RESUMO
BACKGROUND: Flucloxacillin (FLX)-induced liver injury is immune-mediated and highly associated to HLA-B∗57:01 expression. Host factors leading to drug-induced liver injury are not yet well understood. OBJECTIVE: Characterize in vivo immune mechanisms determining the development of CD8+ T cells reactive to FLX in animals expressing the risk human leukocyte antigen (HLA) allotype. METHODS: HLA-B∗57:01 transgenic mice (Tg) or Tg strains with H2-KbDb knockout (Tg/KO) or H2-KbDb/PD-1 double knockout (Tg/DKO) were treated with drug and/or anti-CD4 antibody. Drug-induced liver injury was evaluated on the basis of liver enzyme and histologic changes at day 10 of treatment. FLX-reactive CD8+ T cells were characterized in vitro by release of effector molecules on drug restimulation, gene expression, and flow cytometry analysis, and functionality tested for hepatic cytotoxicity. RESULTS: CD8+ T-cell responses to FLX in Tg were dependent on both HLA and mouse major histocompatibility complex I presentation and in vivo priming. Eliminating H2-KbDb in Tg/KO to allow exclusive presentation of FLX by HLA resulted in a less robust drug-specific CD8+T-cell response unless CD4+ cells, including regulatory T cells, were depleted. Treatment of Tg/KO with anti-CD4 antibody and FLX led to subclinical liver inflammation associated with an increase in PD1+CD8+ T cells in the lymphoid organs and liver. Impaired PD-1 expression in Tg/DKO led to liver histopathologic and transcriptional alterations but without hepatic enzyme elevations. Moreover, effector lymphocytes accumulated in the liver and showed FLX-dependent hepatic cytotoxicity in vitro when tolerogenic liver cells were depleted. CONCLUSIONS: In our in vivo models, FLX primes CD8+ T cells to recognize drug presented by HLA-B∗57:01 and murine major histocompatibility complex I. HLA-B∗57:01-dependent CD8+ T-cell reaction to FLX is limited by the presence of CD4+ cells, presumably regulatory T cells, and PD-1 expression. Tolerogenic hepatic cells limit clinical disease through PD-L1 or additional unexplored mechanisms.
Assuntos
Linfócitos T CD8-Positivos , Doença Hepática Induzida por Substâncias e Drogas , Camundongos , Humanos , Animais , Floxacilina/efeitos adversos , Floxacilina/metabolismo , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/metabolismo , Antígenos de Histocompatibilidade Classe I , Camundongos Transgênicos , Antígenos HLA/genética , Modelos Animais de Doenças , Doença Hepática Induzida por Substâncias e Drogas/genética , Doença Hepática Induzida por Substâncias e Drogas/metabolismoRESUMO
The first case of human transmission of SARS-CoV-2 was reported in China in December 2019. A few months later, this viral infection had spread worldwide and became a pandemic. The disease caused by SARS-CoV-2, termed COVID-19, is multifactorial and associated with both specific antiviral as well as inflammatory responses, the extent of which may determine why some individuals are asymptomatic while others develop serious complications. Here we review possible life-threating immune events that can occur during disease progression to uncover key factors behind COVID-19 severity and provide suggestions for interventions with repurposed drugs in well-controlled and randomized clinical trials. These drugs include therapeutics with potential to inhibit SARS-CoV-2 entry into host cells such as serine protease inhibitors of the cellular protease TMPS2 and drugs targeting the renin-angiotensin system; antivirals with potential to block SARS-CoV-2 replication or factors that could boost the antiviral response; monoclonal antibodies targeting pro-inflammatory cytokines that drive the hyperinflammatory response during COVID-19 progression toward the severe stage and therapeutics that could ameliorate the function of the lungs. Furthermore, in order to help make more informed decisions on the timing of the intervention with the drugs listed in this review, we have grouped these therapeutics according to the stage of COVID-19 progression that we considered most appropriate for their mechanism of action.
Assuntos
Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/imunologia , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/imunologia , Antivirais/uso terapêutico , COVID-19 , Progressão da Doença , Humanos , Inflamação/tratamento farmacológico , Inflamação/imunologia , Pandemias , Tratamento Farmacológico da COVID-19RESUMO
Neoantigen formation due to the interaction of drug molecules with human leukocyte antigen (HLA)-peptide complexes can lead to severe hypersensitivity reactions. Flucloxacillin (FLX), a ß-lactam antibiotic for narrow-spectrum gram-positive bacterial infections, has been associated with severe immune-mediated drug-induced liver injury caused by an influx of T-lymphocytes targeting liver cells potentially recognizing drug-haptenated peptides in the context of HLA-B*57:01. To identify immunopeptidome changes that could lead to drug-driven immunogenicity, we used mass spectrometry to characterize the proteome and immunopeptidome of B-lymphoblastoid cells solely expressing HLA-B*57:01 as MHC-I molecules. Selected drug-conjugated peptides identified in these cells were synthesized and tested for their immunogenicity in HLA-B*57:01-transgenic mice. T cell responses were evaluated in vitro by immune assays. The immunopeptidome of FLX-treated cells was more diverse than that of untreated cells, enriched with peptides containing carboxy-terminal tryptophan and FLX-haptenated lysine residues on peptides. Selected FLX-modified peptides with drug on P4 and P6 induced drug-specific CD8+ T cells in vivo. FLX was also found directly linked to the HLA K146 that could interfere with KIR-3DL or peptide interactions. These studies identify a novel effect of antibiotics to alter anchor residue frequencies in HLA-presented peptides which may impact drug-induced inflammation. Covalent FLX-modified lysines on peptides mapped drug-specific immunogenicity primarily at P4 and P6 suggesting these peptide sites as drivers of off-target adverse reactions mediated by FLX. FLX modifications on HLA-B*57:01-exposed lysines may also impact interactions with KIR or TCR and subsequent NK and T cell function.
Assuntos
Floxacilina/imunologia , Antígenos HLA-B/imunologia , Haptenos/imunologia , Peptídeos/imunologia , Animais , Linhagem Celular , Antígenos HLA-B/genética , Humanos , Camundongos , Camundongos Transgênicos , Peptídeos/genéticaRESUMO
BACKGROUND: The diuretic response has been shown to be a robust independent marker of cardiovascular outcomes in acute heart failure (ADHF) patients. The objectives of this clinical research, will aim are to: a) include diuresis in the formula for diuretic response (R-to-D); b) add to R-to-D the value of a pre-discharged determination of galectin-3 and BNP in predicting mid-term clinical outcome. METHODS: Consecutive patients discharged alive after an ADHF were enrolled. All patients underwent BNP and galectin-3, a 6 min walk test and an echocardiogram together with diuresis and body weight during diuretic administration. Death by any cause, cardiac transplantation and worsening HF requiring readmission to the hospital were considered cardiovascular events. RESULTS: 141 patients (98 males, age 73.8) were analysed (follow-up 17 months). During the follow-up 45 (31.9%) events were scheduled (19 cardiac deaths, 26 re-hospitalisation for HF). Patients who experienced CV-event had a worst renal function (p = 0.003), an higher BNP (p = 0.006) and galectin-3 (p = 0.008). At multivariate analysis, only R-to-D, galectin-3 and BNP showed a significant correlation with worst clinical prognosis (respectively p = 0.043; OR 6.01; p = 0.01; OR 8.9; p = 0.02 OR 10.38), independently of age and renal function. Kaplan-Meier curves depicted a powerful stratification using an R-to-D <1.2 kg/40 mg furosemide (log rank 10.96; p = 0.0009). Adding R-to-D<1.2 mg/40 mg furosemide to galectin-3>17.6 pg/mL and BNP>500 pg/mL the predictive value improved (log rank 23.59; p = 0.0001). CONCLUSION: Adding R-to-D to Gal-3 and BNP, a single pre-discharge strategy testing seemed to obtain a satisfactorily predictive value in alive HF patients discharged after an ADHF episode.
Assuntos
Diuréticos/uso terapêutico , Furosemida/uso terapêutico , Galectina 3/sangue , Insuficiência Cardíaca/tratamento farmacológico , Peptídeo Natriurético Encefálico/sangue , Idoso , Biomarcadores/sangue , Proteínas Sanguíneas , Ecocardiografia , Feminino , Galectinas , Insuficiência Cardíaca/mortalidade , Insuficiência Cardíaca/fisiopatologia , Hospitalização , Humanos , Masculino , Alta do Paciente , Recidiva , Resultado do TratamentoRESUMO
Adverse drug reactions (ADRs) are a major obstacle to drug development, and some of these, including hypersensitivity reactions to the HIV reverse transcriptase inhibitor abacavir (ABC), are associated with HLA alleles, particularly HLA-B*57:01. However, not all HLA-B*57:01+ patients develop ADRs, suggesting that in addition to the HLA genetic risk, other factors may influence the outcome of the response to the drug. To study HLA-linked ADRs in vivo, we generated HLA-B*57:01-Tg mice and show that, although ABC activated Tg mouse CD8+ T cells in vitro in a HLA-B*57:01-dependent manner, the drug was tolerated in vivo. In immunocompetent Tg animals, ABC induced CD8+ T cells with an anergy-like phenotype that did not lead to ADRs. In contrast, in vivo depletion of CD4+ T cells prior to ABC administration enhanced DC maturation to induce systemic ABC-reactive CD8+ T cells with an effector-like and skin-homing phenotype along with CD8+ infiltration and inflammation in drug-sensitized skin. B7 costimulatory molecule blockade prevented CD8+ T cell activation. These Tg mice provide a model for ABC tolerance and for the generation of HLA-B*57:01-restricted, ABC-reactive CD8+ T cells dependent on both HLA genetic risk and immunoregulatory host factors.
Assuntos
Didesoxinucleosídeos/efeitos adversos , Tolerância a Medicamentos/genética , Tolerância a Medicamentos/imunologia , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/genética , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/imunologia , Antígenos HLA-B/genética , Animais , Fármacos Anti-HIV/efeitos adversos , Linfócitos T CD8-Positivos/imunologia , Modelos Animais de Doenças , Hipersensibilidade a Drogas/genética , Hipersensibilidade a Drogas/imunologia , Feminino , Humanos , Técnicas In Vitro , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Inibidores da Transcriptase Reversa/efeitos adversosRESUMO
Galectin-3 demonstrated to be a robust independent marker of cardiovascular mid-term (18-month) outcome in heart failure (HF) patients. The objective of this study was to analyze the value of a predischarged determination of plasma galectin-3 alone and with plasma brain natriuretic peptide (BNP) in predicting mid-term outcome in frequent-flyers (FF) HF (≥2 hospitalization for HF/year)/dead patients discharged after an acute decompensated HF (ADHF) episode.All FF chronic HF subjects discharged alive after an ADHF were enrolled. All patients underwent a determination of BNP and galectin-3, a 6-minute walk test, and an echocardiogram within 48âhours upon hospital discharge. Death by any cause, cardiac transplantation, and worsening HF requiring readmission to hospital were considered cardiovascular events.Eighty-three patients (67 males, age 73.2â±â8.6 years old) were analyzed (mean follow-up 11.6â±â5.2 months; range 4-22 months). During the follow-up 38 events (45.7%) were scheduled: (13 cardiac deaths, 35 rehospitalizations for ADHF). According to medical history, in 33 patients (39.8%) a definition of FF HF patients was performed (range 2-4âhospitalization/year). HF patients who suffered an event (FF or death) demonstrated more impaired ventricular function (P = 0.037), higher plasma BNP (P = 0.005), and Gal-3 at predischarge evaluation (P = 0.027). Choosing adequate cut-off points (BNPâ≥â500âpg/mL and Gal-3â≥â17.6âng/mL), the Kaplan-Meier curves depicted the powerful stratification using BNPâ+âGal-3 in predicting clinical course at mid-term follow-up (log rank 5.65; P = 0.017).Adding Gal-3 to BNP, a single predischarge strategy testing seemed to obtain a satisfactorily predictive value in alive HF patients discharged after an ADHF episode.
Assuntos
Galectina 3/sangue , Insuficiência Cardíaca/sangue , Insuficiência Cardíaca/mortalidade , Peptídeo Natriurético Encefálico/sangue , Idoso , Doença Crônica , Feminino , Humanos , Masculino , Alta do Paciente , Valor Preditivo dos Testes , PrognósticoRESUMO
Oligonucleotide aptamers represent a novel platform for creating ligands with desired specificity, and they offer many potentially significant advantages over monoclonal antibodies in terms of feasibility, cost, and clinical applicability. However, the isolation of high-affinity aptamer ligands from random oligonucleotide pools has been challenging. Although high-throughput sequencing (HTS) promises to significantly facilitate systematic evolution of ligands by exponential enrichment (SELEX) analysis, the enormous datasets generated in the process pose new challenges for identifying those rare, high-affinity aptamers present in a given pool. We show that emulsion PCR preserves library diversity, preventing the loss of rare high-affinity aptamers that are difficult to amplify. We also demonstrate the importance of using reference targets to eliminate binding candidates with reduced specificity. Using a combination of bioinformatics and functional analyses, we show that the rate of amplification is more predictive than prevalence with respect to binding affinity and that the mutational landscape within a cluster of related aptamers can guide the identification of high-affinity aptamer ligands. Finally, we demonstrate the power of this selection process for identifying cross-species aptamers that can bind human receptors and cross-react with their murine orthologs.
Assuntos
Aptâmeros de Nucleotídeos/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , Técnica de Seleção de Aptâmeros/métodos , Animais , Biblioteca Gênica , Humanos , Ligantes , Camundongos , Mutação , Reação em Cadeia da Polimerase , Receptores de Interleucina-10/metabolismo , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismoRESUMO
UNLABELLED: The interplay between dendritic cells (DC) and γδ T lymphocytes represents a network of paracrine and cell contact interactions important for an integrated immune response to pathogens. HIV-1 infection dramatically affects the number and functions of both cell populations, and DC/γδ T cell cross talk may represent a target of virus-induced immune escape. We investigated whether HIV-exposed DC could deliver aberrant signals to interacting γδ T cells. Here we report that the interaction of human γδ T lymphocytes with HIV-1-exposed autologous monocyte-derived DC, but not direct exposure to the virus, impairs lymphocyte expansion and gamma interferon (IFN-γ) production in response to phosphoantigens. This effect is independent of virus strain and occurred in 55% of the donors analyzed. The donor-dependent variation observed relies on the responsiveness of DC to HIV-1 and is strictly related to the capacity of the virus to suppress the maturation-induced expression of interleukin 12 (IL-12). In fact, γδ T cell response to phosphoantigens is almost completely recovered when this cytokine is exogenously added to the DC/lymphocyte cocultures. Interestingly, we show that γδ T lymphocytes are recruited by HIV-1-exposed DC through a CCR5-mediated mechanism and exert a CCL4-mediated control on virus dissemination within DC and susceptible CD4(+) T lymphocytes. These results demonstrate an association between HIV-induced DC dysfunction and alterations of γδ T cell responses. The aberrant cross talk between these two cell populations may contribute to the pathogenesis of HIV infection by further reducing the strength of antiviral immune response. IMPORTANCE: This study provides new evidence on the mechanisms exploited by HIV-1 to evade the host immune response. We report that HIV-1 impairs the cross talk between DC and γδ T lymphocytes, by reducing the capacity of DC to promote functional γδ T cell activation. Interestingly, the virus does not per se interfere with γδ T cell activation, thus highlighting the key role of early DC-HIV-1 interaction in this phenomenon. Furthermore, the results obtained unravel the novel role of γδ T cells in controlling HIV-1 dissemination within the DC population as well as virus transfer to susceptible CD4(+) T lymphocytes. The interactions of DC with innate lymphocytes represent a major control mechanism for an integrated immune response to infection. Understanding how HIV-1 harnesses these pathways may provide important insights on the pathogenesis of disease and offer new opportunities for therapeutic interventions.
Assuntos
Células Dendríticas/imunologia , HIV-1/imunologia , HIV-1/fisiologia , Evasão da Resposta Imune , Linfócitos T/imunologia , Proliferação de Células , Humanos , Interferon gama/metabolismo , Interleucina-12/antagonistas & inibidoresRESUMO
Recognition of microbial components via innate receptors including the C-type lectin receptor Dectin-1, together with the inflammatory environment, programs dendritic cells (DCs) to orchestrate the magnitude and type of adaptive immune responses. The exposure to ß-glucan, a known Dectin-1 agonist and component of fungi, yeasts, and certain immune support supplements, activates DCs to induce T helper (Th)17 cells that are essential against fungal pathogens and extracellular bacteria but may trigger inflammatory pathology or autoimmune diseases. However, the exact mechanisms of DC programming by ß-glucan have not yet been fully elucidated. Using a gene expression/perturbation approach, we demonstrate that in human DCs ß-glucan transcriptionally activates via an interleukin (IL)-1- and inflammasome-mediated positive feedback late-induced genes that bridge innate and adaptive immunity. We report that in addition to its known ability to directly prime T cells toward the Th17 lineage, IL-1 by promoting the transcriptional cofactor inhibitor of κB-ζ (IκB-ζ) also programs ß-glucan-exposed DCs to express cell adhesion and migration mediators, antimicrobial molecules, and Th17-polarizing factors. Interferon (IFN)-γ interferes with the IL-1/IκB-ζ axis in ß-glucan-activated DCs and promotes T cell-mediated immune responses with increased release of IFN-γ and IL-22, and diminished production of IL-17. Thus, our results identify IL-1 and IFN-γ as regulators of DC programming by ß-glucan. These molecular networks provide new insights into the regulation of the Th17 response as well as new targets for the modulation of immune responses to ß-glucan-containing microorganisms.
Assuntos
Células Dendríticas/imunologia , Proteínas I-kappa B/metabolismo , Interferon gama/fisiologia , Interleucina-1/fisiologia , Proteínas Nucleares/metabolismo , beta-Glucanas/farmacologia , Proteínas Adaptadoras de Transdução de Sinal , Células Cultivadas , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/metabolismo , Humanos , Proteína Antagonista do Receptor de Interleucina 1/fisiologia , Subunidade p19 da Interleucina-23/genética , Subunidade p19 da Interleucina-23/metabolismo , Lipopolissacarídeos/farmacologia , Regiões Promotoras Genéticas , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Auxiliares-Indutores/metabolismo , Transcrição Gênica , Ativação Transcricional , TranscriptomaRESUMO
CLRs are predominantly expressed in macrophages and myeloid DCs, where they play a key role in antigen recognition, scavenging, and host defense against pathogens. To identify novel immunoregulatory cytokines and networks involved in the control of these functions, we analyzed the coordinate effects of IFN-ß and IL-3 on CLR expression, antigen uptake, and phagocytosis in human MDMs and MDDCs. We report that these cytokines exert opposite regulatory effects on the expression of CLRs and endocytic/phagocytic activities of MDMs. Specifically, IFN-ß markedly inhibits the expression of MR and Dectin-1 during MDM differentiation and impairs the capacity of MDM to internalize antigens and phagocytose unopsonized Candida albicans. Conversely, IL-3 up-modulates MR, Dectin-1, and DC-SIGN, thus allowing more efficient uptake/phagocytosis. Interestingly, IL-3 counteracts the IFN-ß effect on antigen uptake/processing by fully restoring MR expression in IFN-ß-primed MDMs. In contrast, the phagocytic activity is only partially restored as a result of the failure of IL-3 in counteracting IFN-ß-induced Dectin-1 suppression. Notably, IFN-ß-mediated impairment of CLR expression/function occurs in macrophages but not in MDDCs. These results identify IFN-ß and IL-3 as unrecognized regulators of CLR expression and function, unraveling a novel interaction between these cytokines instrumental for the regulation of the macrophage response to pathogens.
Assuntos
Antígenos/imunologia , Interferon beta/farmacologia , Interleucina-3/farmacologia , Lectinas Tipo C/genética , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Fagocitose/efeitos dos fármacos , Fagocitose/imunologia , Antígenos/metabolismo , Candida albicans/imunologia , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Endocitose/efeitos dos fármacos , Endocitose/imunologia , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Lectinas Tipo C/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Ativação de Macrófagos/imunologia , Macrófagos/metabolismo , Macrófagos/microbiologia , Glicoproteínas de Membrana , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Monócitos/metabolismo , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Receptores Imunológicos/genética , Receptores Imunológicos/metabolismoRESUMO
The gut microbiota influences both local and systemic inflammation. Inflammation contributes to development, progression, and treatment of cancer, but it remains unclear whether commensal bacteria affect inflammation in the sterile tumor microenvironment. Here, we show that disruption of the microbiota impairs the response of subcutaneous tumors to CpG-oligonucleotide immunotherapy and platinum chemotherapy. In antibiotics-treated or germ-free mice, tumor-infiltrating myeloid-derived cells responded poorly to therapy, resulting in lower cytokine production and tumor necrosis after CpG-oligonucleotide treatment and deficient production of reactive oxygen species and cytotoxicity after chemotherapy. Thus, optimal responses to cancer therapy require an intact commensal microbiota that mediates its effects by modulating myeloid-derived cell functions in the tumor microenvironment. These findings underscore the importance of the microbiota in the outcome of disease treatment.
Assuntos
Intestinos/microbiologia , Microbiota/fisiologia , Neoplasias/imunologia , Neoplasias/terapia , Microambiente Tumoral/imunologia , Animais , Antibacterianos/administração & dosagem , Apresentação de Antígeno/genética , Antineoplásicos/uso terapêutico , Bactérias/efeitos dos fármacos , Fenômenos Fisiológicos Bacterianos/efeitos dos fármacos , Regulação para Baixo , Regulação da Expressão Gênica , Vida Livre de Germes , Imunoterapia , Inflamação/genética , Melanoma Experimental , Camundongos , Camundongos Endogâmicos C57BL , Microbiota/efeitos dos fármacos , Transplante de Neoplasias , Neoplasias/microbiologia , Oligodesoxirribonucleotídeos/uso terapêutico , Compostos Organoplatínicos/uso terapêutico , Oxaliplatina , Fagocitose/genética , Espécies Reativas de Oxigênio/metabolismo , Simbiose , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Fungal pathogens elicit cytokine responses downstream of immunoreceptor tyrosine-based activation motif (ITAM)-coupled or hemiITAM-containing receptors and TLRs. The Linker for Activation of B cells/Non-T cell Activating Linker (LAB/NTAL) encoded by Lat2, is a known regulator of ITAM-coupled receptors and TLR-associated cytokine responses. Here we demonstrate that LAB is involved in anti-fungal immunity. We show that Lat2-/- mice are more susceptible to C. albicans infection than wild type (WT) mice. Dendritic cells (DCs) express LAB and we show that it is basally phosphorylated by the growth factor M-CSF or following engagement of Dectin-2, but not Dectin-1. Our data revealed a unique mechanism whereby LAB controls basal and fungal/pathogen-associated molecular patterns (PAMP)-induced nuclear ß-catenin levels. This in turn is important for controlling fungal/PAMP-induced cytokine production in DCs. C. albicans- and LPS-induced IL-12 and IL-23 production was blunted in Lat2-/- DCs. Accordingly, Lat2-/- DCs directed reduced Th1 polarization in vitro and Lat2-/- mice displayed reduced Natural Killer (NK) and T cell-mediated IFN-γ production in vivo/ex vivo. Thus our data define a novel link between LAB and ß-catenin nuclear accumulation in DCs that facilitates IFN-γ responses during anti-fungal immunity. In addition, these findings are likely to be relevant to other infectious diseases that require IL-12 family cytokines and an IFN-γ response for pathogen clearance.
Assuntos
Sistema y+ de Transporte de Aminoácidos/imunologia , Candidíase/imunologia , Células Dendríticas/imunologia , Cadeias Leves da Proteína-1 Reguladora de Fusão/imunologia , Lectinas Tipo C/imunologia , beta Catenina/imunologia , Sistema y+ de Transporte de Aminoácidos/metabolismo , Animais , Candidíase/metabolismo , Células Dendríticas/metabolismo , Modelos Animais de Doenças , Feminino , Cadeias Leves da Proteína-1 Reguladora de Fusão/metabolismo , Interferon gama/biossíntese , Interferon gama/imunologia , Interleucina-12/biossíntese , Interleucina-12/imunologia , Lectinas Tipo C/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/imunologia , beta Catenina/metabolismoRESUMO
Mycobacterium tuberculosis is a virulent intracellular pathogen that survives in macrophages even in the presence of an intact adaptive immune response. Type I IFNs have been shown to exacerbate tuberculosis in mice and to be associated with disease progression in infected humans. Nevertheless, the mechanisms by which type I IFNs regulate the host response to M. tuberculosis infection are poorly understood. In this study, we show that M. tuberculosis induces an IFN-related gene expression signature in infected primary human macrophages, which is dependent on host type I IFN signaling as well as the mycobacterial virulence factor, region of difference-1. We further demonstrate that type I IFNs selectively limit the production of IL-1ß, a critical mediator of immunity to M. tuberculosis. This regulation occurs at the level of IL1B mRNA expression, rather than caspase-1 activation or autocrine IL-1 amplification and appears to be preferentially used by virulent mycobacteria since avirulent M. bovis bacillus Calmette-Guérin (BCG) fails to trigger significant expression of type I IFNs or release of mature IL-1ß protein. The latter property is associated with decreased caspase-1-dependent IL-1ß maturation in the BCG-infected macrophages. Interestingly, human monocytes in contrast to macrophages produce comparable levels of IL-1ß in response to either M. tuberculosis or BCG. Taken together, these findings demonstrate that virulent and avirulent mycobacteria employ distinct pathways for regulating IL-1ß production in human macrophages and reveal that in the case of M. tuberculosis infection the induction of type I IFNs is a major mechanism used for this purpose.
Assuntos
Interferon Tipo I/imunologia , Interleucina-1beta/biossíntese , Macrófagos/imunologia , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/patogenicidade , Transdução de Sinais/imunologia , Western Blotting , Ensaio de Imunoadsorção Enzimática , Expressão Gênica , Regulação da Expressão Gênica/imunologia , Humanos , Interferon Tipo I/metabolismo , Macrófagos/metabolismo , Macrófagos/microbiologia , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Signaling through the adaptor protein myeloid differentiation factor 88 (MyD88) promotes carcinogenesis in several cancer models. In contrast, MyD88 signaling has a protective role in the development of azoxymethane (AOM)/dextran sodium sulfate (DSS) colitis-associated cancer (CAC). The inability of Myd88(-/-) mice to heal ulcers generated upon injury creates an altered inflammatory environment that induces early alterations in expression of genes encoding proinflammatory factors, as well as pathways regulating cell proliferation, apoptosis, and DNA repair, resulting in a dramatic increase in adenoma formation and progression to infiltrating adenocarcinomas with frequent clonal mutations in the beta-catenin gene. Others have reported that toll-like receptor (Tlr) 4-deficient mice have a similar susceptibility to colitis to Myd88-deficient mice but, unlike the latter, are resistant to CAC. We have observed that mice deficient for Tlr2 or Il1r do not show a differential susceptibility to colitis or CAC. However, upon AOM/DSS treatment Il18(-/-) and Il18r1(-/-) mice were more susceptible to colitis and polyp formation than wild-type mice, suggesting that the phenotype of Myd88(-/-) mice is, in part, a result of their inability to signal through the IL-18 receptor. This study revealed a previously unknown level of complexity surrounding MyD88 activities downstream of different receptors that impact tissue homeostasis and carcinogenesis.
Assuntos
Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Interleucina-18/metabolismo , Fator 88 de Diferenciação Mieloide/metabolismo , Transdução de Sinais/fisiologia , Adenocarcinoma/induzido quimicamente , Adenocarcinoma/genética , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Azoximetano/farmacologia , Proliferação de Células/efeitos dos fármacos , Colo/efeitos dos fármacos , Colo/metabolismo , Colo/patologia , Neoplasias do Colo/induzido quimicamente , Neoplasias do Colo/genética , Pólipos do Colo/patologia , Ciclo-Oxigenase 2/genética , Enzimas Reparadoras do DNA/genética , Sulfato de Dextrana/farmacologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/genética , Perfilação da Expressão Gênica , Predisposição Genética para Doença/genética , Doenças Inflamatórias Intestinais/induzido quimicamente , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/metabolismo , Doenças Inflamatórias Intestinais/patologia , Interleucina-18/genética , Subunidade alfa de Receptor de Interleucina-18/genética , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mutação/genética , Fosforilação/efeitos dos fármacos , Receptores Tipo I de Interleucina-1/genética , Fator de Transcrição STAT3/genética , Organismos Livres de Patógenos Específicos , beta Catenina/genéticaRESUMO
Myeloid dendritic cells (DC) and macrophages evolve from a common precursor. However, factors controlling monocyte differentiation toward DC or macrophages are poorly defined. We report that the surface density of the GM-CSF receptor (GM-CSFR) alpha subunit in human peripheral blood monocytes varies among donors. Although no correlation was found between the extent of GM-CSFR and monocyte differentiation into DC driven by GM-CSF and IL-4, GM-CSFR expression strongly influenced the generation of CD1a(+) dendritic-like cells in the absence of IL-4. CD1a(+) cells generated in the presence of GM-CSF express CD40, CD80, MHC class I and II, DC-SIGN, MR, CCR5, and partially retain CD14 expression. Interestingly, they spontaneously induce the expansion of CD4(+) and CD8(+) allogeneic T lymphocytes producing IFN-gamma, and migrate toward CCL4 and CCL19. Upon stimulation with TLR ligands, they acquire the phenotypic features of mature DC. In contrast, the allostimulatory capacity is not further increased upon LPS activation. However, by blocking LPS-induced IL-10, a higher T cell proliferative response and IL-12 production were observed. Interestingly, IL-23 secretion was not affected by endogenous IL-10. These results highlight the importance of GM-CSFR expression in monocytes for cytokine-induced DC generation and point to GM-CSF as a direct player in the generation of functionally distinct DC.
Assuntos
Diferenciação Celular/fisiologia , Células Dendríticas/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/fisiologia , Monócitos/metabolismo , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Anticorpos/imunologia , Anticorpos/farmacologia , Antígenos CD/análise , Antígenos CD1/análise , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Diferenciação Celular/efeitos dos fármacos , Quimiocina CCL1/metabolismo , Quimiocina CCL19/farmacologia , Quimiocina CCL4/farmacologia , Quimiotaxia/efeitos dos fármacos , Citocinas/metabolismo , Células Dendríticas/citologia , Células Dendríticas/efeitos dos fármacos , Citometria de Fluxo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Humanos , Interferon gama/metabolismo , Interleucina-10/imunologia , Interleucina-10/metabolismo , Interleucina-12/metabolismo , Interleucina-4/farmacologia , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Lipopolissacarídeos/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Monócitos/citologia , Monócitos/efeitos dos fármacos , Receptores Toll-Like/agonistasRESUMO
We investigated the interactions between human monocyte-derived dendritic cells (DCs) and Ag-activated circulating TCR-gammadelta-expressing lymphocytes (Vdelta2). Coculture of immature DCs (iDCs) with peripheral blood Vdelta2 T cells activated with either pyrophosphomonoesters (isopentenyl pyrophosphate; IPP) or aminobiphosphonates (pamidronate; PAM) led to a significant up-modulation of CD86 and MHC class I molecules and to the acquisition of functional features typical of activated DCs. DC activation induced by both IPP- and PAM-stimulated gammadelta T cells was mostly mediated by TNF-alpha and IFN-gamma secreted by activated lymphocytes. However, the effect of PAM-activated gammadelta T cells, but not that of IPP-activated cells, required cell-to-cell contact. Reciprocally, activation of Vdelta2 T cells by PAM, but not by IPP, was dependent on cell contact with iDCs. In fact, when PAM-stimulated DC-gammadelta T cell cocultures were separated by a semipermeable membrane or treated with blocking anti-CD86 Abs, induction of CD25 and CD69 as well as IFN-gamma and TNF-alpha secretion by Vdelta2 cells were strongly reduced. These results demonstrate for the first time a bidirectional activating interaction between iDCs and PAM-stimulated gammadelta T lymphocytes, thus suggesting a potential adjuvant role of this early cross-talk in the therapeutic activity of aminobiphosphonate drugs.