RESUMO
Sirtuins are a unique class of NAD(+)-dependent deacetylases that regulate diverse biological functions such as aging, metabolism, and stress resistance. Recently, it has been shown that sirtuins may have anti-inflammatory activities by inhibiting proinflammatory transcription factors such as NF-κB. In contrast, we report in this study that pharmacological inhibition of sirtuins dampens adaptive Th2 responses and subsequent allergic inflammation by interfering with lung dendritic cell (DC) function in a mouse model of airway allergy. Using genetic engineering, we demonstrate that sirtuin 1 represses the activity of the nuclear receptor peroxisome proliferator-activated receptor-γ in DCs, thereby favoring their maturation toward a pro-Th2 phenotype. This study reveals a previously unappreciated function of sirtuin 1 in the regulation of DC function and Th2 responses, thus shedding new light on our current knowledge on the regulation of inflammatory processes by sirtuins.
Assuntos
Asma/imunologia , Células Dendríticas/imunologia , PPAR gama/antagonistas & inibidores , Sirtuína 1/fisiologia , Células Th2/imunologia , Animais , Asma/enzimologia , Asma/patologia , Hiper-Reatividade Brônquica/genética , Hiper-Reatividade Brônquica/imunologia , Hiper-Reatividade Brônquica/patologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Movimento Celular/genética , Movimento Celular/imunologia , Células Cultivadas , Células Dendríticas/metabolismo , Células Dendríticas/patologia , Feminino , Imunofenotipagem , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , PPAR gama/metabolismo , Sirtuína 1/antagonistas & inibidores , Células Th2/enzimologia , Células Th2/patologiaRESUMO
Asthma is a complex disease linked to various pathophysiological events including the activity of proteinases. The multifunctional A disintegrin and metalloproteinases (ADAMs) displaying the ability to cleave membrane-bound mediators or cytokines appear to be key mediators in various inflammatory processes. In the present study, we investigated ADAM-8 expression and production in a mouse model of allergen-induced airway inflammation. In allergen-exposed animals, increased expression of ADAM-8 was found in the lung parenchyma and in DC purified from the lungs. The potential role of ADAM-8 in the development of allergen-induced airway inflammation was further investigated by the use of an anti-ADAM-8 antibody and ADAM-8 knockout animals. We observed a decrease in allergen-induced acute inflammation both in BALF and the peribronchial area in anti-ADAM-8 antibody-treated mice and in ADAM-8-deficient mice (ADAM-8(-/-) ) after allergen exposure. ADAM-8 depletion led to a significant decrease of the CD11c(+) lung DC. We also report lower levels of CCL11 and CCL22 production in antibody-treated mice and ADAM-8- deficient mice that might be explained by decreased eosinophilic inflammation and lower numbers of DC, respectively. In conclusion, ADAM-8 appears to favour allergen-induced acute airway inflammation by promoting DC recruitment and CCL11 and CCL22 production.
Assuntos
Proteínas ADAM/metabolismo , Antígenos CD/metabolismo , Asma/metabolismo , Proteínas de Membrana/metabolismo , Proteínas ADAM/antagonistas & inibidores , Proteínas ADAM/genética , Proteínas ADAM/imunologia , Animais , Anticorpos/imunologia , Anticorpos/farmacologia , Anticorpos/uso terapêutico , Antígenos CD/genética , Antígenos CD/imunologia , Asma/imunologia , Asma/patologia , Líquido da Lavagem Broncoalveolar/citologia , Contagem de Células , Movimento Celular/genética , Movimento Celular/imunologia , Quimiocina CCL11/metabolismo , Quimiocina CCL22/metabolismo , Citocinas/metabolismo , Eosinófilos/metabolismo , Eosinófilos/patologia , Expressão Gênica/genética , Inflamação/imunologia , Inflamação/metabolismo , Inflamação/patologia , Inflamação/prevenção & controle , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/patologia , Macrófagos Alveolares/metabolismo , Macrófagos Alveolares/patologia , Masculino , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ovalbumina/imunologia , VacinaçãoRESUMO
BACKGROUND: Pattern-recognition receptors (PRRs) are critically involved in the pathophysiology of airway allergy, yet most of the signaling pathways downstream of PRRs implicated in allergic airway sensitization remain unknown. OBJECTIVE: We sought to study the effects of genetic depletion of interferon response factor (IRF) 3 and IRF7, important transcription factors downstream of various PRRs, in a murine model of house dust mite (HDM)-induced allergic asthma. METHODS: We compared HDM-induced allergic immune responses in IRF3-deficient (IRF3(-/-)), IRF7(-/-), and wild-type mice. RESULTS: Parameters of airway allergy caused by HDM exposure were strongly attenuated in IRF3(-/-), but not IRF7(-/-), mice compared with those in wild-type mice. Indeed, in HDM-exposed IRF3(-/-) mice HDM-specific T(H)2 cell responses did not develop. This correlated with impaired maturation and migration of IRF3(-/-) lung dendritic cells (DCs) on HDM treatment. Furthermore, adoptive transfer of HDM-loaded DCs indicated that IRF3(-/-) DCs had an intrinsic defect rendering them unable to migrate and to prime HDM-specific T(H)2 responses. Intriguingly, we also show that DC function and allergic airway sensitization in response to HDM were independent of signaling by type I interferons, the main target genes of IRF3. CONCLUSION: Through its role in DC function, IRF3, mainly known as a central activator of antiviral immunity, is essential for the development of T(H)2-type responses to airway allergens.
Assuntos
Antígenos de Dermatophagoides/imunologia , Fator Regulador 3 de Interferon/metabolismo , Pyroglyphidae/imunologia , Hipersensibilidade Respiratória/imunologia , Animais , Antígenos de Dermatophagoides/efeitos adversos , Asma/etiologia , Asma/imunologia , Células Dendríticas/imunologia , Modelos Animais de Doenças , Humanos , Fator Regulador 3 de Interferon/genética , Fator Regulador 7 de Interferon/genética , Fator Regulador 7 de Interferon/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Hipersensibilidade Respiratória/etiologia , Células Th2/imunologiaRESUMO
Dendritic cells (DCs) are professional APCs that have a unique capacity to initiate primary immune responses, including tolerogenic responses. We have genetically engineered bone marrow-derived DCs to express the immunosuppressive cytokine IL-10 and tested the ability of these cells to control experimental asthma. A single intratracheal injection of OVA-pulsed IL-10-transduced DCs (OVA-IL-10-DCs) to naive mice before OVA sensitization and challenge prevented all of the cardinal features of airway allergy, namely, eosinophilic airway inflammation, airway hyperreactivity, and production of mucus, Ag-specific Igs, and IL-4. OVA-IL-10-DCs also reversed established experimental asthma and had long-lasting and Ag-specific effects. We furthermore showed, by using IL-10-deficient mice, that host IL-10 is required for mediating the immunomodulatory effects of OVA-IL-10-DCs and demonstrated a significant increase in the percentage of OVA-specific CD4(+)CD25(+)Foxp3(+)IL-10(+) regulatory T cells in the mediastinal lymph nodes of OVA-IL-10-DC-injected mice. Finally, adoptive transfer of CD4(+) mediastinal lymph node T cells from mice injected with OVA-IL-10-DCs protected OVA-sensitized recipients from airway eosinophilia upon OVA provocation. Our study describes a promising strategy to induce long-lasting Ag-specific tolerance in airway allergy.
Assuntos
Asma/metabolismo , Células Dendríticas/metabolismo , Engenharia Genética/métodos , Tolerância Imunológica , Interleucina-10/metabolismo , Transferência Adotiva , Animais , Antígenos/imunologia , Apoptose/imunologia , Asma/imunologia , Asma/terapia , Células Cultivadas , Células Dendríticas/imunologia , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Imunoterapia/métodos , Interleucina-10/genética , Camundongos , Camundongos Knockout , Ovalbumina/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T Reguladores/imunologia , Transdução GenéticaRESUMO
The gene encoding the cytoskeletal regulator DIAPH3 is lost at high frequency in metastatic prostate cancer, and DIAPH3 silencing evokes a transition to an amoeboid tumor phenotype in multiple cell backgrounds. This amoeboid transformation is accompanied by increased tumor cell migration, invasion, and metastasis. DIAPH3 silencing also promotes the formation of atypically large (> 1 µm) membrane blebs that can be shed as extracellular vesicles (EV) containing bioactive cargo. Whether loss of DIAPH3 also stimulates the release of nano-sized EV (e.g., exosomes) is not established. Here we examined the mechanism of release and potential biological functions of EV shed from DIAPH3-silenced and other prostate cancer cells. We observed that stimulation of LNCaP cells with the prostate stroma-derived growth factor heparin-binding EGF-like growth factor (HB-EGF), combined with p38MAPK inhibition caused EV shedding, a process mediated by ERK1/2 hyperactivation. DIAPH3 silencing in DU145 cells also increased rates of EV production. EV isolated from DIAPH3-silenced cells activated AKT1 and androgen signaling, increased proliferation of recipient tumor cells, and suppressed proliferation of human macrophages and peripheral blood mononuclear cells. DU145 EV contained miR-125a, which suppressed AKT1 expression and proliferation in recipient human peripheral blood mononuclear cells and macrophages. Our findings suggest that EV produced as a result of DIAPH3 loss or growth factor stimulation may condition the tumor microenvironment through multiple mechanisms, including the proliferation of cancer cells and suppression of tumor-infiltrating immune cells.
Assuntos
Micropartículas Derivadas de Células/patologia , Neoplasias da Próstata/patologia , Microambiente Tumoral/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Androgênios/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Forminas , Humanos , Masculino , MicroRNAs/metabolismo , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Microambiente Tumoral/imunologiaRESUMO
We report that programmed death ligand 2 (PD-L2), a known ligand of PD-1, also binds to repulsive guidance molecule b (RGMb), which was originally identified in the nervous system as a co-receptor for bone morphogenetic proteins (BMPs). PD-L2 and BMP-2/4 bind to distinct sites on RGMb. Normal resting lung interstitial macrophages and alveolar epithelial cells express high levels of RGMb mRNA, whereas lung dendritic cells express PD-L2. Blockade of the RGMb-PD-L2 interaction markedly impaired the development of respiratory tolerance by interfering with the initial T cell expansion required for respiratory tolerance. Experiments with PD-L2-deficient mice showed that PD-L2 expression on non-T cells was critical for respiratory tolerance, but expression on T cells was not required. Because PD-L2 binds to both PD-1, which inhibits antitumor immunity, and to RGMb, which regulates respiratory immunity, targeting the PD-L2 pathway has therapeutic potential for asthma, cancer, and other immune-mediated disorders. Understanding this pathway may provide insights into how to optimally modulate the PD-1 pathway in cancer immunotherapy while minimizing adverse events.
Assuntos
Tolerância Imunológica/imunologia , Pulmão/imunologia , Proteínas do Tecido Nervoso/metabolismo , Proteína 2 Ligante de Morte Celular Programada 1/metabolismo , Animais , Proteína Morfogenética Óssea 2/metabolismo , Moléculas de Adesão Celular Neuronais , Linhagem Celular Tumoral , Células Epiteliais/metabolismo , Proteínas Ligadas por GPI , Humanos , Pulmão/metabolismo , Macrófagos Alveolares/metabolismo , Camundongos , Proteínas do Tecido Nervoso/imunologia , Proteína 2 Ligante de Morte Celular Programada 1/imunologia , Ligação ProteicaRESUMO
Staphylococcus (S.) aureus is a major pathogen involved in chronic bovine mastitis. Staphylococcal mastitis is difficult to control due to the ability of S. aureus to invade and survive within host cells. We therefore postulated that induction of CD8(+) cytotoxic T lymphocyte (CTL) responses leading to destruction of infected cells could help in the control of S. aureus mastitis. We demonstrate that immunization of mice with heat-killed S. aureus together with agonistic anti-CD40 monoclonal antibodies elicits strong CTL responses capable of reducing the severity of subsequent staphylococcal mastitis. Our study shows promise for CTL-dependent vaccination against S. aureus mastitis.
Assuntos
Antígenos CD40/imunologia , Linfócitos T CD8-Positivos/imunologia , Mastite/prevenção & controle , Infecções Estafilocócicas/prevenção & controle , Vacinas Antiestafilocócicas/imunologia , Staphylococcus aureus/imunologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/imunologia , Modelos Animais de Doenças , Mastite/imunologia , Mastite/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Infecções Estafilocócicas/imunologia , Infecções Estafilocócicas/patologia , Vacinas Antiestafilocócicas/administração & dosagem , Staphylococcus aureus/patogenicidade , Linfócitos T Citotóxicos/imunologia , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/imunologiaRESUMO
Aluminum-based adjuvants (aluminum salts or alum) are widely used in human vaccination, although their mechanisms of action are poorly understood. Here we report that, in mice, alum causes cell death and the subsequent release of host cell DNA, which acts as a potent endogenous immunostimulatory signal mediating alum adjuvant activity. Furthermore, we propose that host DNA signaling differentially regulates IgE and IgG1 production after alum-adjuvanted immunization. We suggest that, on the one hand, host DNA induces primary B cell responses, including IgG1 production, through interferon response factor 3 (Irf3)-independent mechanisms. On the other hand, we suggest that host DNA also stimulates 'canonical' T helper type 2 (T(H)2) responses, associated with IgE isotype switching and peripheral effector responses, through Irf3-dependent mechanisms. The finding that host DNA released from dying cells acts as a damage-associated molecular pattern that mediates alum adjuvant activity may increase our understanding of the mechanisms of action of current vaccines and help in the design of new adjuvants.
Assuntos
Adjuvantes Imunológicos/metabolismo , Linfócitos B/imunologia , DNA/metabolismo , Transdução de Sinais/imunologia , Células Th2/imunologia , Adjuvantes Imunológicos/farmacologia , Compostos de Alúmen/farmacologia , Animais , Morte Celular/efeitos dos fármacos , Desoxirribonuclease I , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Glicoproteínas/genética , Imunoglobulina E/metabolismo , Imunoglobulina G/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas de Ligação a RNA , Espectrofotometria , Receptor Toll-Like 9/genéticaRESUMO
The respiratory tract is continuously exposed to both innocuous airborne antigens and immunostimulatory molecules of microbial origin, such as LPS. At low concentrations, airborne LPS can induce a lung DC-driven Th2 cell response to harmless inhaled antigens, thereby promoting allergic asthma. However, only a small fraction of people exposed to environmental LPS develop allergic asthma. What prevents most people from mounting a lung DC-driven Th2 response upon exposure to LPS is not understood. Here we have shown that lung interstitial macrophages (IMs), a cell population with no previously described in vivo function, prevent induction of a Th2 response in mice challenged with LPS and an experimental harmless airborne antigen. IMs, but not alveolar macrophages, were found to produce high levels of IL-10 and to inhibit LPS-induced maturation and migration of DCs loaded with the experimental harmless airborne antigen in an IL-10-dependent manner. We further demonstrated that specific in vivo elimination of IMs led to overt asthmatic reactions to innocuous airborne antigens inhaled with low doses of LPS. This study has revealed a crucial role for IMs in maintaining immune homeostasis in the respiratory tract and provides an explanation for the paradox that although airborne LPS has the ability to promote the induction of Th2 responses by lung DCs, it does not provoke airway allergy under normal conditions.