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Bacillus Calmette-Guérin (BCG) vaccination induces a type of immune memory known as "trained immunity", characterized by the immunometabolic and epigenetic changes in innate immune cells. However, the molecular mechanism underlying the strategies for inducing and/or boosting trained immunity in alveolar macrophages remains unknown. Here, we found that mucosal vaccination with the recombinant strain rBCGPPE27 significantly augmented the trained immune response in mice, facilitating a superior protective response against Mycobacterium tuberculosis and non-related bacterial reinfection in mice when compared to BCG. Mucosal immunization with rBCGPPE27 enhanced innate cytokine production by alveolar macrophages associated with promoted glycolytic metabolism, typical of trained immunity. Deficiency of the mammalian target of rapamycin complex 2 and hexokinase 1 abolished the immunometabolic and epigenetic rewiring in mouse alveolar macrophages after mucosal rBCGPPE27 vaccination. Most noteworthy, utilizing rBCGPPE27's higher-up trained effects: The single mucosal immunization with rBCGPPE27-adjuvanted coronavirus disease (CoV-2) vaccine raised the rapid development of virus-specific immunoglobulin G antibodies, boosted pseudovirus neutralizing antibodies, and augmented T helper type 1-biased cytokine release by vaccine-specific T cells, compared to BCG/CoV-2 vaccine. These findings revealed that mucosal recombinant BCG vaccine induces lung-resident memory macrophages and enhances trained immunity via reprogramming mTORC2- and HK-1-mediated aerobic glycolysis, providing new vaccine strategies for improving tuberculosis (TB) or coronavirus variant vaccinations, and targeting innate immunity via mucosal surfaces.
Assuntos
Vacina BCG , Hexoquinase , Memória Imunológica , Pulmão , Macrófagos Alveolares , Alvo Mecanístico do Complexo 2 de Rapamicina , Mycobacterium tuberculosis , Imunidade Treinada , Animais , Camundongos , Vacina BCG/imunologia , Citocinas/metabolismo , Pulmão/imunologia , Macrófagos Alveolares/imunologia , Mycobacterium tuberculosis/imunologia , Vacinas Sintéticas/imunologia , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Hexoquinase/metabolismoRESUMO
Two new austocystin analogs, austocystin P (1) and austocystin Q (2), along with fourteen known compounds (3-16) were isolated from the fermentation extract of Aspergillus sp. WHUF05236. The planar structures of 1 and 2 were elucidated through 1D, 2D NMR and MS analyses. Their absolute configurations were determined by the time-dependent density functional (TDDFT)-ECD calculation. Compounds 3, 11, and 12 exhibited antimicrobial activities against Helicobacter pylori with MIC values ranging from 20.00 to 43.47â µM. Compounds 3, 6, and 7 showed cytotoxicities against the human colon cancer cell lines Hct-116 with IC50 values of 101.79, 65.46, and 36.72â µM, respectively.
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Aspergillus , Fungos , Aspergillus/química , Fungos/química , Humanos , Espectroscopia de Ressonância Magnética , Estrutura MolecularRESUMO
Targeting of Mycobacterium tuberculosis (MTB) PE/PPE antigens that induce type 1 helper T cell (Th1) and Th17 responses represents a crucial strategy for the development of tuberculosis (TB) vaccines. However, only a few PE/PPE antigens induce these responses. Here, we sought to determine how the cell wall-associated antigen PPE60 (Rv3478) activates dendritic cell (DC) maturation and T-cell differentiation. We observed that PPE60 induces DC maturation by augmenting the protein expression of cluster of differentiation 80 (CD80) and CD86 and major histocompatibility complex (MHC) class I and MHC class II on the cell surface. PPE60 also stimulated the production of tumor necrosis factor-α (TNFα), interleukin (IL)-1ß, IL-6, IL-12p70, and IL-23p19 but not IL-10. This induction was mediated by Toll-like receptor 2 (TLR2) and followed by activation of p38, c-Jun N-terminal kinase (JNK), and NF-κB signaling. PPE60 enhanced MHC-II expression and promoted antigen processing by DCs in a TLR2-dependent manner. Moreover, PPE60-stimulated DCs directed naïve CD4+ T cells to produce IFN-γ, IL-2, and IL-17A, expanding the Th1 and Th17 responses, along with activation of T-bet and RAR-related orphan receptor C (RORγt) but not GATA-3. Moreover, PPE60 activated the NLRP3 inflammasome followed by caspase-1-dependent IL-1ß and IL-18 synthesis in DCs. Of note, pharmacological inhibition of NLRP3 activation specifically attenuated IFN-γ and IL-17A secretion into the supernatant from CD4+ T cells cocultured with PPE60-activated DCs. These findings indicate that PPE60 induces Th1 and Th17 immune responses by activating DCs in a TLR2-dependent manner, suggesting PPE60's potential for use in MTB vaccine development.
Assuntos
Antígenos de Bactérias/imunologia , Células Dendríticas/imunologia , Mycobacterium tuberculosis/imunologia , Células Th1/imunologia , Células Th17/imunologia , Receptor 2 Toll-Like/metabolismo , Animais , Antígenos de Bactérias/metabolismo , Proteínas de Bactérias/imunologia , Proteínas de Bactérias/metabolismo , Diferenciação Celular/imunologia , Células Dendríticas/citologia , Feminino , Regulação da Expressão Gênica/imunologia , Antígenos de Histocompatibilidade Classe II/metabolismo , Inflamassomos/metabolismo , Ligantes , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NF-kappa B/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Transdução de Sinais/imunologia , Células Th1/citologia , Células Th17/citologiaRESUMO
The discovery of promising cytokines and clarification of their immunological mechanisms in controlling the intracellular fate of Mycobacterium tuberculosis (Mtb) are necessary to identify effective diagnostic biomarkers and therapeutic targets. To escape immune clearance, Mtb can manipulate and inhibit the normal host process of phagosome maturation. Phagosome maturation arrest by Mtb involves multiple effectors and much remains unknown about this important aspect of Mtb pathogenesis. In this study, we found that interleukin 16 (IL-16) is elevated in the serum samples of Tuberculosis (TB) patients and can serve as a specific target for treatment TB. There was a significant difference in IL-16 levels among active TB, latent TB infection (LTBI), and non-TB patients. This study first revealed that macrophages are the major source of IL-16 production in response to Mtb infection, and elucidated that IL-16 can promote Mtb intracellular survival by inhibiting phagosome maturation and suppressing the expression of Rev-erbα which can inhibit IL-10 secretion. The experiments using zebrafish larvae infected with M. marinum and mice challenged with H37Rv demonstrated that reducing IL-16 levels resulted in less severe pathology and improved survival, respectively. In conclusion, this study provided direct evidence that Mtb hijacks the host macrophages-derived interleukin 16 to enhance intracellular growth. It is suggesting the immunosuppressive role of IL-16 during Mtb infection, supporting IL-16 as a promising therapeutic target.
Assuntos
Interleucina-16 , Mycobacterium tuberculosis , Tuberculose , Animais , Humanos , Camundongos , Interleucina-16/metabolismo , Macrófagos/microbiologia , Mycobacterium tuberculosis/fisiologia , Fagossomos/metabolismo , Fagossomos/microbiologia , Tuberculose/microbiologia , Peixe-ZebraRESUMO
AIM: To elucidate the molecular mechanisms underlying the immunosuppressive effects of emodin isolated from Rheum palmatum L. METHODS: Human T cells were isolated from the peripheral venous blood of 10 healthy adult donors. Cell viability was analyzed with MTT assay. AO/EB and Annexin V/PI staining and DNA damage assay were used to detect cell apoptosis. Fluorescence staining was used to detect the levels of ROS, the mitochondrial membrane potential and intracellular Ca(2+). Colorimetry was used to detect the levels of MDA and total SOD and GSH/GSSG ratio. The expression and activity of caspase-3, -4, and -9 were detected with Western blotting and a fluorometric assay. Western blotting was also used to detect the expression of Bcl-2, Bax, cytochrome C, and endoplasmic reticulum (ER) markers. RESULTS: Emodin (1, 10, and 100 µmol/L) inhibited the growth of human T cells and induced apoptosis in dose- and time dependent manners. Emodin triggered ER stress and significantly elevated intracellular free Ca(2+) in human T cells. It also disrupted mitochondrial membrane potential, and increased cytosolic level of cytochrome C, and the levels of activated cleavage fragments of caspase-3, -4, and -9 in human T cells. Furthermore, emodin significantly increased the levels of ROS and MDA, inhibited both SOD level and GSH/GSSG ratio in human T cells, whereas co-incubation with the ROS scavenger N-acetylcysteine (NAC, 20 µmol/L) almost completely blocked emodin-induced ER stress and mitochondrial dysfunction in human T cells, and decreased the caspase cascade-mediated apoptosis. CONCLUSION: Emodin exerts immunosuppressive actions at least partly by inducing apoptosis of human T cells, which is triggered by ROS-mediated ER stress and mitochondrial dysfunction.
Assuntos
Apoptose/efeitos dos fármacos , Emodina/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Linfócitos T/efeitos dos fármacos , Apoptose/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Estresse do Retículo Endoplasmático/fisiologia , Humanos , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/fisiologia , Mitocôndrias/fisiologia , Linfócitos T/fisiologiaRESUMO
Metabolites of microorganisms have long been considered as potential sources for drug discovery. In this study, five new depsidone derivatives, talaronins A-E (1-5) and three new xanthone derivatives, talaronins F-H (6-8), together with 16 known compounds (9-24), were isolated from the ethyl acetate extract of the mangrove-derived fungus Talaromyces species WHUF0362. The structures were elucidated by analysis of spectroscopic data and chemical methods including alkaline hydrolysis and Mosher's method. Compounds 1 and 2 each attached a dimethyl acetal group at the aromatic ring. A putative biogenetic relationship of the isolated metabolites was presented and suggested that the depsidones and the xanthones probably had the same biosynthetic precursors such as chrysophanol or rheochrysidin. The antimicrobial activity assay indicated that compounds 5, 9, 10, and 14 showed potent activity against Helicobacter pylori with minimum inhibitory concentration (MIC) values in the range of 2.42-36.04 µmol/L. While secalonic acid D (19) demonstrated significant antimicrobial activity against four strains of H. pylori with MIC values in the range of 0.20 to 1.57 µmol/L. Furthermore, secalonic acid D (19) exhibited cytotoxicity against cancer cell lines Bel-7402 and HCT-116 with IC50 values of 0.15 and 0.19 µmol/L, respectively. The structure-activity relationship of depsidone derivatives revealed that the presence of the lactone ring and the hydroxyl at C-10 was crucial to the antimicrobial activity against H. pylori. The depsidone derivatives are promising leads to inhibit H. pylori and provide an avenue for further development of novel antibiotics. Supplementary Information: The online version contains supplementary material available at 10.1007/s42995-023-00170-5.
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With conserved structural scaffold and divergent electrophysiological functions, animal toxins are considered powerful tools for investigating the basic structure-function relationship of voltage-gated sodium channels. Jingzhaotoxin-III (ß-TRTX-Cj1α) is a unique sodium channel gating modifier from the tarantula Chilobrachys jingzhao, because the toxin can selectively inhibit the activation of cardiac sodium channel but not neuronal subtypes. However, the molecular basis of JZTX-III interaction with sodium channels remains unknown. In this study, we showed that JZTX-III was efficiently expressed by the secretory pathway in yeast. Alanine-scanning analysis indicated that 2 acidic residues (Asp1, Glu3) and an exposed hydrophobic patch, formed by 4 Trp residues (residues 8, 9, 28 and 30), play important roles in the binding of JZTX-III to Nav1.5. JZTX-III docked to the Nav1.5 DIIS3-S4 linker. Mutations S799A, R800A, and L804A could additively reduce toxin sensitivity of Nav1.5. We also demonstrated that the unique Arg800, not emerging in other sodium channel subtypes, is responsible for JZTX-III selectively interacting with Nav1.5. The reverse mutation D816R in Nav1.7 greatly increased the sensitivity of the neuronal subtype to JZTX-III. Conversely, the mutation R800D in Nav1.5 decreased JZTX-III's IC50 by 72-fold. Therefore, our results indicated that JZTX-III is a site 4 toxin, but does not possess the same critical residues on sodium channels as other site 4 toxins. Our data also revealed the underlying mechanism for JZTX-III to be highly specific for the cardiac sodium channel.
Assuntos
Peptídeos/toxicidade , Canais de Sódio/metabolismo , Venenos de Aranha/toxicidade , Aranhas/fisiologia , Substituição de Aminoácidos , Animais , Regulação da Expressão Gênica , Células HEK293 , Humanos , Modelos Moleculares , Mutagênese Sítio-Dirigida , Mutação , Canal de Sódio Disparado por Voltagem NAV1.5 , Técnicas de Patch-Clamp , Peptídeos/química , Peptídeos/genética , Conformação Proteica , Canais de Sódio/genética , Venenos de Aranha/química , Venenos de Aranha/genética , Relação Estrutura-AtividadeRESUMO
Ischemic stroke has been reported to cause significant changes to memory, thinking, and behavior. Intriguingly, recently reported studies have indicated the association of Trimethylamine N-oxide (TMAO) with the acute phase of ischemic stroke. However, the comprehensive underlying mechanism remained unknown. The objective of the present study was to investigate the association between TMAO and recovery of neurological function after ischemic stroke. For this purpose, a middle cerebral artery occlusion/reperfusion (MCAO/R) rat model was established and treated with TMAO or/and sh-ALK5, followed by the neurological function evaluation. Behaviors of rats were observed through staircase and cylinder tests. Moreover, the expression of Smurf2 and ALK5 was detected by immunohistochemistry while expression of GFAP, Neurocan, and Phosphacan in brain tissues was determined by immunofluorescence. Thereafter, gain- and loss-of-function assays in astrocytes, the proliferation, viability, and migration were evaluated by the EdU, CCK-8, and Transwell assays. Besides, Smurf2 mRNA expression was determined by the RT-qPCR, whereas, Smurf2, ALK5, GFAP, Neurocan, and Phosphacan expression was evaluated by the Western blotting. Finally, the interaction of Smurf2 with ALK5 and ALK5 ubiquitination was assessed by the co-immunoprecipitation. Notably, our results showed that TMAO promoted the proliferation of reactive astrocyte and formation of glial scar in MCAO/R rats. However, this effect was abolished by the Smurf2 overexpression or ALK5 silencing. We further found that TMAO upregulated the ALK5 expression by inhibiting the ubiquitination role of Smurf2. Overexpression of ALK5 reversed the inhibitory effect of Smurf2 on astrocyte proliferation, migration, and viability. Collectively, our work identifies the evolutionarily TMAO/Smurf2/ALK5 signaling as a major genetic factor in the control of reactive astrocyte proliferation and glial scar formation in ischemic stroke, thus laying a theoretical foundation for the identification of ischemic stroke.
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Trained monocytes and macrophages produce reactive oxygen species (ROS), which trigger antioxidative glutathione (GSH) response to buffer the rising ROS. However, whether and how the trained immunity is shaped by GSH synthesis remains unknown. Here, we report that ß-glucan-trained macrophages from mice harboring a myeloid-specific deletion of the catalytic subunit of glutamate-cysteine ligase (Gclc) showed impaired GSH synthesis and decreased proinflammatory cytokine production in response to lipopolysaccharide challenge. Gclc deficiency compromised the activation of mammalian target of rapamycin-1 (mTOR) and expression of c-Myc transcription factors, abrogating the energy utilization and the metabolic reprogramming that allows ß-glucan-trained macrophages to switch to glycolysis and glutaminolysis. Furthermore, Gclc deletion repressed effective H3K27me3 demethylation in the promoters of immunometabolic genes, such as Gls, Hk2, and Glut1, in ß-glucan-trained macrophages by promoting the methyltransferase enhancer of zeste homolog 2 (EZH2). In vivo, myeloid-specific ablation of Gclc decreased the secretion of proinflammatory cytokines upon rechallenge with Candida albicans and these animals were less protected against the infection, compared with control littermates. Moreover, pharmacological inhibition of EZH2 enhanced the trained immunity response against Candida infection in Gclc-deficient mouse and human peripheral blood mononuclear cells treated with GCLC inhibitor buthionine sulfoximine (BSO). Thus, antioxidative GSH synthesis supports an environment conducive to ß-glucan-induced metabolic and epigenetic reprogramming in trained immunity, allowing exploration of its functional consequences in autoimmune or inflammatory disease.
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Xiaoyaosan is one of the famous formulas treating for liver, spleen and blood deficiency syndrome along Chinese history. Their five main components, paeoniflorin, ferulic acid, glycyrrhizic acid, liquiritin, and atractylenolide I are believed to partly representative of this formula. Ultra performance liquid chromatography method was newly established, UPLC BEH-C18 column (2.1 × 100 mm, 1.7 µm) with acetonitrile - 0.1% phosphate acid gradient elution system, in 0.4 mL/min and the temperature was 30 °C; Detection wavelengths were optimized separately. The five components were linear within their linear range (r ≥ 0.9991), average recovery 97.63%â¼102.83%, RSD 1.88%â¼4.38%. The newly established method is accurate, rapid, and convenient, with satisfied separation performance to quantify multi-components in formula and preparations of Xiaoyaosan. It will provide a reliable reference for the quality evaluation of both Xiaoyaosan formula and preparations during the process of manufacture, administration and clinical application in the future.
Assuntos
Medicamentos de Ervas Chinesas/química , Compostos Fitoquímicos/análise , Calibragem , Ácidos Cumáricos/análise , Ácidos Cumáricos/química , Flavanonas/análise , Flavanonas/química , Glucosídeos/análise , Glucosídeos/química , Ácido Glicirrízico/análise , Ácido Glicirrízico/química , Lactonas/análise , Lactonas/química , Monoterpenos/análise , Monoterpenos/química , Compostos Fitoquímicos/química , Reprodutibilidade dos Testes , Sesquiterpenos/análise , Sesquiterpenos/químicaRESUMO
BACKGROUND: Acute lymphoblastic leukaemia (ALL) is often characterized by broad clinical and biological heterogeneity, as well as recurrent genetic aberrations. Despite remarkable improvements in the treatment outcome in paediatric ALL over the past several decades, it remains a leading cause of morbidity and mortality among children. Cytokines have been extensively studied in haematologic diseases; however, the mechanisms by which cytokines contribute to ALL pathogenesis remain poorly understood. METHODS: IL-33 levels were measured by enzyme-linked immunosorbent assay (ELISA). IL1RL1 expression on ALL cell surface was accessed by flow cytometry. Expression of phosphorylated p38 MAPK, p38, pAKT, AKT and GAPDH were quantified by western blot. Cell survival signals were evaluated by apoptosis using flow cytometry. RESULTS: BM samples from ALL patients at diagnosis upregulated their cell surface expression of IL1RL1, and a higher interleukin (IL)-33 level in the serum was observed as compared to the healthy individuals. Moreover, exogenous IL-33 treatment significantly inhibited apoptosis by activating p38 mitogen-activated protein kinase (MAPK) and AKT pathway, while the inhibitor for p38 MAPK, SB203580, counteracted IL-33-induced anti-apoptosis via inactivation of p38 MAPK and AKT. Furthermore, IL-33 negatively regulates cyclin B1 protein level while increasing the expression of CDK1, with SB203580 inhibiting the effect. CONCLUSION: Our study reveals an important role for IL-33/IL1RL1 axis in supporting ALL which may represent a novel treatment for paediatric patients.KEY MESSAGESBoth IL-33 and IL1RL1 levels are upregulated in primary ALL samples.IL-33 increased both p38 MAPK and AKT activation in ALL.IL-33 promotes survival and cell cycle progression of ALL cells via activating p38 MAPK.
Assuntos
Apoptose/efeitos dos fármacos , Inflamação/metabolismo , Interleucina-33/farmacologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Proteínas Proto-Oncogênicas c-akt , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Criança , Ensaio de Imunoadsorção Enzimática , Humanos , Interleucina-33/sangue , Recidiva Local de Neoplasia , Fosforilação , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismoRESUMO
Trained immunity, induced by ß-glucan in monocytes, is mediated by activating metabolic pathways that result in epigenetic rewiring of cellular functional programs; however, molecular mechanisms underlying these changes remain unclear. Here, we report a key immunometabolic and epigenetic pathway mediated by the miR-9-5p-isocitrate dehydrogenase 3α (IDH3α) axis in trained immunity. We found that ß-glucan-trained miR-9-5p-/- monocytes showed decreased IL-1ß, IL-6, and TNF-α production after LPS stimulation. Trained miR-9-5p-/- mice produced decreased levels of proinflammatory cytokines upon rechallenge in vivo and had worse protection against Candida albicans infection. miR-9-5p targeted IDH3α and reduced α-ketoglutarate (α-KG) levels to stabilize HIF-1α, which promoted glycolysis. Accumulating succinate and fumarate via miR-9-5p action integrated immunometabolic circuits to induce histone modifications by inhibiting KDM5 demethylases. ß-Glucan-trained monocytes exhibited low IDH3α levels, and IDH3α overexpression blocked the induction of trained immunity by monocytes. Monocytes with IDH3α variants from autosomal recessive retinitis pigmentosa patients showed a trained immunity phenotype at immunometabolic and epigenetic levels. These findings suggest that miR-9-5p and IDH3α act as critical metabolic and epigenetic switches in trained immunity.
Assuntos
Epigênese Genética/genética , Imunidade Inata/genética , Memória Imunológica/genética , Isocitrato Desidrogenase/metabolismo , Redes e Vias Metabólicas/genética , MicroRNAs/genética , Monócitos/metabolismo , Animais , Candida albicans , Candidíase/genética , Candidíase/imunologia , Epigênese Genética/imunologia , Fumaratos/metabolismo , Glicólise/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Imunidade Inata/imunologia , Memória Imunológica/imunologia , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Ácidos Cetoglutáricos/metabolismo , Lipopolissacarídeos/farmacologia , Redes e Vias Metabólicas/imunologia , Camundongos , Camundongos Knockout , MicroRNAs/metabolismo , Monócitos/efeitos dos fármacos , Retinose Pigmentar/genética , Retinose Pigmentar/metabolismo , Ácido Succínico/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , beta-Glucanas/imunologiaRESUMO
Breath volatile biomarkers are capable of distinguishing patients with various cancers. However, high throughput analytical technology is a prerequisite to a large-cohort study intended to discover reliable breath biomarkers for cancer diagnosis. Single-photon ionization (SPI) is a universal ionization technology, and SPI-mass spectrometry (SPI-MS) shows a remarkable advantage in the comprehensive detection of volatile organic compounds (VOCs), in particular, nonpolar compounds. In this study, we have introduced SPI-MS coupled with on-line thermal desorption (TD-SPI-MS) to demonstrate nontarget analysis of breath VOCs for gastric cancer patients. The breath fingerprints of the gastric cancer patients were significantly distinct from that of the control group. Acetone, isoprene, 1,3-dioxolan-2-one, phenol, meta-xylene, 1,2,3-trimethylbenzene, and phenyl acetate showed higher relative peak intensities in the breath profiles of gastric cancer patients. A diagnostic prediction model was further developed by using a training set (121 samples) and validated with a test set (53 samples). The predication accuracy of the developed model was 96.2%, and the area under the curve (AUC) of the receiver operator characteristic curve (ROC) was 0.997, indicating a satisfactory prediction ability of the developed model. Thus, by taking gastric cancer as an example, we have shown that TD-SPI-MS will be a promising tool for high throughput analysis of breath samples to discover characteristic VOCs in patients with various cancers.
Assuntos
Biomarcadores Tumorais/análise , Testes Respiratórios/métodos , Espectrometria de Massas/métodos , Neoplasias Gástricas/diagnóstico , Compostos Orgânicos Voláteis/análise , Acetatos/análise , Acetona/análise , Butadienos/análise , Estudos de Coortes , Dioxolanos/análise , Expiração , Hemiterpenos/análise , Humanos , Fenol/análise , Fenóis/análise , Curva ROC , Xilenos/análiseRESUMO
Acute myeloid leukemia (AML) is a fatal disease characterized by the accumulation of immature myeloid blasts in the bone marrow (BM). Cytokine provide signals for leukemia cells to improve their survival in the BM microenvironment. Previously, we identified interleukin-33 (IL-33) as a promoter of cell survival in a human AML cell line and primary mouse leukemia cells. In this study, we report that the cell surface expression of IL-33-specific receptor, Interleukin 1 Receptor Like 1 (IL1RL1), is elevated in BM cells from AML patients at diagnosis, and the serum level of IL-33 in AML patients is higher than that of healthy donor controls. Moreover, IL-33 levels are found to be positively associated with IL-6 levels in pediatric patients with AML. In vitro, IL-33 treatment increased IL-6 mRNA expression and protein level in BM and peripheral blood (PB) cells from AML patients. Evidence was also provided that IL-33 inhibits cell apoptosis by activating p38 mitogen-activated protein kinase (MAPK) pathway using human AML cell line and AML patient samples. Finally, we confirmed that IL-33 activated IL-6 expression in a manner that required p38 MAPK pathway using clinical AML samples. Taken together, we identified a potential mechanism of IL-33-mediated survival involving p38 MAPK in pediatric AML patients that would facilitate future drug development.
Assuntos
Regulação Leucêmica da Expressão Gênica/imunologia , Interleucina-33/imunologia , Interleucina-6/imunologia , Leucemia Mieloide Aguda/imunologia , Proteínas de Neoplasias/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/imunologia , Sobrevivência Celular/imunologia , Criança , Feminino , Humanos , Leucemia Mieloide Aguda/patologia , MasculinoRESUMO
BACKGROUND: Whether the mesenteric lymphatic system could serve as a route of transport by which gut-derived inflammatory mediators contribute to the induction of remote organ injuries is uncertain. We therefore made a gut-induced lung injury canine model by portal vein occlusion and reperfusion (PV O/R) and studied the role of mesenteric lymphatic ducts ligation (ML) to gut-induced lung injury with this model. MATERIAL AND METHODS: Eighteen mongrel dogs were divided into control, PV O/R, and PV O/R + ML groups. Cytokines and endotoxin levels in the portal vein and lymph from thoracic duct in different groups were tested. The permeability, myeloperoxidase activity, and histopathological investigation of intestine and lung were evaluated. RESULTS: Cytokines and endotoxin levels in the portal vein were significantly increased in experimental groups compared with control group (P < 0.05), and that in the lymph from thoracic duct were significantly increased in PV O/R group compared with control and PV O/R + ML group (P < 0.05). Lung permeability and MPO activity in PV O/R group were significantly higher than those in control and PV O/R + ML group (P < 0.05); intestinal permeability in experimental groups were significantly higher with respect to control group. The lung injury score in PV O/R group was significantly higher than those in control and PV O/R + ML group (P < 0.05) and the intestinal injury scores in experimental groups were significantly higher than control group (P < 0.05). CONCLUSIONS: The gut-induced lung injury canine model made by PV O/R is successful, and mesenteric lymphatic ducts ligation decreases the degree of gut-induced lung injury in this model.
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Intestinos/irrigação sanguínea , Lesão Pulmonar/etiologia , Doenças Linfáticas/cirurgia , Veia Porta/cirurgia , Traumatismo por Reperfusão/prevenção & controle , Doenças Vasculares/cirurgia , Animais , Modelos Animais de Doenças , Cães , Intestinos/patologia , Ligadura/métodos , Ligadura/veterinária , Lesão Pulmonar/cirurgia , ReperfusãoRESUMO
The myobacterial factors and the associated mechanism by which Mycobacterium tuberculosis (Mtb) evades the host immune surveillance system remain widely unexplored. Here, we found that overexpressing Rv1016c, a mannosylated protein of M. tuberculosis in BCG (rBCG-Rv1016c) led to increased virulence of the recombined BCG in the severe-combined immunodeficient (SCID) mice model and to a loss of protective efficacy in a zebrafish-M. marinum model, compared to wild type BCG. Further investigations on the effects of rBCG-Rv1016c on the host innate immunity revealed that rBCG-Rv1016c decreased the production of cytokines IL-2, IL-12p70, TGF-ß, IL-6 as well as of the co-stimulatory molecules CD80, CD86, MHC-I and MHC-II by the infected DCs. These effects were mimicked by rBCG-Rv1016cHis, which carried an extra 6-His tag at the C-terminus of Rv1016c. Relatively to BCG infected DCs, the rBCG-Rv1016c-infected DCs failed to polarize naïve T cells to Th1- and Th17-type cells to secret IFN-γ and IL-17. Additionally, T lymphocytes from BCG- infected mice showed significantly less proliferation and production of IFN-γ and IL-17. Similarly, rBCG-Rv1016c mice released a higher level of IL-10 in response to rBCG-Rv1016c stimulation than wild type BCG infected mice. Furthermore, DCs from TLR-2 knockout mice showed no reduction in IL-6, IL-12 p70 and TGF-ß secretion in response to rBCG-Rv1016c infection, compared to DCs infected with BCG. We propose that Rv1016c interferes in differentiation of the DCs by targeting suppressor of cytokine signaling (SOCS) 1 and SOCS3 expression, which subsequently leads to the reduction in STAT-1 and STAT-6 phosphorylation. These findings open new perspectives regarding the immunosuppressive strategies adopted by Mtb to survive in the host.
Assuntos
Proteínas de Bactérias/metabolismo , Diferenciação Celular , Células Dendríticas/imunologia , Glicoproteínas/metabolismo , Mycobacterium tuberculosis/fisiologia , Células Th1/imunologia , Células Th17/imunologia , Tuberculose/imunologia , Animais , Polaridade Celular , Citocinas/metabolismo , Células Dendríticas/microbiologia , Feminino , Camundongos , Mycobacterium bovis/patogenicidade , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais , Baço/patologia , Proteína 3 Supressora da Sinalização de Citocinas/metabolismo , Receptor 2 Toll-Like/metabolismo , Tuberculose/microbiologia , Regulação para CimaRESUMO
Targeting of Mycobacterium tuberculosis (MTB) latent antigens comprises a crucial strategy for the development of alternative tuberculosis (TB) vaccine(s) that protects against TB reactivation. Here, we generated a multistage DNA vaccine, A39, containing the early antigens Ag85A and Rv3425 as well as the latency-associated protein Rv2029c, which conferred protective immunity in a pre-exposure mouse model. Moreover, administration of the A39 vaccination after MTB exposure inhibited reactivation and resulted in significantly lower bacterial loads in the lungs and spleen of mice, compared to those in the control population. Subsequently, we investigated the effect of Rv2029c on innate immunity and characterized the molecular details of the interaction of this protein with the host via iTRAQ proteomic and biochemical assay analyses. Rv2029c activated macrophages, triggered the production of pro-inflammatory cytokines, and promoted toll-like receptor/mitogen-activated protein kinase (TLR/MAPK)-dependent macrophage apoptosis. Furthermore, Rv2029c treatment enhanced the ability of Mycobacterium bovis Bacillus Calmette-Guérin (BCG)-infected macrophages to present antigens to CD4+ T cells in vitro, which correlated with an increase in MHC-II expression. Lastly, Rv2029c-treated macrophages activated T cells, effectively polarized CD4+ and CD8+ T cells to secrete IFN-γ and IL-2, and specifically expanded a population of CD44highCD62LlowCD4+/CD8+ effector/memory cells, indicating that Rv2029c, as a specific recall antigen, contributes to Th1 polarization in T cell immunity. These results suggest that Rv2029c and A39 comprise promising targets for the development of next-generation clinical TB therapeutic vaccines.
RESUMO
TLR2-dependent cellular signaling in Mycobacterium tuberculosis-infected macrophages causes apoptosis and inhibits class II major histocompatibility complex (MHC-II) molecules antigen processing, leading to evasion of surveillance. Mycobacterium tuberculosis (MTB) lipoproteins are an important class of Toll-like receptor (TLR) ligand, and identified as specific components that mediate these effects. In this study, we identified and characterized MTB lipoprotein Rv1016c (lpqT) as a cell wall associated-protein that was exposed on the cell surface and enhanced the survival of recombinants M. smegmatis_Rv1016c under stress conditions. We found that Rv1016c lipoprotein was a novel TLR2 ligand and able to induce macrophage apoptosis in a both dose- and time-dependent manner. Additionally, apoptosis induced by Rv1016c was reserved in THP-1 cells blocked with anti-TLR-2 Abs or in TLR2-/- mouse macrophages, indicating that Rv1016c-induced apoptosis is dependent on TLR2. Moreover, we demonstrated that Rv1016c lipoprotein inhibited IFN-γ-induced MHC-II expression and processing of soluble antigens in a TLR2 dependent manner. Class II transactivator (CIITA) regulates MHC II expression. In this context, Rv1016c lipoprotein diminished IFN-γ-induced expression of CIITA IV through TLR2 and MAPK Signaling. TLR2-dependent apoptosis and inhibition of MHC-II Ag processing induced by Rv1016c during mycobacteria infection may promote the release of residual bacilli from apoptotic cells and decrease recognition by CD4+ T cells. These mechanisms may allow intracellular MTB to evade immune surveillance and maintain chronic infection.
Assuntos
Apresentação de Antígeno/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Proteínas de Bactérias/farmacologia , Antígenos de Histocompatibilidade Classe II/efeitos dos fármacos , Lipoproteínas/genética , Macrófagos/microbiologia , Mycobacterium tuberculosis/metabolismo , Proteínas Recombinantes/farmacologia , Receptor 2 Toll-Like/metabolismo , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Linhagem Celular , Parede Celular/química , DNA Bacteriano , Modelos Animais de Doenças , Regulação da Expressão Gênica , Genes Bacterianos/genética , Humanos , Interferon gama/efeitos dos fármacos , Lipoproteínas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mycobacterium smegmatis , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/imunologia , Proteínas Nucleares/efeitos dos fármacos , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Receptores de Superfície Celular/fisiologia , Proteínas Recombinantes/genética , Transdução de Sinais/efeitos dos fármacos , Análise de Sobrevida , Receptor 2 Toll-Like/genética , Receptores Toll-Like/classificação , Receptores Toll-Like/metabolismo , Transativadores/efeitos dos fármacos , Tuberculose/microbiologiaRESUMO
In this study, a flexible asymmetrical all-solid-state supercapacitor with high electrochemical performance was fabricated with Ni/MnO2-filter paper (FP) as the positive electrode and Ni/active carbon (AC)-filter paper as negative electrode, separated with poly(vinyl alcohol) (PVA)-Na2SO4 electrolyte. A simple procedure, such as electroless plating, was introduced to prepare the Ni/MnO2-FP electrode on the conventional laboratory FP, combined with the subsequent step of electrodeposition. Electrochemical results show that the as-prepared electrodes display outstanding areal specific capacitance (1900 mF/cm(2) at 5 mV/s) and excellent cycling performance (85.1% retention after 1000 cycles at 20 mA/cm(2)). Such a flexible supercapacitor assembled asymmetrically in the solid state exhibits a large volume energy density (0.78 mWh/cm(3)) and superior flexibility under different bending conditions. It has been demonstrated that the supercapacitors could be used as a power source to drive a 3 V light-emitting diode indicator. This study may provide an available method for designing and fabricating flexible supercapacitors with high performance in the application of wearable and portable electronics based on easily available materials.