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1.
Commun Biol ; 7(1): 817, 2024 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-38965342

RESUMO

Macrophages play a pivotal role in orchestrating the immune response against pathogens. While the intricate interplay between macrophage activation and metabolism remains a subject of intense investigation, the role of glutamate oxaloacetate transaminase 1 (Got1) in this context has not been extensively assessed. Here, we investigate the impact of Got1 on macrophage polarization and function, shedding light on its role in reactive oxygen species (ROS) production, pathogen defense, and immune paralysis. Using genetically modified mouse models, including both myeloid specific knockout and overexpression, we comprehensively demonstrate that Got1 depletion leads to reduced ROS production in macrophages. Intriguingly, this impairment in ROS generation does not affect the resistance of Got1 KO mice to pathogenic challenges. Furthermore, Got1 is dispensable for M2 macrophage differentiation and does not influence the onset of LPS-induced immune paralysis. Our findings underscore the intricate facets of macrophage responses, suggesting that Got1 is dispensable in discrete immunological processes.


Assuntos
Diferenciação Celular , Macrófagos , Camundongos Knockout , Espécies Reativas de Oxigênio , Animais , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Espécies Reativas de Oxigênio/metabolismo , Camundongos Endogâmicos C57BL , Aspartato Aminotransferase Citoplasmática/genética , Aspartato Aminotransferase Citoplasmática/metabolismo , Ativação de Macrófagos/genética , Lipopolissacarídeos/farmacologia
2.
J Leukoc Biol ; 2024 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-38973642

RESUMO

HHLA2, a member of the B7 family of co-signaling molecules, is aberrantly expressed in various human cancers and has emerged as a promising target for cancer immunotherapy. It exhibits a unique structure and tissue distribution pattern compared to other B7 family members, where its expression is regulated by the complex physiological and tumor microenvironment. HHLA2 plays a crucial but contradictory role in immune modulation, and is thereby associated with heterogeneous prognostic implications across different cancer types. It interacts with two distinct receptors: TMIGD2, which is predominantly expressed on naïve T and NK cells to deliver co-stimulatory signals to T cells and NK cells; and KIR3DL3, which is prevalent on terminally differentiated T and CD56dim CD16+ NK cells to transmit inhibitory signals. The expression dynamics of these receptors on immune cells contribute to the maintenance of immune response homeostasis. Therapeutic strategies targeting the HHLA2 immune checkpoint aim to selectively inhibit the immunosuppressive HHLA2-KIR3DL3 pathway while preserving the HHLA2-TMIGD2 signaling. Several anti-HHLA2 and anti-KIR3DL3 antibodies are currently under investigation in early clinical trials, building upon encouraging results observed in humanized mouse models. Notably, the non-overlapping expression of HHLA2 and PD-L1 in tumors suggests potential synergistic benefits of combining HHLA2-KIR3DL3 targeted therapies with PD-1/PD-L1 blockade or anti-CTLA-4 to augment antitumor activity.

3.
Commun Biol ; 7(1): 281, 2024 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-38448655

RESUMO

Rosamine-based mitochondrial dyes, such as Mitotracker Red, have commonly been employed to visualize mitochondrial localization within cells due to their preferential accumulation in organelles with membrane potential. Consequently, Mitotracker Red has often served as a surrogate indicator for tracking mitochondrial movement between neighboring cells. However, it is important to note that the presence of membrane potential in the cell membrane and other organelles may lead to the non-specific partial enrichment of Mitotracker Red in locations other than mitochondria. This study comprehensively investigates the reliability of mitochondrial dye as a marker for studying horizontal mitochondrial transfer (HMT). By meticulous replicating of previous experiments and comparing the efficiency of mitochondrial dye transfer with that of mito-targeted GFP, our findings confirm that HMT occurs at significantly lower efficiency than previously indicated by Mitotracker dye. Subsequent experiments involving mitochondria-deficient cells robustly demonstrates the non-specificity of mitochondrial dye as indicator for mitochondria. We advocate for a thorough reevaluation of existing literature in this field and propose exploration of alternative techniques to enhance the investigation of HMT. By addressing these pivotal aspects, we can advance our understanding of cellular dynamics and pave the way for future explorations in this captivating field.


Assuntos
Corantes , Mitocôndrias , Reprodutibilidade dos Testes , Membrana Celular , Potenciais da Membrana
4.
Nat Commun ; 14(1): 6639, 2023 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-37863917

RESUMO

Type 1 conventional dendritic cells (cDC1) are the most efficient cross-presenting cells that induce protective cytotoxic T cell response. However, the regulation of their homeostasis and function is incompletely understood. Here we observe a selective reduction of splenic cDC1 accompanied by excessive cell death in mice with Zeb1 deficiency in dendritic cells, rendering the mice more resistant to Listeria infection. Additionally, cDC1 from other sources of Zeb1-deficient mice display impaired cross-presentation of exogenous antigens, compromising antitumor CD8+ T cell responses. Mechanistically, Zeb1 represses the expression of microRNA-96/182 that target Cybb mRNA of NADPH oxidase Nox2, and consequently facilitates reactive-oxygen-species-dependent rupture of phagosomal membrane to allow antigen export to the cytosol. Cybb re-expression in Zeb1-deficient cDC1 fully restores the defective cross-presentation while microRNA-96/182 overexpression in Zeb1-sufficient cDC1 inhibits cross-presentation. Therefore, our results identify a Zeb1-microRNA-96/182-Cybb pathway that controls cross-presentation in cDC1 and uncover an essential role of Zeb1 in cDC1 homeostasis.


Assuntos
MicroRNAs , Fatores de Transcrição , Animais , Camundongos , Antígenos/metabolismo , Linfócitos T CD8-Positivos , Células Dendríticas , Homeostase , MicroRNAs/genética , MicroRNAs/metabolismo , Fatores de Transcrição/metabolismo
5.
Nat Commun ; 14(1): 4117, 2023 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-37433761

RESUMO

The emergence of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) variants and "anatomical escape" characteristics threaten the effectiveness of current coronavirus disease 2019 (COVID-19) vaccines. There is an urgent need to understand the immunological mechanism of broad-spectrum respiratory tract protection to guide broader vaccines development. Here we investigate immune responses induced by an NS1-deleted influenza virus vectored intranasal COVID-19 vaccine (dNS1-RBD) which provides broad-spectrum protection against SARS-CoV-2 variants in hamsters. Intranasal delivery of dNS1-RBD induces innate immunity, trained immunity and tissue-resident memory T cells covering the upper and lower respiratory tract. It restrains the inflammatory response by suppressing early phase viral load post SARS-CoV-2 challenge and attenuating pro-inflammatory cytokine (Il6, Il1b, and Ifng) levels, thereby reducing excess immune-induced tissue injury compared with the control group. By inducing local cellular immunity and trained immunity, intranasal delivery of NS1-deleted influenza virus vectored vaccine represents a broad-spectrum COVID-19 vaccine strategy to reduce disease burden.


Assuntos
COVID-19 , Vacinas contra Influenza , Influenza Humana , Animais , Cricetinae , Humanos , Vacinas contra COVID-19 , SARS-CoV-2 , COVID-19/prevenção & controle
6.
Cell Rep ; 42(5): 112471, 2023 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-37149865

RESUMO

T helper type 2 (Th2) cytokine-activated M2 macrophages contribute to inflammation resolution and wound healing. This study shows that IL-4-primed macrophages exhibit a stronger response to lipopolysaccharide stimulation while maintaining M2 signature gene expression. Metabolic divergence between canonical M2 and non-canonical proinflammatory-prone M2 (M2INF) macrophages occurs after the IL-4Rα/Stat6 axis. Glycolysis supports Hif-1α stabilization and proinflammatory phenotype of M2INF macrophages. Inhibiting glycolysis blunts Hif-1α accumulation and M2INF phenotype. Wdr5-dependent H3K4me3 mediates the long-lasting effect of IL-4, with Wdr5 knockdown inhibiting M2INF macrophages. Our results also show that the induction of M2INF macrophages by IL-4 intraperitoneal injection and transferring of M2INF macrophages confer a survival advantage against bacterial infection in vivo. In conclusion, our findings highlight the previously neglected non-canonical role of M2INF macrophages and broaden our understanding of IL-4-mediated physiological changes. These results have immediate implications for how Th2-skewed infections could redirect disease progression in response to pathogen infection.


Assuntos
Interleucina-4 , Macrófagos , Humanos , Interleucina-4/farmacologia , Interleucina-4/metabolismo , Macrófagos/metabolismo , Inflamação/metabolismo , Citocinas/metabolismo , Glicólise/fisiologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo
8.
Oncogene ; 40(30): 4847-4858, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34155349

RESUMO

Small cell lung cancer (SCLC) continues to cause poor clinical outcomes due to limited advances in sustained treatments for rapid cancer cell proliferation and progression. The transcriptional factor Forkhead Box M1 (FOXM1) regulates cell proliferation, tumor initiation, and progression in multiple cancer types. However, its biological function and clinical significance in SCLC remain unestablished. Analysis of the Cancer Cell Line Encyclopedia and SCLC datasets in the present study disclosed significant upregulation of FOXM1 mRNA in SCLC cell lines and tissues. Gene set enrichment analysis (GSEA) revealed that FOXM1 is positively correlated with pathways regulating cell proliferation and DNA damage repair, as evident from sensitization of FOXM1-depleted SCLC cells to chemotherapy. Furthermore, Foxm1 knockout inhibited SCLC formation in the Rb1fl/flTrp53fl/flMycLSL/LSL (RPM) mouse model associated with increased levels of neuroendocrine markers, Ascl1 and Cgrp, and decrease in Yap1. Consistently, FOXM1 depletion in NCI-H1688 SCLC cells reduced migration and enhanced apoptosis and sensitivity to cisplatin and etoposide. SCLC with high FOXM1 expression (N = 30, 57.7%) was significantly correlated with advanced clinical stage, extrathoracic metastases, and decrease in overall survival (OS), compared with the low-FOXM1 group (7.90 vs. 12.46 months). Moreover, the high-FOXM1 group showed shorter progression-free survival after standard chemotherapy, compared with the low-FOXM1 group (3.90 vs. 8.69 months). Our collective findings support the utility of FOXM1 as a prognostic biomarker and potential molecular target for SCLC.


Assuntos
Biomarcadores Tumorais , Proteína Forkhead Box M1/genética , Neoplasias Pulmonares/etiologia , Neoplasias Pulmonares/mortalidade , Carcinoma de Pequenas Células do Pulmão/etiologia , Carcinoma de Pequenas Células do Pulmão/mortalidade , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Linhagem Celular Tumoral , Proliferação de Células , Modelos Animais de Doenças , Feminino , Proteína Forkhead Box M1/metabolismo , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/diagnóstico , Masculino , Camundongos , Camundongos Transgênicos , Pessoa de Meia-Idade , Gradação de Tumores , Estadiamento de Neoplasias , Prognóstico , Carcinoma de Pequenas Células do Pulmão/diagnóstico , Microtomografia por Raio-X , Ensaios Antitumorais Modelo de Xenoenxerto
9.
Sci Rep ; 9(1): 12900, 2019 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-31501504

RESUMO

Sleep apnea has been associated with a variety of diseases, but its impact on sepsis outcome remains unclear. This study investigated the effect of intermittent hypoxia [IH]-the principal feature of sleep apnea-on murine sepsis. 5-week-old male C57BL6 mice were assigned to groups receiving severe IH (O2 fluctuating from room air to an O2 nadir of 5.7% with a cycle length of 90 seconds), mild IH (room air to 12%, 4 minutes/cycle), or room air for 3 weeks. Sepsis was induced by cecal ligation and puncture and survival was monitored. Sepsis severity was evaluated by murine sepsis scores, blood bacterial load, plasma tumor necrosis factor-α [TNF-α]/interleukin-6 [IL-6] levels and histopathology of vital organs. Compared with normoxic controls, mice subjected to severe IH had earlier mortality, a lower leukocyte count, higher blood bacterial load, higher plasma TNF-α and IL-6 levels, more severe inflammatory changes in the lung, spleen and small intestine. Mice subjected to mild IH did not differ from normoxic controls, except a higher IL-6 level after sepsis induced. The adverse impact of severe IH was reversed following a 10-day normoxic recovery. In conclusion, severe IH, not mild IH, contributed to poorer outcomes in a murine sepsis model.


Assuntos
Hipóxia/metabolismo , Sepse/metabolismo , Sepse/mortalidade , Animais , Biomarcadores , Biópsia , Modelos Animais de Doenças , Mediadores da Inflamação , Masculino , Camundongos , Prognóstico , Sepse/diagnóstico , Sepse/etiologia
10.
Front Immunol ; 9: 526, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29593748

RESUMO

High-mobility group nucleosome-binding protein 1 (HMGN1) functions as a non-histone chromatin-binding protein in the cell nucleus. However, extracellular HMGN1 acts as an endogenous danger-associated inflammatory mediator (also called alarmin). We demonstrated that HMGN1 not only directly stimulated cytokine production but also had the capacity to induce immune tolerance by a TLR4-dependent pathway, similar to lipopolysaccharide (LPS)-induced tolerance. HMGN1-induced tolerance was accompanied by a metabolic shift associated with the inhibition of the induction of Warburg effect (aerobic glycolysis) and histone deacetylation via Sirtuin-1. In addition, HMGN1 pre-challenge of mice also downregulated TNF production similar to LPS-induced tolerance in vivo. In conclusion, HMGN1 is an endogenous TLR4 ligand that can induce both acute stimulation of cytokine production and long-term tolerance, and thus it might play a modulatory role in sterile inflammatory processes such as those induced by infection, trauma, or ischemia.


Assuntos
Proteína HMGN1/imunologia , Leucócitos Mononucleares/imunologia , Sirtuína 1/imunologia , Receptor 4 Toll-Like/imunologia , Animais , Citocinas/sangue , Feminino , Humanos , Tolerância Imunológica , Imunidade Inata , Ligantes , Camundongos Endogâmicos C57BL
11.
J Crohns Colitis ; 12(5): 582-588, 2018 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-29361163

RESUMO

Background and Aims: Crohn's disease [CD] is a chronic inflammatory disease with unpredictable behaviour. More than half of CD patients eventually develop complications such as stenosis, for which they then require endoscopic dilatation or surgery, as no anti-fibrotic drugs are currently available. We aim to identify disease-modifying genes associated with fibrostenotic CD. Methods: We performed a within-case analysis comparing 'extreme phenotypes' using the Immunochip and replication of the top single nucleotide polymorphisms [SNPs] with Agena Bioscience in two independent case-control cohorts totalling 322 cases with fibrostenotis [recurrent after surgery] and 619 cases with purely inflammatory CD. Results: Combined meta-analysis resulted in a genome-wide significant signal for SNP rs11861007 [p = 6.0910-11], located on chromosome 16, in lncRNA RP11-679B19.1, an lncRNA of unknown function, and close to exon 9 of the WWOX gene, which codes for WW domain-containing oxidoreductase. We analysed mRNA expression of TGF-ß and downstream genes in ileocecal resection material from ten patients with and without the WWOX risk allele. Patients carrying the risk allele [A] showed enhanced colonic expression of TGF-ß compared to patients homozygous for the wild-type [G] allele [p = 0.0079]. Conclusion: We have identified a variant in WWOX and in lncRNA RP11-679B19.1 as a disease-modifying genetic variant associated with recurrent fibrostenotic CD and replicated this association in an independent cohort. WWOX can potentially play a crucial role in fibrostenosis in CD, being positioned at the crossroads of inflammation and fibrosis.


Assuntos
Doença de Crohn/genética , Doença de Crohn/metabolismo , RNA Mensageiro/metabolismo , Proteínas Supressoras de Tumor/genética , Oxidorredutase com Domínios WW/genética , Adolescente , Adulto , Alelos , Estudos de Casos e Controles , Constrição Patológica/etiologia , Doença de Crohn/complicações , Feminino , Fibrose , Estudo de Associação Genômica Ampla , Genômica , Humanos , Masculino , Fenótipo , Polimorfismo de Nucleotídeo Único , RNA Longo não Codificante/genética , Fator de Crescimento Transformador beta/genética , Adulto Jovem
12.
Cell Metab ; 24(6): 807-819, 2016 12 13.
Artigo em Inglês | MEDLINE | ID: mdl-27866838

RESUMO

Induction of trained immunity (innate immune memory) is mediated by activation of immune and metabolic pathways that result in epigenetic rewiring of cellular functional programs. Through network-level integration of transcriptomics and metabolomics data, we identify glycolysis, glutaminolysis, and the cholesterol synthesis pathway as indispensable for the induction of trained immunity by ß-glucan in monocytes. Accumulation of fumarate, due to glutamine replenishment of the TCA cycle, integrates immune and metabolic circuits to induce monocyte epigenetic reprogramming by inhibiting KDM5 histone demethylases. Furthermore, fumarate itself induced an epigenetic program similar to ß-glucan-induced trained immunity. In line with this, inhibition of glutaminolysis and cholesterol synthesis in mice reduced the induction of trained immunity by ß-glucan. Identification of the metabolic pathways leading to induction of trained immunity contributes to our understanding of innate immune memory and opens new therapeutic avenues.


Assuntos
Epigênese Genética , Fumaratos/metabolismo , Glutamina/metabolismo , Imunidade Inata/genética , Colesterol/metabolismo , Glucose/metabolismo , Glicólise , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Tolerância Imunológica , Macrófagos/metabolismo , Modelos Biológicos , Via de Pentose Fosfato/genética , Proteólise
13.
Eur J Immunol ; 46(11): 2574-2586, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27624090

RESUMO

Cells in homeostasis metabolize glucose mainly through the tricarboxylic acid cycle and oxidative phosphorylation, while activated cells switch their basal metabolism to aerobic glycolysis. In this study, we examined whether metabolic reprogramming toward aerobic glycolysis is important for the host response to Mycobacterium tuberculosis (Mtb). Through transcriptional and metabolite analysis we show that Mtb induces a switch in host cellular metabolism toward aerobic glycolysis in human peripheral blood mononuclear cells (PBMCs). The metabolic switch is TLR2 dependent but NOD2 independent, and is mediated in part through activation of the AKT-mTOR (mammalian target of rapamycin) pathway. We show that pharmacological inhibition of the AKT/mTOR pathway inhibits cellular responses to Mtb both in vitro in human PBMCs, and in vivo in a model of murine tuberculosis. Our findings reveal a novel regulatory layer of host responses to Mtb that will aid understanding of host susceptibility to Mtb, and which may be exploited for host-directed therapy.


Assuntos
Glicólise , Leucócitos Mononucleares/metabolismo , Mycobacterium tuberculosis/imunologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Animais , Antibacterianos/farmacologia , Perfilação da Expressão Gênica , Glucose/metabolismo , Glicólise/genética , Interações Hospedeiro-Patógeno , Humanos , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/microbiologia , Camundongos , Fosforilação Oxidativa , Transdução de Sinais/efeitos dos fármacos , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/imunologia , Receptor 2 Toll-Like/imunologia , Tuberculose/imunologia , Tuberculose/metabolismo , Tuberculose/microbiologia
14.
Nat Immunol ; 17(4): 406-13, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26950237

RESUMO

The acute phase of sepsis is characterized by a strong inflammatory reaction. At later stages in some patients, immunoparalysis may be encountered, which is associated with a poor outcome. By transcriptional and metabolic profiling of human patients with sepsis, we found that a shift from oxidative phosphorylation to aerobic glycolysis was an important component of initial activation of host defense. Blocking metabolic pathways with metformin diminished cytokine production and increased mortality in systemic fungal infection in mice. In contrast, in leukocytes rendered tolerant by exposure to lipopolysaccharide or after isolation from patients with sepsis and immunoparalysis, a generalized metabolic defect at the level of both glycolysis and oxidative metabolism was apparent, which was restored after recovery of the patients. Finally, the immunometabolic defects in humans were partially restored by therapy with recombinant interferon-γ, which suggested that metabolic processes might represent a therapeutic target in sepsis.


Assuntos
Citocinas/imunologia , Endotoxemia/imunologia , Metabolismo Energético/imunologia , Tolerância Imunológica/imunologia , Imunidade Inata/imunologia , Macrófagos/imunologia , Monócitos/imunologia , Sepse/imunologia , Trifosfato de Adenosina/metabolismo , Adulto , Animais , Antifúngicos/uso terapêutico , Aspergilose/tratamento farmacológico , Aspergilose/imunologia , Aspergilose/metabolismo , Candidíase Invasiva/tratamento farmacológico , Candidíase Invasiva/imunologia , Candidíase Invasiva/metabolismo , Endotoxemia/metabolismo , Infecções por Escherichia coli/imunologia , Infecções por Escherichia coli/metabolismo , Feminino , Glicólise , Humanos , Immunoblotting , Interferon gama/uso terapêutico , Ácido Láctico/metabolismo , Leucócitos/imunologia , Leucócitos/metabolismo , Lipopolissacarídeos/imunologia , Macrófagos/metabolismo , Masculino , Camundongos , Pessoa de Meia-Idade , Monócitos/metabolismo , NAD/metabolismo , Fosforilação Oxidativa , Consumo de Oxigênio , Estudos Prospectivos , Sepse/tratamento farmacológico , Sepse/metabolismo , Transcriptoma , Adulto Jovem
15.
J Biol Chem ; 291(15): 7961-72, 2016 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-26887946

RESUMO

The immune system is essential to maintain the mutualistic homeostatic interaction between the host and its micro- and mycobiota. Living as a commensal,Saccharomyces cerevisiaecould potentially shape the immune response in a significant way. We observed thatS. cerevisiaecells induce trained immunity in monocytes in a strain-dependent manner through enhanced TNFα and IL-6 production upon secondary stimulation with TLR ligands, as well as bacterial and fungal commensals. Differential chitin content accounts for the differences in training properties observed among strains, driving induction of trained immunity by increasing cytokine production and direct antimicrobial activity bothin vitroandin vivo These chitin-induced protective properties are intimately associated with its internalization, identifying a critical role of phagosome acidification to facilitate microbial digestion. This study reveals how commensal and passenger microorganisms could be important in promoting health and preventing mucosal diseases by modulating host defense toward pathogens and thus influencing the host microbiota-immune system interactions.


Assuntos
Quitina/imunologia , Imunidade Inata , Monócitos/microbiologia , Saccharomyces cerevisiae/imunologia , Animais , Parede Celular/imunologia , Humanos , Interleucina-6/imunologia , Camundongos Endogâmicos C57BL , Monócitos/imunologia , Fagocitose , Fator de Necrose Tumoral alfa/imunologia
16.
Immunity ; 43(4): 715-26, 2015 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-26488816

RESUMO

CARD9 is a central component of anti-fungal innate immune signaling via C-type lectin receptors, and several immune-related disorders are associated with CARD9 alterations. Here, we used a rare CARD9 variant that confers protection against inflammatory bowel disease as an entry point to investigating CARD9 regulation. We showed that the protective variant of CARD9, which is C-terminally truncated, acted in a dominant-negative manner for CARD9-mediated cytokine production, indicating an important role for the C terminus in CARD9 signaling. We identified TRIM62 as a CARD9 binding partner and showed that TRIM62 facilitated K27-linked poly-ubiquitination of CARD9. We identified K125 as the ubiquitinated residue on CARD9 and demonstrated that this ubiquitination was essential for CARD9 activity. Furthermore, we showed that similar to Card9-deficient mice, Trim62-deficient mice had increased susceptibility to fungal infection. In this study, we utilized a rare protective allele to uncover a TRIM62-mediated mechanism for regulation of CARD9 activation.


Assuntos
Proteínas Adaptadoras de Sinalização CARD/fisiologia , Candidíase Invasiva/imunologia , Receptores de Angiotensina/fisiologia , Receptores de Endotelina/fisiologia , Ubiquitina-Proteína Ligases/fisiologia , Adjuvantes Imunológicos/farmacologia , Animais , Proteínas Adaptadoras de Sinalização CARD/química , Proteínas Adaptadoras de Sinalização CARD/deficiência , Proteínas Adaptadoras de Sinalização CARD/genética , Candidíase Invasiva/genética , Colite/induzido quimicamente , Colite/genética , Colite/prevenção & controle , Citocinas/biossíntese , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Genes Dominantes , Predisposição Genética para Doença , Células HEK293 , Células HeLa , Humanos , Doenças Inflamatórias Intestinais/genética , Camundongos , Camundongos da Linhagem 129 , Camundongos Knockout , Mapeamento de Interação de Proteínas , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/fisiologia , Processamento de Proteína Pós-Traducional , Estrutura Terciária de Proteína , Receptores de Angiotensina/química , Receptores de Angiotensina/deficiência , Receptores de Endotelina/química , Receptores de Endotelina/deficiência , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais , Organismos Livres de Patógenos Específicos , Proteínas com Motivo Tripartido , Ubiquitina-Proteína Ligases/química , Ubiquitinação
17.
Cytokine ; 76(2): 465-472, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26297607

RESUMO

Although it is known that Borrelia species express sugar-like structures on their outer surface, not much is known about the role of these structures in immune recognition by host cells. Fungi, like Candida albicans, are mainly recognized by C-type lectin receptors, in specific Dectin-1 and Dectin-2. In this study we assessed the role of Dectin-1 and Dectin-2 in the recognition process of Borrelia spirochetes. Using specific inhibitors against these receptors on human cells did not influenced cytokine production. Individuals carrying a SNP leading to an early stop codon in the DECTIN-1 gene also did not lead to differential induction of Borrelia-dependent cytokines. After injection of live Borrelia into knee joints of Dectin-2 deficient mice a trend towards lower inflammation was observed. Inhibition of Syk in human cells resulted in lower cytokine production after Borrelia stimulation. In conclusion, Dectin-1 and Dectin-2 seem not to play a major role in Borrelia recognition or Borrelia-induced inflammation. However, Syk seems to be involved in Borrelia-induced cytokine production.


Assuntos
Borrelia burgdorferi/fisiologia , Citocinas/biossíntese , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Lectinas Tipo C/metabolismo , Proteínas Tirosina Quinases/metabolismo , Animais , Feminino , Lectinas Tipo C/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Quinase Syk
18.
Eur J Clin Microbiol Infect Dis ; 34(5): 963-974, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25579795

RESUMO

The induction of host defense against Candida species is initiated by recognition of the fungi by pattern recognition receptors and activation of downstream pathways that produce inflammatory mediators essential for infection clearance. In this study, we present complementary evidence based on transcriptome analysis, genetics, and immunological studies in knockout mice and humans that the cytosolic RIG-I-like receptor MDA5 (IFIH1) has an important role in the host defense against C. albicans. Firstly, IFIH1 expression in macrophages is specifically induced by invasive C. albicans hyphae, and patients suffering from chronic mucocutaneous candidiasis (CMC) express lower levels of MDA5 than healthy controls. Secondly, there is a strong association between missense variants in the IFIH1 gene (rs1990760 and rs3747517) and susceptibility to systemic Candida infections. Thirdly, cells from Mda5 knockout mice and human peripheral blood mononuclear cells (PBMCs) with different IFIH1 genotypes display an altered cytokine response to C. albicans. These data strongly suggest that MDA5 is involved in immune responses to Candida infection. As a receptor for viral RNA, MDA5 until now has been linked to antiviral host defense, but these novel studies show unexpected effects in antifungal immunity as well. Future studies are warranted to explore the potential of MDA5 as a novel target for immunotherapeutic strategies.


Assuntos
Candida/imunologia , Candidemia/imunologia , RNA Helicases DEAD-box/metabolismo , Adulto , Animais , Células Cultivadas , Estudos de Coortes , RNA Helicases DEAD-box/deficiência , Suscetibilidade a Doenças , Humanos , Helicase IFIH1 Induzida por Interferon , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/microbiologia , Camundongos Knockout , Polimorfismo de Nucleotídeo Único
19.
Cytokine ; 71(2): 215-22, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25461401

RESUMO

BACKGROUND: Candida albicans is an opportunistic fungal pathogen that induces strong proinflammatory responses, such as IL-1ß production. Much less is known about the induction of immune modulatory cytokines, such as the IL-1 receptor antagonist (IL-1Ra) that is the main natural antagonist of IL-1, by C. albicans. METHODS: Peripheral blood mononuclear cells (PBMC) of healthy individuals were stimulated with C. albicans and different components of the fungal cell wall. The role of pathogen recognition receptors (PRRs) for the induction of IL-1ß and IL-1Ra was investigated by using specific blockers or in PBMC from Dectin-1 deficient patients. RESULTS: C. albicans induced a strong IL-1Ra response, and this induction was primarily induced by the cell-wall component ß-glucan. Blocking IL-1Ra significantly increased C. albicans ß-glucan hyphae induced IL-1ß and IL-6 production. Surprisingly, blocking the ß-glucan receptor Dectin-1 or the downstream Syk or Raf-1 pathways only marginally reduced C. albicans-induced IL-1Ra production, while blocking of the complement receptor 3 (CR3), TLR2 or TLR4 had no effect. In line with this, blocking MAP kinases had little effect on Candida-induced IL-1Ra production. PBMC isolated from Dectin-1 deficient patients produced normal IL-1Ra amounts in response to C. albicans stimulation. Interestingly, the IL-1Ra synthesis induced by ß-glucan was blocked by inhibitors of the Akt/PI3K pathway. CONCLUSIONS: ß-glucan of C. albicans induces a strong IL-1Ra response, which is independent of the ß-glucan receptors dectin-1 and CR3. These data strongly argue for the existence of an unknown ß-glucan receptor that specifically induces an Akt/PI3K-dependent anti-inflammatory IL-1Ra response upon recognition of C. albicans.


Assuntos
Candida albicans/imunologia , Proteína Antagonista do Receptor de Interleucina 1/imunologia , Lectinas Tipo C/imunologia , Antígeno de Macrófago 1/imunologia , beta-Glucanas/imunologia , Candida albicans/fisiologia , Células Cultivadas , Ensaio de Imunoadsorção Enzimática , Interações Hospedeiro-Patógeno/imunologia , Humanos , Proteína Antagonista do Receptor de Interleucina 1/metabolismo , Interleucina-1beta/imunologia , Interleucina-1beta/metabolismo , Lectinas Tipo C/metabolismo , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/microbiologia , Antígeno de Macrófago 1/metabolismo , Fosfatidilinositol 3-Quinases/imunologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/imunologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/imunologia
20.
Proc Natl Acad Sci U S A ; 111(42): E4478-84, 2014 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-25288745

RESUMO

Toll-like receptor (TLR)10 is the only pattern-recognition receptor without known ligand specificity and biological function. We demonstrate that TLR10 is a modulatory receptor with mainly inhibitory effects. Blocking TLR10 by antagonistic antibodies enhanced proinflammatory cytokine production, including IL-1ß, specifically after exposure to TLR2 ligands. Blocking TLR10 after stimulation of peripheral blood mononuclear cells with pam3CSK4 (Pam3Cys) led to production of 2,065 ± 106 pg/mL IL-1ß (mean ± SEM) in comparison with 1,043 ± 51 pg/mL IL-1ß after addition of nonspecific IgG antibodies. Several mechanisms mediate the modulatory effects of TLR10: on the one hand, cotransfection in human cell lines showed that TLR10 acts as an inhibitory receptor when forming heterodimers with TLR2; on the other hand, cross-linking experiments showed specific induction of the anti-inflammatory cytokine IL-1 receptor antagonist (IL-1Ra, 16 ± 1.7 ng/mL, mean ± SEM). After cross-linking anti-TLR10 antibody, no production of IL-1ß and other proinflammatory cytokines could be found. Furthermore, individuals bearing TLR10 polymorphisms displayed an increased capacity to produce IL-1ß, TNF-α, and IL-6 upon ligation of TLR2, in a gene-dose-dependent manner. The modulatory effects of TLR10 are complex, involving at least several mechanisms: there is competition for ligands or for the formation of heterodimer receptors with TLR2, as well as PI3K/Akt-mediated induction of the anti-inflammatory cytokine IL-1Ra. Finally, transgenic mice expressing human TLR10 produced fewer cytokines when challenged with a TLR2 agonist. In conclusion, to our knowledge we demonstrate for the first time that TLR10 is a modulatory pattern-recognition receptor with mainly inhibitory properties.


Assuntos
Inflamação/metabolismo , Receptores de Reconhecimento de Padrão/metabolismo , Receptor 10 Toll-Like/metabolismo , Animais , Citocinas/metabolismo , Células HEK293 , Humanos , Proteína Antagonista do Receptor de Interleucina 1/metabolismo , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Leucócitos Mononucleares/citologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Polimorfismo Genético , Polimorfismo de Nucleotídeo Único , Interferência de RNA , Transdução de Sinais , Receptor 2 Toll-Like/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Regulação para Cima
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