Linfocitos/efectos de los fármacos , Activación de Linfocitos/inmunología , Neutrófilos/inmunología , Neutrófilos/efectos de los fármacos , Neutrófilos/metabolismo , Modelos Animales de Enfermedad , Bibliotecas de Moléculas Pequeñas/farmacología , Evaluación Preclínica de Medicamentos , Ratones NoqueadosAsunto(s)
Antipsicóticos , Neutrófilos , Humanos , Antipsicóticos/uso terapéutico , Neutrófilos/efectos de los fármacos , Masculino , Adulto , Recuento de Leucocitos , Femenino , Sistema del Grupo Sanguíneo Duffy/genética , Esquizofrenia/tratamiento farmacológico , Esquizofrenia/sangre , Persona de Mediana EdadRESUMEN
Neutrophil extracellular trap formation has been identified as a new cell death mediator, termed NETosis, which is distinct from apoptosis and necrosis. NETs capture foreign substances, such as bacteria, by releasing DNA into the extracellular environment, and have been associated with inflammatory diseases and altered immune responses. Short-chain fatty acids, such as acetate, are produced by the gut microbiota and reportedly enhance innate immune responses; however, the underlying molecular mechanisms remain unclear. Here, we investigated the effects of sodium acetate, which has the highest SCFA concentration in the blood and gastrointestinal tract, on NETosis by focusing on the mechanisms associated with histone acetylation in neutrophil-like HL-60 cells. Sodium acetate enhanced NETosis, as shown by fluorescence staining with SYTOX green, and the effect was directly proportional to the treatment duration (16-24 h). Moreover, the addition of sodium acetate significantly enhanced the acetylation of Ace-H3, H3K9ace, and H3K14ace. Sodium acetate-induced histone acetylation rapidly decreased upon stimulation with the calcium ionophore A23187, whereas histone citrullination markedly increased. These results demonstrate that sodium acetate induces NETosis via histone acetylation in neutrophil-like HL-60 cells, providing new insights into the therapeutic effects based on the innate immunity-enhancing effect of dietary fiber.
Asunto(s)
Trampas Extracelulares , Histonas , Neutrófilos , Acetato de Sodio , Humanos , Células HL-60 , Trampas Extracelulares/metabolismo , Trampas Extracelulares/efectos de los fármacos , Histonas/metabolismo , Acetilación/efectos de los fármacos , Neutrófilos/metabolismo , Neutrófilos/efectos de los fármacos , Acetato de Sodio/farmacologíaRESUMEN
BACKGROUND: Acute lung injury (ALI) is a devastating condition caused by sepsis, pneumonia, trauma, and more recently, COVID-19. SH003, an herbal formula consisted of Astragalus membranaceus, Angelica gigas and Trichosanthes kirilowii, is known for its effects on cancer and immunoregulation. HYPOTHESIS/PURPOSE: Previous studies show SH003 exerts a promising anti-inflammatory effect. This study investigates the effect of modified SH003 on ALI using in silico, in vivo, and in vitro models. STUDY DESIGN AND METHODS: We performed in silico-based analysis of SH003 on ALI-related pathways. C57BL/6 mice were intraperitoneally subjected to lipopolysaccharide (LPS) to induce septic ALI, followed by oral administration of SH003 for 2 weeks. Dexamethasone was used as the positive control. Human peripheral blood-derived polymorphonuclear neutrophils (PMN) were used to investigate the effect and mechanisms of SH003 on neutrophil extracellular trap (NET) formation. RESULTS: Network pharmacology analysis suggested SH003 regulates lung inflammation by modulating NET formation. SH003 significantly reduced mortality in sepsis in vivo by inhibiting local and systemic inflammation, likely via nuclear factor kappa B and mitogen-activated protein kinase pathways-mediated inflammasome suppression. SH003 also decreased NET-related markers in lung tissues and inhibited LPS- and phorbol myristate acetate-induced NET formation in PMN. Cytometry time-of-flight analysis confirmed regulation of NETosis-related pathways by SH003. CONCLUSION: SH003 effectively inhibits excessive immune responses in the lung by suppressing inflammasome activation and NET formation. These findings suggest SH003 as a potential therapeutic agent for septic ALI.
Asunto(s)
Lesión Pulmonar Aguda , Angelica , Astragalus propinquus , Trampas Extracelulares , Inflamasomas , Lipopolisacáridos , Ratones Endogámicos C57BL , Neutrófilos , Animales , Lesión Pulmonar Aguda/tratamiento farmacológico , Lesión Pulmonar Aguda/inducido químicamente , Trampas Extracelulares/efectos de los fármacos , Ratones , Neutrófilos/efectos de los fármacos , Humanos , Inflamasomas/metabolismo , Inflamasomas/efectos de los fármacos , Astragalus propinquus/química , Masculino , Angelica/química , Medicamentos Herbarios Chinos/farmacología , Antiinflamatorios/farmacología , Modelos Animales de EnfermedadRESUMEN
Besnoitia besnoiti is an obligate intracellular apicomplexan parasite and the causal agent of bovine besnoitiosis. Bovine besnoitiosis has a considerable economic impact in Africa and Asia due to reduced milk production, abortions, and bull infertility. In Europe, bovine besnoitiosis is classified as an emerging disease. Polymorphonuclear neutrophils (PMN) are one of the most abundant leukocytes in cattle blood and amongst the first immunological responders toward invading pathogens. In the case of B. besnoiti, bovine PMN produce reactive oxygen species (ROS), release neutrophil extracellular traps (NETs), and show increased autophagic activities upon exposure to tachyzoite stages. In that context, the general processes of NETosis and autophagy were previously reported as associated with AMP-activated protein kinase (AMPK) activation. Here, we study the role of AMPK in B. besnoiti tachyzoite-induced NET formation, thereby expanding the analysis to both upstream proteins, such as the calcium/calmodulin-dependent protein kinase kinase 2 (CAMKK), and downstream signaling and effector molecules, such as the autophagy-related proteins ULK-1 and Beclin-1. Current data revealed early AMPK activation (<30 min) in both B. besnoiti-exposed and AMPK activator (AICAR)-treated bovine PMN. This finding correlated with upstream responses on the level of CAMKK activation. Moreover, these reactions were accompanied by an augmented autophagic activity, as represented by enhanced expression of ULK-1 but not of Beclin-1. Referring to neutrophil effector functions, AICAR treatments induced both AMPK phosphorylation and NET formation, without affecting cell viability. In B. besnoiti tachyzoite-exposed PMN, AICAR treatments failed to affect oxidative responses, but led to enhanced NET formation, thereby indicating that AMPK and autophagic activation synergize with B. besnoiti-driven NETosis.
Asunto(s)
Proteínas Quinasas Activadas por AMP , Quinasa de la Proteína Quinasa Dependiente de Calcio-Calmodulina , Trampas Extracelulares , Neutrófilos , Sarcocystidae , Transducción de Señal , Animales , Bovinos , Neutrófilos/metabolismo , Neutrófilos/efectos de los fármacos , Neutrófilos/inmunología , Proteínas Quinasas Activadas por AMP/metabolismo , Quinasa de la Proteína Quinasa Dependiente de Calcio-Calmodulina/metabolismo , Trampas Extracelulares/metabolismo , Sarcocystidae/metabolismo , Transducción de Señal/efectos de los fármacos , Autofagia/efectos de los fármacos , Coccidiosis/parasitología , Coccidiosis/veterinaria , Coccidiosis/inmunología , Enfermedades de los Bovinos/parasitología , Enfermedades de los Bovinos/metabolismo , Enfermedades de los Bovinos/inmunología , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Interaction of microbiota with hybrid vaterite-pectin microparticles as an attractive multifunctional vehicle for mucosal delivery should not provoke inflammation. Our purpose was to study the reaction of bacteria E. coli strain Mg1655 and isolate SharL from a patient with Crohn disease on the cultivation with hybrid microparticles and vaterite, and the subsequent activation of neutrophils. Vaterite-pectin microparticles enhanced leakage of ATP from bacteria. For E. coli Mg1655, the concentration of DNA decreased, while intracellular ATP increased. For E. coli SharL, the intracellular ATP decreased with simultaneous growth of DNA. Bacteria and microparticles together did not enhance activation of neutrophils in comparison with the particles per se in the medium without serum and in comparison with bacteria in the medium supplemented with serum; microparticles did not reduce functional activity of neutrophils.
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Escherichia coli , Neutrófilos , Pectinas , Humanos , Neutrófilos/efectos de los fármacos , Neutrófilos/metabolismo , Escherichia coli/efectos de los fármacos , Pectinas/farmacología , Adenosina Trifosfato/metabolismo , Carbonato de Calcio/farmacología , Carbonato de Calcio/química , Enfermedad de Crohn/microbiología , Enfermedad de Crohn/patología , Activación Neutrófila/efectos de los fármacosRESUMEN
In an experimental model of Alzheimer's disease in mice, oral administration of trehalose disaccharide reduces neuroinflammation assessed by the expression level of microglia activation marker Iba1 and affects the neutrophil degranulation activity. A potential anti-inflammatory effect of 4% trehalose solution associated with a decrease in the activity of leukocyte elastase in plasma was revealed.
Asunto(s)
Enfermedad de Alzheimer , Modelos Animales de Enfermedad , Microglía , Trehalosa , Animales , Trehalosa/farmacología , Trehalosa/uso terapéutico , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Microglía/efectos de los fármacos , Microglía/metabolismo , Ratones , Proteínas de Microfilamentos/metabolismo , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Masculino , Proteínas de Unión al Calcio/metabolismo , Elastasa de Leucocito/metabolismo , Neutrófilos/efectos de los fármacos , Neutrófilos/metabolismo , Disacáridos/farmacología , Antiinflamatorios/farmacologíaRESUMEN
Stroke is a leading cause of permanent disability worldwide. Despite intensive research over the last decades, key anti-inflammatory strategies that have proven beneficial in pre-clinical animal models have often failed in translation. The importance of neutrophils as pro- and anti-inflammatory peripheral immune cells has often been overlooked in ischemic stroke. However, neutrophils rapidly infiltrate into the brain parenchyma after stroke and secrete an array of pro-inflammatory factors including reactive oxygen species, proteases, cytokines, and chemokines exacerbating damage. In this study, we demonstrate the neuroprotective and anti-inflammatory effect of benserazide, a clinically used DOPA decarboxylase inhibitor, using both in vitro models of inflammation and in vivo mouse models of focal cerebral ischemia. Benserazide significantly attenuated PMA-induced NETosis in isolated human neutrophils. Furthermore, benserazide was able to protect both SH-SY5Y and iPSC-derived human cortical neurons when challenged with activated neutrophils demonstrating the clinical relevance of this study. Additional in vitro data suggest the ability of benserazide to polarize macrophages towards M2-phenotypes following LPS stimulation. Neuroprotective effects of benserazide are further demonstrated by in vivo studies where peripheral administration of benserazide significantly attenuated neutrophil infiltration into the brain, altered microglia/macrophage phenotypes, and improved the behavioral outcome post-stroke. Overall, our data suggest that benserazide could serve as a drug candidate for the treatment of ischemic stroke. The importance of our results for future clinical trials is further underlined as benserazide has been approved by the European Medicines Agency as a safe and effective treatment in Parkinson's disease when combined with levodopa.
Asunto(s)
Benserazida , Accidente Cerebrovascular Isquémico , Fármacos Neuroprotectores , Neutrófilos , Benserazida/farmacología , Animales , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Humanos , Accidente Cerebrovascular Isquémico/tratamiento farmacológico , Accidente Cerebrovascular Isquémico/inmunología , Accidente Cerebrovascular Isquémico/metabolismo , Ratones , Neutrófilos/efectos de los fármacos , Neutrófilos/inmunología , Neutrófilos/metabolismo , Modelos Animales de Enfermedad , Recuperación de la Función/efectos de los fármacos , Masculino , Ratones Endogámicos C57BL , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/metabolismo , Microglía/efectos de los fármacos , Microglía/metabolismo , Neuronas/efectos de los fármacos , Neuronas/metabolismoRESUMEN
Severe alcohol-associated hepatitis (AH) is a life-threatening form of alcohol-associated liver disease. Liver neutrophil infiltration is a hallmark of AH, yet the effects of alcohol on neutrophil functions remain elusive. Identifying therapeutic targets to reduce neutrophil-mediated liver damage is essential. Bruton's tyrosine kinase (BTK) plays an important role in neutrophil development and function; however, the role of BTK in AH is unknown. Using RNA sequencing of circulating neutrophils, we found an increase in Btk expression (P = 0.05) and phosphorylated BTK (pBTK) in patients with AH compared with healthy controls. In vitro, physiologically relevant doses of alcohol resulted in a rapid, TLR4-mediated induction of pBTK in neutrophils. In a preclinical model of AH, administration of a small-molecule BTK inhibitor (evobrutinib) or myeloid-specific Btk knockout decreased proinflammatory cytokines and attenuated neutrophil-mediated liver damage. We found that pBTK was essential for alcohol-induced bone marrow granulopoiesis and liver neutrophil infiltration. In vivo, BTK inhibition or myeloid-specific Btk knockout reduced granulopoiesis, circulating neutrophils, liver neutrophil infiltration, and liver damage in a mouse model of AH. Mechanistically, using liquid chromatography-tandem mass spectrometry, we identified CD84 as a kinase target of BTK, which is involved in granulopoiesis. In vitro, CD84 promoted alcohol-induced interleukin-1ß and tumor necrosis factor-α in primary human neutrophils, which was inhibited by CD84-blocking antibody treatment. Our findings define the role of BTK and CD84 in regulating neutrophil inflammation and granulopoiesis, with potential therapeutic implications in AH.
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Agammaglobulinemia Tirosina Quinasa , Hepatopatías Alcohólicas , Neutrófilos , Agammaglobulinemia Tirosina Quinasa/metabolismo , Agammaglobulinemia Tirosina Quinasa/antagonistas & inhibidores , Animales , Humanos , Neutrófilos/metabolismo , Neutrófilos/efectos de los fármacos , Hepatopatías Alcohólicas/metabolismo , Hepatopatías Alcohólicas/patología , Inhibidores de Proteínas Quinasas/farmacología , Ratones , Masculino , Hígado/patología , Hígado/metabolismo , Hígado/efectos de los fármacos , Granulocitos/metabolismo , Granulocitos/efectos de los fármacos , Ratones Endogámicos C57BL , Antígenos CD/metabolismo , Ratones Noqueados , Receptor Toll-Like 4/metabolismo , Fosforilación/efectos de los fármacosRESUMEN
The present study focused on the efficacy and role of triptolide (TPL) in relieving symptoms of acute gouty arthritis (AGA) in vivo and in vitro. The effects of TPL in AGA were investigated in monosodium urate (MSU)-treated rat ankles, RAW264.7 macrophages, and neutrophils isolated from mouse peritoneal cavity. Observation of pathological changes in the ankle joint of rats. Enzyme-linked immunosorbent assay and real-time quantitative polymerase chain reaction (RT-qPCR) were performed to detect the expression levels of inflammatory factors and chemokines. The levels of the indicators of macrophage M1/M2 polarization, and the mechanistic targets of Akt and rapamycin complex 2, were determined via western blotting and RT-qPCR. The expression levels of CD86 and CD206 were detected using immunohistochemistry. Neutrophil migration was observed via air pouch experiments in vivo and Transwell cell migration assay in vitro. Myeloperoxidase (MPO) and Neutrophil elastase (NE) release was analyzed by via immunohistochemistry and immunofluorescence. The expression levels of beclin-1, LC3B, Bax, Bcl-2, and cleaved caspase-3 in neutrophils were determined via western blotting and immunofluorescence. Neutrophil apoptosis was detected using the terminal deoxynucleotidyl transferase dUTP nick end labeling assay. Our results suggest that TPL inhibited inflammatory cell infiltration in rat ankle joints and inflammatory factor and chemokine secretion in rat serum, regulated macrophage polarization through the PI3K/AKT signaling pathway, suppressed inflammatory factor and chemokine expression in neutrophils, and inhibited neutrophil migration, neutrophil extracellular trap formation, transitional autophagy, and apoptosis. This suggests that TPL can prevent and treat MSU-induced AGA by regulating macrophage polarization through the PI3K/Akt pathway and modulating neutrophil activity.
Asunto(s)
Artritis Gotosa , Diterpenos , Compuestos Epoxi , Macrófagos , Neutrófilos , Fenantrenos , Ácido Úrico , Animales , Diterpenos/farmacología , Diterpenos/uso terapéutico , Neutrófilos/inmunología , Neutrófilos/efectos de los fármacos , Neutrófilos/metabolismo , Ratas , Fenantrenos/farmacología , Fenantrenos/uso terapéutico , Artritis Gotosa/tratamiento farmacológico , Artritis Gotosa/inducido químicamente , Artritis Gotosa/metabolismo , Artritis Gotosa/inmunología , Macrófagos/inmunología , Macrófagos/metabolismo , Macrófagos/efectos de los fármacos , Ratones , Compuestos Epoxi/farmacología , Masculino , Modelos Animales de Enfermedad , Células RAW 264.7 , Transducción de Señal/efectos de los fármacos , Activación de Macrófagos/efectos de los fármacos , Apoptosis/efectos de los fármacos , Activación Neutrófila/efectos de los fármacos , Humanos , Ratas Sprague-Dawley , Movimiento Celular/efectos de los fármacosRESUMEN
Excessive neutrophil infiltration into the tumor microenvironment (TME) is an important factor that contributes to tumor overgrowth and limited immunotherapy efficacy. Neutrophils activate various receptors involved in tumor progression, while suppressing the infiltration and activity of cytotoxic T cells and creating optimal conditions for tumor growth. Therefore, the appropriate control of neutrophil infiltration is an effective strategy for tumor treatment. In the present study, 1-palmitoyl-2-linoleoyl-3-acetyl-rac-glycerol (PLAG) inhibited tumor overgrowth by suppressing excessive neutrophil infiltration, resulting in >74.97â¯% reduction in tumor size in a Lewis lung carcinoma (LLC-1) mouse model. All subjects in the positive control group died during the 90-day survival period, whereas only four subjects in the PLAG treatment group survived. PLAG had a significantly higher tumor growth inhibitory effect and survival rate than other neutrophil infiltration-targeting inhibitors (e.g., Navarixin, lymphocyte antigen 6 complex locus G6D antibody [aLy6G]). The ability of PLAG to regulate neutrophil infiltration and inhibit tumor growth depends on thioredoxin-interacting protein (TXNIP). In tumors lacking TXNIP expression, PLAG failed to control neutrophil infiltration and infiltration-related factor release, and the inhibitory effect of PLAG on tumor growth was reduced. PLAG-mediated inhibition of neutrophil infiltration enhances the efficacy of immune checkpoint inhibitors (ICIs), increasing the antitumor efficacy and survival rate by 30â¯%. In conclusion, PLAG could be a novel alternative to anti-tumor drugs that effectively targets excessive neutrophil infiltration into cancer tissues.
Asunto(s)
Carcinoma Pulmonar de Lewis , Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Ratones Endogámicos C57BL , Infiltración Neutrófila , Microambiente Tumoral , Animales , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/inmunología , Carcinoma Pulmonar de Lewis/tratamiento farmacológico , Carcinoma Pulmonar de Lewis/patología , Carcinoma Pulmonar de Lewis/inmunología , Ratones , Infiltración Neutrófila/efectos de los fármacos , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/patología , Carcinoma de Pulmón de Células no Pequeñas/inmunología , Microambiente Tumoral/efectos de los fármacos , Diglicéridos/farmacología , Línea Celular Tumoral , Neutrófilos/efectos de los fármacos , Neutrófilos/inmunología , Neutrófilos/metabolismo , Modelos Animales de Enfermedad , Masculino , Antineoplásicos/farmacología , GlicéridosRESUMEN
Cathepsin C (CatC) is an enzyme which regulates the maturation of neutrophil serine proteases (NSPs) essential for neutrophil activation. Activated neutrophils are key players in the innate immune system, and are also implicated in the etiology of various inflammatory diseases. This study aims to demonstrate a therapeutic potential for CatC inhibitors against disorders in which activated neutrophil-derived neutrophil extracellular traps (NETs) play a significant role. We demonstrate that a CatC inhibitor, MOD06051, dose-dependently suppresses the cellular activity of NSPs, including neutrophil elastase (NE), in vitro. Neutrophils derived from MOD06051-administered rats exhibit significantly lower NE activity and NET-forming ability than controls. Furthermore, MOD06051 dose-dependently ameliorates vasculitis and significantly decreases NETs when administered to a rat model of myeloperoxidase (MPO)-antineutrophil cytoplasmic antibody-associated vasculitis (AAV). These findings suggest that CatC inhibition is a promising strategy to reduce neutrophil activation and improve activated neutrophil-mediated diseases such as MPO-AAV.
Asunto(s)
Catepsina C , Trampas Extracelulares , Elastasa de Leucocito , Activación Neutrófila , Neutrófilos , Peroxidasa , Catepsina C/metabolismo , Catepsina C/antagonistas & inhibidores , Animales , Neutrófilos/inmunología , Neutrófilos/efectos de los fármacos , Trampas Extracelulares/efectos de los fármacos , Trampas Extracelulares/inmunología , Trampas Extracelulares/metabolismo , Activación Neutrófila/efectos de los fármacos , Humanos , Ratas , Elastasa de Leucocito/metabolismo , Elastasa de Leucocito/antagonistas & inhibidores , Masculino , Peroxidasa/metabolismo , Peroxidasa/antagonistas & inhibidores , Serina Proteasas/metabolismo , Ratas Sprague-Dawley , Modelos Animales de Enfermedad , Vasculitis Asociada a Anticuerpos Citoplasmáticos Antineutrófilos/tratamiento farmacológico , Vasculitis Asociada a Anticuerpos Citoplasmáticos Antineutrófilos/inmunologíaRESUMEN
Intravenous thrombolysis using recombinant tissue plasminogen activator (tPA) remains the primary treatment for patients with acute ischemic stroke (AIS). However, the mechanism of tPA-related hemorrhagic transformation (HT) remains poorly understood. Elevation of histidine-rich glycoprotein (HRG) expression was detected by nano-liquid chromatography tandem mass spectrometry at 1 h following tPA infusion as compared to baseline prior to tPA infusion (discovery cohort, n = 10), which was subsequently confirmed in a validation cohort (n = 157) by ELISA. Surprisingly, no elevation of HRG was detected in individuals who subsequently developed HT. During in vitro experiments, HRG reduced neutrophil NETosis, inflammatory cytokine production, and migration across the blood-brain barrier induced by tPA. In a photothrombotic murine AIS model, HRG administration ameliorated HT with delayed thrombolysis, by inhibiting neutrophil immune infiltration and downregulating pro-inflammatory signaling pathways. Neutrophil depletion or NETosis inhibition also alleviated HT, whereas HRG siRNA treatment exacerbated HT. In conclusion, fluctuations in HRG levels may reflect tPA therapy and its associated HT. The inhibitory effect of HRG on neutrophils may counteract tPA-induced immune abnormalities and HT in patients with AIS.
Asunto(s)
Neutrófilos , Activador de Tejido Plasminógeno , Animales , Neutrófilos/metabolismo , Neutrófilos/efectos de los fármacos , Neutrófilos/inmunología , Humanos , Ratones , Activador de Tejido Plasminógeno/metabolismo , Activador de Tejido Plasminógeno/efectos adversos , Masculino , Femenino , Anciano , Persona de Mediana Edad , Proteínas/metabolismo , Terapia Trombolítica/efectos adversos , Terapia Trombolítica/métodos , Modelos Animales de Enfermedad , Fibrinolíticos/farmacología , Glicoproteínas/metabolismo , Ratones Endogámicos C57BL , Accidente Cerebrovascular Isquémico/tratamiento farmacológicoRESUMEN
Type 2 diabetes mellitus (T2DM) is accompanied by halogenative stress resulting from the excessive activation of neutrophils and neutrophilic myeloperoxidase (MPO) generating highly reactive hypochlorous acid (HOCl). HOCl in blood plasma modifies serum albumin (Cl-HSA). We studied the formation of neutrophil extracellular traps (NETs) in the whole blood and by isolated neutrophils under the action of Cl-HSA. It was found that Cl-HSA induces neutrophil priming and NETosis. MPO-containing as well as MPO-free NETs were found. These NETs with different composition can be a product of NETosis of one and the same neutrophil. NET formation in neutrophils with vacuolated cytoplasm was detected. In the presence of Cl-HSA, acceleration of NET degradation was observed. Accelerated NET degradation and neutrophil priming can be the factors contributing to the development of complications in T2DM.
Asunto(s)
Trampas Extracelulares , Ácido Hipocloroso , Neutrófilos , Peroxidasa , Ácido Hipocloroso/metabolismo , Ácido Hipocloroso/farmacología , Neutrófilos/metabolismo , Neutrófilos/efectos de los fármacos , Trampas Extracelulares/metabolismo , Trampas Extracelulares/efectos de los fármacos , Humanos , Peroxidasa/metabolismo , Diabetes Mellitus Tipo 2/sangre , Albúmina Sérica/metabolismo , MasculinoRESUMEN
Glucocorticoids are a major class of therapeutic anti-inflammatory and immunosuppressive drugs prescribed to patients with inflammatory diseases, to avoid transplant rejection, and as part of cancer chemotherapy. However, exposure to these drugs increases the risk of opportunistic infections such as with the fungus Aspergillus fumigatus, which causes mortality in >50% of infected patients. The mechanisms by which glucocorticoids increase susceptibility to A. fumigatus are poorly understood. In this article, we used a zebrafish larva Aspergillus infection model to identify innate immune mechanisms altered by glucocorticoid treatment. Infected larvae exposed to dexamethasone succumb to infection at a significantly higher rate than control larvae. However, both macrophages and neutrophils are still recruited to the site of infection, and dexamethasone treatment does not significantly affect fungal spore killing. Instead, the primary effect of dexamethasone manifests later in infection with treated larvae exhibiting increased invasive hyphal growth. In line with this, dexamethasone predominantly inhibits neutrophil function rather than macrophage function. Dexamethasone-induced mortality also depends on the glucocorticoid receptor. Dexamethasone partially suppresses NF-κB activation at the infection site by inducing the transcription of IκB via the glucocorticoid receptor. Independent CRISPR/Cas9 targeting of IKKγ to prevent NF-κB activation also increases invasive A. fumigatus growth and larval mortality. However, dexamethasone treatment of IKKγ crispant larvae further increases invasive hyphal growth and host mortality, suggesting that dexamethasone may suppress other pathways in addition to NF-κB to promote host susceptibility. Collectively, we find that dexamethasone acts through the glucocorticoid receptor to suppress NF-κB-mediated neutrophil control of A. fumigatus hyphae in zebrafish larvae.