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Defective autophagy is linked to proinflammatory diseases. However, the mechanisms by which autophagy limits inflammation remain elusive. Here, we found that the pan-FGFR inhibitor LY2874455 efficiently activated autophagy and suppressed expression of proinflammatory factors in macrophages stimulated by lipopolysaccharide (LPS). Multiplex proteomic profiling identified the immunoproteasome, which is a specific isoform of the 20s constitutive proteasome, as a substrate that is degraded by selective autophagy. SQSTM1/p62 was found to be a selective autophagy-related receptor that mediated this degradation. Autophagy deficiency or p62 knockdown blocked the effects of LY2874455, leading to the accumulation of immunoproteasomes and increases in inflammatory reactions. Expression of proinflammatory factors in autophagy-deficient macrophages could be reversed by immunoproteasome inhibitors, confirming the pivotal role of immunoproteasome turnover in the autophagy-mediated suppression on the expression of proinflammatory factors. In mice, LY2874455 protected against LPS-induced acute lung injury and dextran sulfate sodium (DSS)-induced colitis and caused low levels of proinflammatory cytokines and immunoproteasomes. These findings suggested that selective autophagy of the immunoproteasome was a key regulator of signaling via the innate immune system.
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Lipopolisacáridos , Proteómica , Animales , Ratones , Lipopolisacáridos/farmacología , Autofagia , Inflamación , Macrófagos , Complejo de la Endopetidasa ProteasomalRESUMEN
Ultra-sensitive detection of cancer-related biomarkers in serum is of great significance for early diagnosis, treatment, prognosis, and staging of cancer. In this work, we proposed a surface-enhanced Raman scattering and fluorescence (SERS/FL) dual-mode biosensor for hepatocellular carcinoma (HCC)-related miRNA (miR-224) detection using the composition of well-arranged Au nanoarrays (Au NAs) substrate coupled with the target-catalyzed hairpin assembly (CHA) strategy. The hot spots densely and uniformly distributed on the Au array offers considerably enhanced and reproducible SERS signals, along with their wide and open surface to facilitate miR-224 adsorption. By this sensing strategy, the target miR-224 can be detected in a wide linear range (1 fM to 1 nM) with a limit of detection of 0.34 fM in the SERS mode and 0.39 fM in the FL mode. Meanwhile, this biosensor with exceptional specificity and anti-interference ability can discriminate target miR-224 from other interference miRNAs. Practical analysis of human blood samples also demonstrated considerable reliability and repeatability of our developed strategy. Furthermore, this biosensor can distinguish HCC cancer subjects from normal ones and monitor HCC patients before and after hepatectomy as well as guide the distinct Barcelona clinic liver cancer (BCLC) stages. Overall, benefiting from a well-arranged Au nanoarray, CHA amplification strategy, and SERS/metal enhanced fluorescence effect, this established biosensor opens new avenues for the early prediction, warning, monitoring, and staging of HCC.
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Técnicas Biosensibles , Carcinoma Hepatocelular , Neoplasias Hepáticas , Nanopartículas del Metal , MicroARNs , Humanos , Carcinoma Hepatocelular/diagnóstico , Oro/química , Límite de Detección , Neoplasias Hepáticas/diagnóstico , Nanopartículas del Metal/química , Nanoestructuras , Reproducibilidad de los Resultados , Espectrometría Raman , Colorantes Fluorescentes/químicaRESUMEN
A new lateral flow immunoassay strip (LFIA) combining sensitive detection and identification of multiple bacteria remains a huge challenge. In this study, we first developed multifunctional urchin-shaped Au-Ag@Pt nanoparticles (UAA@P NPs) with a unique combination of colorimetric-SERS-photothermal-catalytic (CM/SERS/PT/CL) properties and integrated them with LFIA for multiplexed detection and specific discrimination of pathogenic bacteria in blood samples. Unlike the conventional LFIA that relied on antibody (Ab), this novel LFIA introduced 4-mercaptophenylboronic acid (4-MPBA) as an ideal Ab replacer that was functionalized on UAA@P NPs (UAA@P/M NPs) with outstanding binding and enrichment capacities toward bacteria. Taking Staphylococcus aureus (S. aureus) as model bacteria, the limit of detection (LOD) was 3 CFU/mL for SERS-LFIA, 27 CFU/mL for PT-LFIA, and 18 CFU/mL for CL-LFIA, three of which were over 330-fold, 37-fold, and 55-fold more sensitive than ordinary visual CM-LFIA, respectively. Besides, this SERS-LFIA is capable of identifying three types of bacterial spiked blood samples (E. coli, S. aureus, and P. aeruginosa) effectively according to specific bacterial Raman "fingerprints" by partial least-squares-discriminant analysis (PLS-DA). More importantly, this LFIA was successfully applied to blood samples with satisfactory recoveries from 90.3% to 108.8% and capable of identifying the infected patients (N = 4) from healthy subjects (N = 2) with great accuracy. Overall, the multimodal LFIA incorporates bacteria discrimination and quantitative detection, offering an avenue for early warning and diagnosis of bacterial infection.
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Infecciones Bacterianas , Nanopartículas del Metal , Humanos , Escherichia coli , Staphylococcus aureus , Inmunoensayo , Bacterias , Anticuerpos , Infecciones Bacterianas/diagnóstico , Límite de Detección , Nanopartículas del Metal/química , Oro/químicaRESUMEN
α4 integrin plays a crucial role in retention and release of neutrophils from bone marrow. Although α4 integrin is known to be a potential target of reactive oxygen species (ROS)-induced cysteine glutathionylation, the physiological significance and underlying regulatory mechanism of this event remain elusive. Here, using in vitro and in vivo biochemical and cell biology approaches, we show that physiological ROS-induced glutathionylation of α4 integrin in neutrophils increases the binding of neutrophil-associated α4 integrin to vascular cell adhesion molecule 1 (VCAM-1) on human endothelial cells. This enhanced binding was reversed by extracellular glutaredoxin 1 (Grx1), a thiol disulfide oxidoreductase promoting protein deglutathionylation. Furthermore, in a murine inflammation model, Grx1 disruption dramatically elevated α4 glutathionylation and subsequently enhanced neutrophil egress from the bone marrow. Corroborating this observation, intravenous injection of recombinant Grx1 into mice inhibited α4 glutathionylation and thereby suppressed inflammation-induced neutrophil mobilization from the bone marrow. Taken together, our results establish ROS-elicited glutathionylation and its modulation by Grx1 as pivotal regulatory mechanisms controlling α4 integrin affinity and neutrophil mobilization from the bone marrow under physiological conditions.
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Médula Ósea/metabolismo , Glutarredoxinas/metabolismo , Integrina alfa4/metabolismo , Neutrófilos/metabolismo , Regulación hacia Arriba , Molécula 1 de Adhesión Celular Vascular/metabolismo , Animales , Médula Ósea/patología , Modelos Animales de Enfermedad , Glutarredoxinas/genética , Células HL-60 , Humanos , Inflamación/genética , Inflamación/metabolismo , Inflamación/patología , Integrina alfa4/genética , Ratones Noqueados , Neutrófilos/patología , Molécula 1 de Adhesión Celular Vascular/genéticaRESUMEN
Non-tuberculous mycobacteria (NTM), also known as an environmental and atypical mycobacteria, can cause the chronic pulmonary infectious diseases. Macrophages have been suggested as the main host cell to initiate the innate immune responses to NTM infection. However, the molecular mechanism to regulate the antimicrobial immune responses to NTM is still largely unknown. Current study showed that the NTM clinical groups, Mycobacterium abscessus and Mycobacterium smegmatis, significantly induced the M1 macrophage polarization with the characteristic production of nitric oxide (NO) and marker gene expression of iNOS, IFNγ, TNF-α, IL1-ß and IL-6. Interestingly, a non-histone nuclear protein, HMGN2 (high-mobility group N2), was found to be spontaneously induced during NTM-activated M1 macrophage polarization. Functional studies revealed that HMGN2 deficiency in NTM-infected macrophage promotes the expression of M1 markers and the production of NO via the enhanced activation of NF-κB and MAPK signalling. Further studies exhibited that HMGN2 knock-down also enhanced IFNγ-induced M1 macrophage polarization. Finally, we observed that silencing HMGN2 affected the survival of NTM in macrophage, which might largely relevant to enhanced macrophage polarization into M1 phenotype under the NTM infection. Collectively, current studies thus suggested a novel function of HMGN2 in regulating the anti-non-tuberculous mycobacteria innate immunity of macrophage.
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Proteína HMGN2/metabolismo , Activación de Macrófagos/genética , Macrófagos/metabolismo , Infecciones por Mycobacterium/inmunología , Micobacterias no Tuberculosas/crecimiento & desarrollo , Animales , Supervivencia Celular/genética , Técnicas de Silenciamiento del Gen , Silenciador del Gen , Proteína HMGN2/genética , Humanos , Inmunidad Innata , Interferón gamma/metabolismo , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Sistema de Señalización de MAP Quinasas/genética , Ratones , Mycobacterium abscessus/inmunología , Mycobacterium abscessus/aislamiento & purificación , Mycobacterium smegmatis/inmunología , Mycobacterium smegmatis/aislamiento & purificación , FN-kappa B/metabolismo , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo II/metabolismo , Células RAW 264.7 , Interferencia de ARN , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Urinary tract infection is one of the most common bacterial infections which is mainly caused by Escherichia coli (UPEC). Autophagy plays a key role in immune response to eliminate invading pathogens. Exploring the effect of autophagy on UPEC infection and the molecular mechanisms will be benefit for the treatment of urinary tract infection. High-mobility group protein N2 (HMGN2), a highly conserved nuclear protein and an antibacterial peptide, has been associated with bacterial infection induced immune response; however, whether this function is due to the regulation of autophagy remains unclear. In this study, we demonstrate for the first time that HMGN2 is upregulated in UPEC infection of bladder epithelial cell line 5637 (BEC 5637). Furthermore, HMGN2 enhances autophagy in BEC 5637 via activation of AMPK and ULK1, whereas UPEC suppresses autophagy. In addition, the enhanced autophagy activity by HMGN2 overexpression or rapamycin boosts the proliferation of UPEC J96 in BEC 5637. In summary, our data indicate that HMGN2 activates autophagy via AMPK/ULK1 pathway which can be utilized by UPEC J96 for their proliferation within bladder epithelial cells.
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Proteínas Quinasas Activadas por AMP/metabolismo , Homólogo de la Proteína 1 Relacionada con la Autofagia/metabolismo , Infecciones por Escherichia coli/metabolismo , Proteína HMGN2/metabolismo , Vejiga Urinaria/microbiología , Infecciones Urinarias/metabolismo , Animales , Autofagia , Línea Celular , Proliferación Celular , Células Epiteliales/citología , Células Epiteliales/metabolismo , Células Epiteliales/microbiología , Escherichia coli/crecimiento & desarrollo , Escherichia coli/metabolismo , Infecciones por Escherichia coli/microbiología , Femenino , Humanos , Ratones Endogámicos C57BL , Transducción de Señal , Vejiga Urinaria/citología , Vejiga Urinaria/metabolismo , Infecciones Urinarias/microbiologíaRESUMEN
BACKGROUND: Inhalation of environmental (nano) particles (NP) as well as persistent herpesvirus-infection are potentially associated with chronic lung disease and as both are omnipresent in human society a coincidence of these two factors is highly likely. We hypothesized that NP-exposure of persistently herpesvirus-infected cells as a second hit might disrupt immune control of viral latency, provoke reactivation of latent virus and eventually lead to an inflammatory response and tissue damage. RESULTS: To test this hypothesis, we applied different NP to cells or mice latently infected with murine gammaherpesvirus 68 (MHV-68) which provides a small animal model for the study of gammaherpesvirus-pathogenesis in vitro and in vivo. In vitro, NP-exposure induced expression of the typically lytic viral gene ORF50 and production of lytic virus. In vivo, lytic viral proteins in the lung increased after intratracheal instillation with NP and elevated expression of the viral gene ORF50 could be detected in cells from bronchoalveolar lavage. Gene expression and metabolome analysis of whole lung tissue revealed patterns with striking similarities to acute infection. Likewise, NP-exposure of human cells latently infected with Epstein-Barr-Virus also induced virus production. CONCLUSIONS: Our results indicate that NP-exposure of persistently herpesvirus-infected cells - murine or human - restores molecular signatures found in acute virus infection, boosts production of lytic viral proteins, and induces an inflammatory response in the lung - a combination which might finally result in tissue damage and pathological alterations.
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Gammaherpesvirinae/efectos de los fármacos , Infecciones por Herpesviridae/virología , Nanopartículas/toxicidad , Activación Viral/efectos de los fármacos , Animales , Línea Celular , Cricetinae , Gammaherpesvirinae/fisiología , Ratones , Células 3T3 NIH , Latencia del Virus , Replicación ViralRESUMEN
Apoptosis is a tightly regulated cellular process that plays an essential role in the development, aging, cancer biology, immune response, and pathogenesis of various diseases. Herein, we report a new SERS sensing strategy for in vitro sensitive detection of early apoptotic cells. The principle of this method is to in situ synthesize silver nanoparticles (AgNPs) on the phosphatidylserine (PS) of the apoptotic cell membrane during the early apoptosis, which enables distinguishing normal and apoptotic cells. The total assay time of the presented method is only 10 min, thus being faster, cheaper and simpler than current techniques for the detection of apoptosis. The intrinsic mechanism was verified by different approaches based on externalized phosphatidylserine. In addition, the detection process is real-time and label-free; i.e., the intrinsic SERS spectra from the cellular membrane are directly employed for apoptosis real-time detection, which avoids using additional chemical or biological reagents as external signal indicators. Therefore, our SERS approach may serve as a potentially practical tool for sensitive and real-time detection of early cell apoptosis, complementing the state-of-the-art strategies, e.g. flow cytometry. While further investigation is required to better understand the intrinsic mechanism of the in situ coating method, the current results may provide another choice for real-time detection of early apoptosis.
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BACKGROUND: Carbonaceous nanoparticles (CNP) represent a major constituent of urban particulate air pollution, and inhalation of high CNP levels has been described to trigger a pro-inflammatory response of the lung. While several studies identified specific particle characteristics driving respiratory toxicity of low-solubility and low-toxicity particles such as CNP, the major lung cell type, which initiates and drives that response, remains still uncertain. Since alveolar macrophages (AM) are known to effectively phagocytose inhaled particles and play a crucial role for the initiation of pulmonary inflammation caused by invading microbes, we aimed to determine their role for sterile stimuli such as CNP by profiling the primary alveolar cell compartments of the lung. We exposed C57BL/6 mice to 20 µg CNP by intratracheal instillation and comprehensively investigated the expression of the underlying mediators during a time span of 3 to 72 h in three different lung cell populations: CD45- (negative) structural cells, CD45+ (positive) leukocytes, and by BAL recovered cells. RESULTS: Bronchoalveolar lavage (BAL) analysis revealed an acute inflammatory response characterized by the most prominent culmination of neutrophil granulocytes from 12 to 24 h after instillation, which declined to basal levels by day 7. As early as 3 h after CNP exposure 50 % of the AM revealed particle laden. BAL concentrations and lung gene expression profiles of TNFα, and the neutrophil chemoattractants CXCL1,-2 and-5 preceded the neutrophil recruitment and showed highest levels after 12 h of CNP exposure, pointing to a significant activation of the inflammation-evoking lung cells at this point of time. AM, isolated from lungs 3 to 12 h after CNP instillation, however, did not show a pro-inflammatory signature. On the contrary, gene expression analysis of different lung cell populations isolated 12 h after CNP instillation revealed CD45-, mainly representing alveolar epithelial type II (ATII) cells as major producer of inflammatory CXCL cytokines. Particularly by CD45- cells expressed Cxcl5 proved to be the most abundant chemokine, being 12 h after CNP exposure 24 (±11) fold induced. CONCLUSION: Our data suggests that AM are noninvolved in the initiation of the inflammatory response. ATII cells, which induced highest CXCL levels early on, might in contrast be the driver of acute neutrophilic inflammation upon pulmonary CNP exposure.
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Carbono/toxicidad , Macrófagos Alveolares/citología , Nanopartículas/toxicidad , Neumonía/inducido químicamente , Enfermedad Aguda , Animales , Líquido del Lavado Bronquioalveolar , Carbono/química , Quimiocinas/metabolismo , Ratones , Nanopartículas/química , Neutrófilos/citología , Neumonía/metabolismoRESUMEN
Uropathogenic Escherichia coli (UPEC) biofilm formation enables the organism to avoid the host immune system, resist antibiotics, and provide a reservoir for persistent infection. Once the biofilm is established, eradication of the infection becomes difficult. Therefore, strategies against UPEC biofilm are urgently required. In this study, we investigated the effect of allicin, isolated from garlic essential oil, on UPEC CFT073 and J96 biofilm formation and dispersal, along with its effect on UPEC adhesion ability and swimming motility. Sub-inhibitory concentrations (sub-MICs) of allicin decreased UPEC biofilm formation and affected its architecture. Allicin was also capable of dispersing biofilm. Furthermore, allicin decreased the bacterial adhesion ability and swimming motility, which are important for biofilm formation. Real-time quantitative polymerase chain reaction (RT-qPCR) revealed that allicin decreased the expression of UPEC type 1 fimbriae adhesin gene fimH. Docking studies suggested that allicin was located within the binding pocket of heptyl α-d-mannopyrannoside in FimH and formed hydrogen bonds with Phe1 and Asn135. In addition, allicin decreased the expression of the two-component regulatory systems (TCSs) cognate response regulator gene uvrY and increased the expression of the RNA binding global regulatory protein gene csrA of UPEC CFT073, which is associated with UPEC biofilm. The findings suggest that sub-MICs of allicin are capable of affecting UPEC biofilm formation and dispersal, and decreasing UPEC adhesion ability and swimming motility.
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Biopelículas/efectos de los fármacos , Ácidos Sulfínicos/farmacología , Escherichia coli Uropatógena/efectos de los fármacos , Escherichia coli Uropatógena/fisiología , Adhesinas de Escherichia coli/genética , Adhesinas de Escherichia coli/metabolismo , Adhesión Bacteriana/efectos de los fármacos , Disulfuros , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Regulación Bacteriana de la Expresión Génica/genética , Escherichia coli Uropatógena/crecimiento & desarrollo , Escherichia coli Uropatógena/metabolismoRESUMEN
This paper describes a novel double-stranded DNA detection method through resonance between SYBR Green I and DNA with the surface-enhanced resonance Raman scattering (SERRS) assay, which opens an avenue to the quantitative and reliable application of SERRS in DNA detection.
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ADN/análisis , Colorantes Fluorescentes/química , Compuestos Orgánicos/química , Reacción en Cadena de la Polimerasa/métodos , Espectrometría Raman/métodos , Benzotiazoles , Diaminas , QuinolinasRESUMEN
BACKGROUND: Persons with cystic fibrosis (CF) are at-risk for health effects from ambient air pollution but little is known about the interaction of nanoparticles (NP) with CF lungs. Here we study the distribution of inhaled NP in a murine CF model and aim to reveal mechanisms contributing to adverse effects of inhaled particles in susceptible populations. METHODS: Chloride channel defective CftrTgH (neoim) Hgu mice were used to analyze lung function, lung distribution and whole body biokinetics of inhaled NP, and inflammatory responses after intratracheal administration of NP. Distribution of 20-nm titanium dioxide NP in lungs was assessed on ultrathin sections immediately and 24 h after a one-hour NP inhalation. NP biokinetics was deduced from total and regional lung deposition and from whole body translocation of inhaled 30-nm iridium NP within 24 h after aerosol inhalation. Inflammatory responses were assessed within 7 days after carbon NP instillation. RESULTS: Cftr mutant females had moderately reduced lung compliance and slightly increased airway resistance compared to wild type mice. We found no genotype dependent differences in total, regional and head deposition or in secondary-organ translocation of inhaled iridium NP. Titanium dioxide inhalation resulted in higher NP uptake by alveolar epithelial cells in Cftr mutants. Instillation of carbon NP induced a comparable acute and transient inflammatory response in both genotypes. The twofold increase of bronchoalveolar lavage (BAL) neutrophils in Cftr mutant compared to wild type mice at day 3 but not at days 1 and 7, indicated an impaired capacity in inflammation resolution in Cftr mutants. Concomitant to the delayed decline of neutrophils, BAL granulocyte-colony stimulating factor was augmented in Cftr mutant mice. Anti-inflammatory 15-hydroxyeicosatetraenoic acid was generally significantly lower in BAL of Cftr mutant than in wild type mice. CONCLUSIONS: Despite lacking alterations in lung deposition and biokinetics of inhaled NP, and absence of significant differences in lung function, higher uptake of NP by alveolar epithelial cells and prolonged, acute inflammatory responses to NP exposure indicate a moderately increased susceptibility of lungs to adverse effects of inhaled NP in Cftr mutant mice and provides potential mechanisms for the increased susceptibility of CF patients to air pollution.
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Contaminantes Atmosféricos/farmacocinética , Contaminantes Atmosféricos/toxicidad , Fibrosis Quística/patología , Nanopartículas/toxicidad , Contaminación del Aire , Animales , Líquido del Lavado Bronquioalveolar , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Modelos Animales de Enfermedad , Femenino , Inmunohistoquímica , Exposición por Inhalación , Iridio/farmacocinética , Iridio/toxicidad , Radioisótopos de Iridio , Pulmón/efectos de los fármacos , Pulmón/patología , Masculino , Ratones , Ratones Endogámicos CFTR , Ratones Transgénicos , Microscopía Electrónica de Transmisión , Neumonía/inducido químicamente , Neumonía/patología , Pruebas de Función Respiratoria , Titanio/toxicidadRESUMEN
OBJECTIVES: Acute lung injury (ALI) is a highly inflammatory condition with the involvement of M1 alveolar macrophages (AMs) polarization, eventually leading to the development of non-cardiogenic edema in alveolar and interstitial regions, accompanied by persistent hypoxemia. Given the significant mortality rate associated with ALI, it is imperative to investigate the underlying mechanisms of this condition so as to identify potential therapeutic targets. The therapeutic effects of the inhibition of bromodomain containing protein 4 (BRD4), an epigenetic reader, has been proven with high efficacy in ameliorating various inflammatory diseases through mediating immune cell activation. However, little is known about the therapeutic potential of BRD4 degradation in acute lung injury. METHODS: This study aimed to assess the protective efficacy of ARV-825, a novel BRD4-targeted proteolysis targeting chimera (PROTAC), against ALI through histopathological examination in lung tissues and biochemical analysis in bronchoalveolar lavage fluid (BALF). Additionally, the underlying mechanism by which BRD4 regulated M1 AMs was elucidated by using CUT & Tag assay. RESULTS: In this study, we found the upregulation of BRD4 in a lipopolysaccharide (LPS)-induced ALI model. Furthermore, we observed that intraperitoneal administration of ARV-825, significantly alleviated LPS-induced pulmonary pathological changes and inflammatory responses. These effects were accompanied by the suppression of M1 AMs. In addition, our findings revealed that the administration of ARV-825 effectively suppressed M1 AMs by inhibiting the expression of IRF7, a crucial transcriptional factor involved in M1 macrophages. CONCLUSION: Our study suggested that targeting BRD4 using ARV-825 is a potential therapeutic approach for ALI.
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Lesión Pulmonar Aguda , Proteínas que Contienen Bromodominio , Lipopolisacáridos , Macrófagos Alveolares , Factores de Transcripción , Animales , Lesión Pulmonar Aguda/tratamiento farmacológico , Lesión Pulmonar Aguda/inducido químicamente , Lesión Pulmonar Aguda/patología , Lesión Pulmonar Aguda/inmunología , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Macrófagos Alveolares/efectos de los fármacos , Macrófagos Alveolares/inmunología , Macrófagos Alveolares/metabolismo , Ratones , Masculino , Ratones Endogámicos C57BL , Humanos , Proteolisis/efectos de los fármacos , Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico , Pulmón/patología , Pulmón/efectos de los fármacos , Pulmón/inmunología , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Modelos Animales de Enfermedad , Activación de Macrófagos/efectos de los fármacosRESUMEN
Hyper-IgE syndrome (HIES) is a primary immunodeficiency characterized by, among others, the excessive production of IgE and repetitive bacterial/fungal infections. Mutations in STAT3, a transcription factor that orchestrates immune responses, may cause HIES, but the underlying mechanisms are not fully understood. Here, we used multi-omic approaches to comprehensively decipher the immune disturbance in a male HIES patient harboring STAT3-V637M. In his peripheral blood mononuclear cell (PBMC) we found significant clonal expansion of CD8 T cells (with increased CD8 subunits expression, potentially enhancing responsiveness to MHC I molecules), but not in his CD4 T cells and B cells. Although his B cells exhibited a higher potential in producing immunoglobulin, elevated SPIC binding might bias the products toward IgE isotype. Immune checkpoint inhibitors, including CTLA4, LAG3, were overexpressed in his PBMC-CD4 T cells, accompanied by reduced CD28 and IL6ST (gp130) expression. In his CD4 T cells, integrative analyses predicted upstream transcription factors (including ETV6, KLF13, and RORA) for LAG3, IL6ST, and CD28, respectively. The down-regulation of phagocytosis and nitric oxide synthesis-related genes in his PBMC-monocytes seem to be the culprit of his disseminated bacterial/fungal infection. Counterintuitively, in his PBMC we predicted increased STAT3 binding in both naïve and mature CD4 compartments, although this was not observed in most of his PBMC. In his bronchoalveolar lavage fluid (BALF), we found two macrophage subtypes with anti-bacterial properties, which were identified by CXCL8/S100A8/S100A9, or SOD2, respectively. Together, we described how the immune cell landscape was disturbed in STAT3-V637M HIES, providing a resource for further studies.
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Síndrome de Job , Leucocitos Mononucleares , Humanos , Masculino , Antígenos CD28 , Síndrome de Job/genética , Multiómica , Inmunoglobulina E , Factor de Transcripción STAT3/genéticaRESUMEN
BACKGROUND: Ovarian cancer (OC) typically develops an immunosuppressive microenvironment by funtional changes of host immune cells. Dysregulated m6A level is associated with cancer progression via the intrinsic oncogenic pathways. However, the role of m6A in regulating host immune cell function during anti-tumor immunity needs comprehensive analysis. This study aimed to investigate the role of METTL3, a catalytic subunit of the methyltransferase complex, in regulating host immune cell response against OC. METHODS: In this study, myeloid-specific Mettl3 gene knockout (Mettl3-cKO) mice were bred using the Cre-LoxP system. Intraperitoneally injection of ID8 cells was used as a syngeneic OC model. Furthermore, the compositions of immune cell populations were analyzed by flow cytometry and single-cell sequencing. Moreover, chemokines and cytokines secretion were assessed using ELISA. Lastly, the role of METTL3 in regulating IL-1ß secretion and inflammasome activation in bone marrow-derived macrophages cocultured with ID8 cells was specified by ELISA and immunoblotting. RESULTS: It was revealed that OC cell growth was enhanced in Mettl3-cKO mice. Furthermore, a shift of decreased M1 to increased M2 macrophage polarization was observed during OC progression. Moreover, Mettl3 depletion in myeloid lineage cells increased secretion of CCL2 and CXCL2 in peritoneal lavage fluild. Interestingly, Mettl3 deficiency enhanced IL-1ß secretion induced by viable ID8 cells independent of inflammasome activation and cell death. Therefore, OC cells in tumor-bearing mice trigger a slight inflammatory response with a low-to-moderate secretion of pro-inflammatory cytokines and chemokines. CONCLUSION: This study provides new insights into METTL3-mediated m6A methylation, which regulates host immune response against OC.
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Macrophages exist in various tissues, several body cavities, and around mucosal surfaces and are a vital part of the innate immune system for host defense against many pathogens and cancers. Macrophages possess binary M1/M2 macrophage polarization settings, which perform a central role in an array of immune tasks via intrinsic signal cascades and, therefore, must be precisely regulated. Many crucial questions about macrophage signaling and immune modulation are yet to be uncovered. In addition, the clinical importance of tumor-associated macrophages is becoming more widely recognized as significant progress has been made in understanding their biology. Moreover, they are an integral part of the tumor microenvironment, playing a part in the regulation of a wide variety of processes including angiogenesis, extracellular matrix transformation, cancer cell proliferation, metastasis, immunosuppression, and resistance to chemotherapeutic and checkpoint blockade immunotherapies. Herein, we discuss immune regulation in macrophage polarization and signaling, mechanical stresses and modulation, metabolic signaling pathways, mitochondrial and transcriptional, and epigenetic regulation. Furthermore, we have broadly extended the understanding of macrophages in extracellular traps and the essential roles of autophagy and aging in regulating macrophage functions. Moreover, we discussed recent advances in macrophages-mediated immune regulation of autoimmune diseases and tumorigenesis. Lastly, we discussed targeted macrophage therapy to portray prospective targets for therapeutic strategies in health and diseases.
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Epigénesis Genética , Neoplasias , Humanos , Macrófagos , Neoplasias/genética , Neoplasias/terapia , Inmunoterapia , Transducción de Señal , Microambiente Tumoral/genéticaRESUMEN
In situ and real-time monitoring of responsive drug release is critical for the assessment of pharmacodynamics in chemotherapy. In this study, a novel pH-responsive nanosystem is proposed for real-time monitoring of drug release and chemo-phototherapy by surface-enhanced Raman spectroscopy (SERS). The Fe3O4@Au@Ag nanoparticles (NPs) deposited graphene oxide (GO) nanocomposites with a high SERS activity and stability are synthesized and labeled with a Raman reporter 4-mercaptophenylboronic acid (4-MPBA) to form SERS probes (GO-Fe3O4@Au@Ag-MPBA). Furthermore, doxorubicin (DOX) is attached to SERS probes through a pH-responsive linker boronic ester (GO-Fe3O4@Au@Ag-MPBA-DOX), accompanying the 4-MPBA signal change in SERS. After the entry into tumor, the breakage of boronic ester in the acidic environment gives rise to the release of DOX and the recovery of 4-MPBA SERS signal. Thus, the DOX dynamic release can be monitored by the real-time changes of 4-MPBA SERS spectra. Additionally, the strong T2 magnetic resonance (MR) signal and NIR photothermal transduction efficiency of the nanocomposites make it available for MR imaging and photothermal therapy (PTT). Altogether, this GO-Fe3O4@Au@Ag-MPBA-DOX can simultaneously fulfill the synergistic combination of cancer cell targeting, pH-sensitive drug release, SERS-traceable detection and MR imaging, endowing it great potential for SERS/MR imaging-guided efficient chemo-phototherapy on cancer treatment.
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Despite the potential indicating role of tyrosinase (TYR) in cutaneous melanoma, how to capture the real changes of TYR in suspicious skin remains a major challenge. Unlike the traditional human serum test, this study reports a sensing platform that incorporates a wearable microneedle (MN) patch and trimetallic Au@Ag-Pt nanoparticles (NPs) for surface-enhanced Raman scattering (SERS) and colorimetric dual-mode detecting TYR in human skin in situ toward potential melanoma screening. In the presence of TYR, catechol immobilized on MN is preferentially oxidized to benzoquinone, which competitively impedes the interaction of MN and Au@Ag-Pt NPs, triggering the SERS-colorimetric signal reciprocal switch. Using a B16F10 mouse melanoma model, our platform is capable of noninvasively piercing the skin surface and detecting TYR levels before and during anti-PD-1 antibody treatment, which would be highly informative for prognostic judgment and illness monitoring of melanoma. Through in situ sensing for capturing the metabolic changes of TYR in advance, this platform was successfully applied to discriminate the melanoma subjects from skin moles and normal ones (p < 0.001), as well as screen potential melanoma from lactate dehydrogenase (LDH)-negative patients. Melanoma growth and prognosis can still be monitored through recording the continuous change of TYR levels. More importantly, the well-defined flexible and stretchable characteristics of the MN patch allow robustly adhering to the skin without inducing chemical or physical irritation. We believe this platform integrating MN-based in situ sensing, TYR responsiveness, and SERS/colorimetric dual-readout strategy will have high clinical importance in early diagnosis and monitoring of cutaneous melanoma.
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Melanoma , Nanopartículas del Metal , Neoplasias Cutáneas , Dispositivos Electrónicos Vestibles , Animales , Ratones , Humanos , Melanoma/diagnóstico , Melanoma/metabolismo , Monofenol Monooxigenasa/metabolismo , Neoplasias Cutáneas/diagnóstico , Espectrometría Raman , Oro , Melanoma Cutáneo MalignoRESUMEN
Peritoneal macrophages (PMs), which resided in peritoneal cavity, are crucial to maintain tissue homeostasis and immunity. Macrophage self-renewal and polarization states are critical for PM population homeostasis and function. However, the underlying molecular mechanism that regulates self-renewal and polarization of PMs is still unclear and needs to be explored. Here, we demonstrated that PMs self-renewal was stimulated by granulocyte macrophage colony-stimulating factor (GM-CSF), but not by macrophage colony-stimulating factor (M-CSF). Pharmacological inhibition of Bromodomain & Extraterminal (BET) Proteins by either JQ1 or ARV-825 significantly reduced GM-CSF-dependent peritoneal macrophage self-renewal by abrogating cell proliferation and decreasing self-renewal-related gene expression, such as MYC and Klf4, at transcriptional and protein levels. In addition, transcriptomic analysis showed that JQ1 blocked alternative PMs polarization by downregulating key transcriptional factor IRF4 expression, but not the activation of AKT or STAT6 in PMs. These findings illustrated that the significance of BET family proteins in GM-CSF-induced PMs self-renewal and IL-4-induced alternative polarization.
RESUMEN
[This corrects the article DOI: 10.1016/j.apsb.2022.08.024.].