RESUMO
BACKGROUND: Dermatomyositis (DM) is an autoimmune disease primarily affecting skin and muscle. OBJECTIVES: The purpose of this study was to determine whether an association exists between clinical skin disease activity as measured by the validated Cutaneous Dermatomyositis Disease Area and Severity Index (CDASI) and type 1 interferon (IFN) pathway biomarkers in the blood of patients with DM. METHODS: Forty-two patients with DM and 25 healthy volunteers were prospectively enrolled. CDASI scores were obtained, and serum and blood RNA were isolated from all participants. Associations between CDASI activity and type 1 IFN-inducible gene signature were assessed cross-sectionally in all patient samples and longitudinally on 13 paired visits via transcriptional profiling analyses. RESULTS: By RNAseq analysis, type 1 IFN-inducible genes were the most highly differentially regulated. A CDASI activity threshold of 12 was correlated with an elevated type 1 IFN gene signature and with serum IFN-ß, but not with IFN-α protein. Expression analysis showed that all patients with mild disease activity had a low type 1 IFN gene signature, while 93% of patients with moderate-to-high disease activity had elevated gene signature. In longitudinal analysis, changes in CDASI activity showed nonsignificant trends with concordant directional changes in gene signature. CONCLUSIONS: A type 1 IFN pathway signature biomarker in blood is highly correlated with CDASI activity scores in DM, and may be a promising surrogate clinical trial end point. The correlation of serum IFN-ß, but not IFN-α, with both a gene signature and CDASI suggests that IFN-ß drives disease activity in DM.
Assuntos
Dermatomiosite/genética , Interferon Tipo I/genética , Interferon beta/genética , Biomarcadores/metabolismo , Quimiocina CXCL10/metabolismo , Estudos Transversais , Dermatomiosite/sangue , Feminino , Voluntários Saudáveis , Humanos , Interferon Tipo I/metabolismo , Interferon beta/metabolismo , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , RNA Mensageiro/metabolismo , Índice de Gravidade de DoençaRESUMO
A cancer diagnosis is often associated with loss of agency and control that can adversely affect well-being. Patients may try to regain control through dietary change aimed at preventing progression and/or recurrence. Evidence for the effectiveness of post-diagnostic dietary change in prostate cancer is not conclusive, which can cause uncertainty among patients and health professionals. This qualitative study explored how eight men in the U.K., who had been diagnosed with prostate cancer in the previous 5 years, accounted for any post-diagnostic changes they made to their diet. Data were generated through semi-structured telephone interviews and were subjected to thematic analysis. This yielded two themes concerning the perceived nature and importance of dietary change and the perceived determinants of dietary change. The latter focused on internal dimensions such as agency and external dimensions such as the perceived role of relationships with health professionals, the availability of credible dietary information and family influences. The study points to the importance of the family context in enabling the men to implement dietary change. It is suggested that, even if health professionals can only offer qualified, general advice about diet, this may provide men with a focus for action and a means of regaining control.
Assuntos
Dietoterapia/psicologia , Dieta/psicologia , Neoplasias da Próstata/psicologia , Idoso , Atitude Frente a Saúde , Humanos , Masculino , Pessoa de Meia-Idade , Percepção , Neoplasias da Próstata/diagnóstico , Pesquisa Qualitativa , Reino UnidoRESUMO
We investigated the role of granulocyte-macrophage colony-stimulating factor (GM-CSF) in a subchronic exposure model of cigarette smoke (CS)-induced inflammation using antibodies directed against GM-CSF or the GM-CSF receptor (GM-CSFR) α-chain. CS-induced mononuclear and neutrophilic inflammation following 4 days of CS exposure in BALB/c mice was assessed in bronchoalveolar lavage (BAL) fluid. An increase in mature dendritic cells (DCs) (CD11c+ and major histocompatibility complex II+) and Gr-1-high neutrophils was also observed by flow cytometric analysis of whole-lung tissue. Daily i.p. injection of 400 µg GM-CSF or GM-CSFR antibody prior to daily smoke exposure attenuated the accumulation of neutrophils within the BAL by 60%. A reduction in mature DCs was also observed. Anti-GM-CSFR antibody administration did not have an effect on the percentage of lung T-cells; however, a significant decrease in activated CD69+ CD8+ T-cells was observed. Anti-GM-CSFR antibody administration decreased the mRNA and protein expression of interleukin-12 p40 and matrix metalloproteinase 12. Taken together, intervention with this receptor antibody implicates the GM-CSF pathway as an important mediator of smoke-induced inflammation.
Assuntos
Anticorpos/imunologia , Neutrófilos/imunologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/imunologia , Fumar/imunologia , Animais , Antígenos CD/imunologia , Antígenos de Diferenciação de Linfócitos T/imunologia , Líquido da Lavagem Broncoalveolar/imunologia , Antígeno CD11c/imunologia , Linfócitos T CD8-Positivos/imunologia , Células Dendríticas/imunologia , Feminino , Genes MHC da Classe II/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Subunidade p40 da Interleucina-12/biossíntese , Subunidade p40 da Interleucina-12/imunologia , Lectinas Tipo C/imunologia , Pulmão/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Receptores de Quimiocinas/imunologia , Receptores de Interleucina-1/genética , Receptores de Interleucina-1/imunologiaRESUMO
Interleukin (IL)-2 and IL-15 are redundant in stimulating T-cell proliferation in vitro. Their precise role in vivo in governing T-cell expansion and T-cell homeostasis is less clear. Each may have distinct functions and regulate distinct aspects of T-cell activation. The functional receptors for IL-2 and IL-15 consist of a private alpha-chain, which defines the binding specificity for IL-2 or IL-15, and shared IL-2 receptor beta- and gamma-chains. The gamma-chain is also a critical signaling component of IL-4, IL-7 and IL-9 receptors. Thus, the gamma-chain is called the common gamma or gamma-c. As these receptor subunits can be expressed individually or in various combinations resulting in the formation of receptors with different affinities, distinct signaling capabilities or both, we hypothesized that differential expression of IL-2 and IL-15 receptor subunits on cycling T cells in vivo may direct activated T cells to respond to IL-2 or IL-15, thereby regulating the homeostasis of T-cell response in vivo. By observing in vivo T-cell divisions and expression of IL-2 and IL-15 receptor subunits, we demonstrate that IL-15 is a critical growth factor in initiating T cell divisions in vivo, whereas IL-2 limits continued T-cell expansion via downregulation of the gamma-c expression. Decreased gamma-c expression on cycling T cells reduced sustained Bcl-2 expression and rendered cells susceptible to apoptotic cell death. Our study provides data that IL-2 and IL-15 regulate distinct aspects of primary T-cell expansion in vivo.
Assuntos
Apoptose/fisiologia , Interleucina-15/fisiologia , Interleucina-2/fisiologia , Linfócitos T/citologia , Animais , Ativação Linfocitária/fisiologia , CamundongosRESUMO
The recruitment of eosinophils into the airways after allergen exposure is dependent on interleukin (IL) 5 secreted from antigen-specific CD4+ T cells of the T helper cell (Th) 2 subset. However, while it is established that costimulation through CD28 is required for TCR-mediated activation and IL-2 production, the importance of this mechanism for the induction of a Th2 immune response is less clear. In the present study, we administered the fusion protein CTLA-4 immunoglobulin (Ig) into the lungs before allergen provocation to determine whether CD28/CTLA-4 ligands are required for allergen-induced eosinophil accumulation and the production of Th2 cytokines. Administration of CTLA-4 Ig inhibited the recruitment of eosinophils into the lungs by 75% and suppressed IgE in the bronchoalveolar lavage fluid. CTLA-4 Ig also inhibited the production of IL-4, IL-5, and IL-10 by 70-80% and enhanced interferon-gamma production from CD3-T cell receptor-activated lung Thy1.2+ cells. Allergen exposure upregulated expression of B7-2, but not B7-1, on B cells from the lung within 24 h. Moreover, airway administration of an anti-B7-2 monoclonal antibody (mAb) inhibited eosinophil infiltration, IgE production, and Th2 cytokine secretion comparable in magnitude to that observed with CTLA-4 Ig. Treatment with an anti-B7-1 mAb had a small, but significant effect on eosinophil accumulation, although was less effective in inhibiting Th2 cytokine production. The anti-B7-2, but not anti-B7-1, mAb also inhibited antigen-induced airway hyperresponsiveness in vivo. In all of the parameters assessed, the combination of both the anti-B7-1 and anti-B7-2 mAb was no more effective than anti-B7-2 mAb treatment alone. We propose that strategies aimed at inhibition of CD28 interactions with B7-2 molecules may represent a novel therapeutic target for the treatment of lung mucosal allergic inflammation.
Assuntos
Antígenos CD/fisiologia , Imunidade nas Mucosas , Imunoconjugados , Pulmão/imunologia , Glicoproteínas de Membrana/fisiologia , Células Th2/imunologia , Abatacepte , Alérgenos , Animais , Antígenos de Diferenciação/farmacologia , Antígeno B7-1/fisiologia , Antígeno B7-2 , Antígenos CD28/fisiologia , Antígeno CTLA-4 , Células Cultivadas , Quimiotaxia de Leucócito , Citocinas/biossíntese , Eosinófilos/imunologia , Imunoglobulina E/metabolismo , Interferon gama/biossíntese , Camundongos , Ventilação Pulmonar , Regulação para CimaRESUMO
Elevated levels of immunoglobulin (Ig) E are associated with bronchial asthma, a disease characterized by eosinophilic inflammation of the airways. Activation of antigen-specific T helper (Th) 2 cells in the lung with the subsequent release of interleukin (IL) 4 and IL-5 is believed to play an important role in the pathogenesis of this disease. In this study, we have used a non-anaphylactogenic anti-mouse-IgE antibody to investigate the relationship between IgE, airway eosinophil infiltration, and the production of Th2 cytokines. Immunization of mice with house dust mite antigen increased serum levels of IgE and IgG. Antigen challenge of immunized but not control mice induced an infiltration of eosinophils in the bronchoalveolar lavage associated with the production of IL-4 and IL-5 from lung purified Thy1.2+ cells activated through the CD3-T cell receptor complex. Administration of the anti-IgE monoclonal antibody (mAb) 6h before antigen challenge neutralized serum IgE but not IgG and inhibited the recruitment of eosinophils into the lungs and the production of IL-4 and IL-5 but not interferon gamma. Studies performed using an anti-CD23 mAb, CD23 deficient and mast cell deficient mice suggest that anti-IgE mAb suppresses eosinophil infiltration and Th2 cytokine production by inhibiting IgE-CD23-facilitated antigen presentation to T cells. Our results demonstrate that IgE-dependent mechanisms are important in the induction of a Th2 immune response and the subsequent infiltration of eosinophils into the airways. Neutralization of IgE, for example, non-anaphylactogenic anti-IgE mAbs may provide a novel therapeutic approach to the treatment of allergic airway disease.
Assuntos
Asma/imunologia , Eosinófilos/imunologia , Imunoglobulina E/imunologia , Pulmão/imunologia , Células Th2/imunologia , Anafilaxia/imunologia , Animais , Anticorpos Monoclonais/farmacologia , Movimento Celular , Citocinas/biossíntese , Inflamação/imunologia , Pulmão/citologia , Mastócitos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Receptores de IgERESUMO
Virus infections of the lung are thought to predispose individuals to asthma, a disease characterized by eosinophil infiltration of the airways. CD8+ T cells are an important part of the host response to virus infection, however, they have no reported role in eosinophil recruitment. We developed a mouse model of virus peptide-stimulated CD8+ T cell immune responses in the lung. We found that bystander CD4+ T helper cell type 2 immune responses to ovalbumin switched the virus peptide-specific CD8+ T cells in the lung to interleukin (IL) 5 production. Furthermore, when such IL-5-producing CD8 T cells were challenged via the airways with virus peptide, a significant eosinophil infiltration was induced. In vitro studies indicated that IL-4 could switch the virus-specific CD8+ T cells to IL-5 production. These results could explain the link between virus infection and acute exacerbation of asthma and, perhaps more importantly, they indicate an IL-4-dependent mechanism that would impair CD8+ T cell responses and delay viral clearance from the host.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Eosinofilia/etiologia , Interleucina-5/biossíntese , Pulmão/imunologia , Vírus da Coriomeningite Linfocítica/imunologia , Animais , Imunização , Interleucina-4/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina/imunologia , Células Th2/imunologiaRESUMO
T cells secreting interleukin (IL)-4 and IL-5 (T helper cell type 2 [Th2] cells) play a detrimental role in a variety of diseases, but specific methods of regulating their activity remain elusive. T1/ST2 is a surface ligand of the IL-1 receptor family, expressed on Th2- but not on interferon (IFN)-gamma-producing Th1 cells. Prior exposure of BALB/c mice to the attachment (G) or fusion (F) protein of respiratory syncytial virus (RSV) increases illness severity during intranasal RSV challenge, due to Th2-driven lung eosinophilia and exuberant Th1-driven pulmonary infiltration, respectively. We used these polar models of viral illness to study the recruitment of T1/ST2 cells to the lung and to test the effects of anti-T1/ST2 treatment in vivo. T1/ST2 was present on a subset of CD4(+) cells from mice with eosinophilic lung disease. Monoclonal anti-T1/ST2 treatment reduced lung inflammation and the severity of illness in mice with Th2 (but not Th1) immunopathology. These results show that inhibition of T1/ST2 has a specific effect on virally induced Th2 responses and suggests that therapy targeted at this receptor might be of value in treating Th2-driven illness.
Assuntos
Proteínas de Membrana , Proteínas/imunologia , Receptores de Interleucina-1/imunologia , Infecções por Vírus Respiratório Sincicial/imunologia , Linfócitos T Auxiliares-Indutores , Animais , Eosinofilia/imunologia , Eosinofilia/terapia , Feminino , Proteína 1 Semelhante a Receptor de Interleucina-1 , Camundongos , Camundongos Endogâmicos BALB C , Receptores de Interleucina , Infecções por Vírus Respiratório Sincicial/terapia , Células Th1 , Células Th2RESUMO
It has been shown that certain pathogens can trigger efficient T cell responses in the absence of CD28, a key costimulatory receptor expressed on resting T cells. Inducible costimulator protein (ICOS) is an inducible costimulator structurally and functionally related to CD28. Here, we show that in the absence of CD28 both T helper cell type 1 (Th1) and Th2 responses were impaired but not abrogated after infection with lymphocytic choriomeningitis virus (LCMV), vesicular stomatitis virus (VSV), and the nematode Nippostrongylus brasiliensis. Inhibition of ICOS in CD28-deficient mice further reduced Th1/Th2 polarization. Blocking of ICOS alone had a limited but significant capacity to downregulate Th subset development. In contrast, cytotoxic T lymphocyte (CTL) responses, which are regulated to a minor and major extent by CD28 after LCMV and VSV infection, respectively, remained unaffected by blocking ICOS. Together, our results demonstrate that ICOS regulates both CD28-dependent and CD28-independent CD4(+) subset (Th1 and Th2) responses but not CTL responses in vivo.
Assuntos
Antígenos de Diferenciação de Linfócitos T/fisiologia , Antígenos CD28/fisiologia , Vírus da Coriomeningite Linfocítica/imunologia , Nippostrongylus/imunologia , Células Th1/imunologia , Células Th2/imunologia , Vírus da Estomatite Vesicular Indiana/imunologia , Animais , Antígenos CD28/genética , Polaridade Celular , Imunoglobulina G/imunologia , Isotipos de Imunoglobulinas/imunologia , Proteína Coestimuladora de Linfócitos T Induzíveis , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/parasitologia , Subpopulações de Linfócitos T/virologia , Células Th1/citologia , Células Th1/parasitologia , Células Th1/virologia , Células Th2/citologia , Células Th2/parasitologia , Células Th2/virologiaRESUMO
Isolated peripheral blood CD4 cells from allergic individuals express CC chemokine receptor (CCR)3 and CCR4 after expansion in vitro. In addition, human T helper type 2 (Th2) cells polarized in vitro selectively express CCR3 and CCR4 at certain stages of activation/differentiation and respond preferentially to the ligands eotaxin and monocyte-derived chemokine (MDC). However, controversy arises when the in vivo significance of this distinct expression is discussed. To address the functional role of CCR3/eotaxin and CCR4/MDC during the in vivo recruitment of Th2 cells, we have transferred effector Th cells into naive mice to induce allergic airway disease. Tracking of these cells after repeated antigen challenge has established that both CCR3/eotaxin and CCR4/MDC axes contribute to the recruitment of Th2 cells to the lung, demonstrating the in vivo relevance of the expression of these receptors on Th2 cells. We have shown that involvement of the CCR3/eotaxin pathway is confined to early stages of the response in vivo, whereas repeated antigen stimulation results in the predominant use of the CCR4/MDC pathway. We propose that effector Th2 cells respond to both CCR3/eotaxin and CCR4/MDC pathways initially, but that a progressive increase in CCR4-positive cells results in the predominance of the CCR4/MDC axis in the long-term recruitment of Th2 cells in vivo.
Assuntos
Pulmão/imunologia , Receptores de Quimiocinas/imunologia , Células Th2/imunologia , Transferência Adotiva , Alérgenos/imunologia , Animais , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Células Cultivadas , Feminino , Expressão Gênica , Humanos , Pulmão/citologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Ovalbumina/imunologia , Receptores CCR3 , Receptores CCR4 , Receptores de Quimiocinas/genética , Receptores de Quimiocinas/metabolismo , Células Th1/imunologia , Células Th2/metabolismoRESUMO
The complex pathophysiology of lung allergic inflammation and bronchial hyperresponsiveness (BHR) that characterize asthma is achieved by the regulated accumulation and activation of different leukocyte subsets in the lung. The development and maintenance of these processes correlate with the coordinated production of chemokines. Here, we have assessed the role that different chemokines play in lung allergic inflammation and BHR by blocking their activities in vivo. Our results show that blockage of each one of these chemokines reduces both lung leukocyte infiltration and BHR in a substantially different way. Thus, eotaxin neutralization reduces specifically BHR and lung eosinophilia transiently after each antigen exposure. Monocyte chemoattractant protein (MCP)-5 neutralization abolishes BHR not by affecting the accumulation of inflammatory leukocytes in the airways, but rather by altering the trafficking of the eosinophils and other leukocytes through the lung interstitium. Neutralization of RANTES (regulated upon activation, normal T cell expressed and secreted) receptor(s) with a receptor antagonist decreases significantly lymphocyte and eosinophil infiltration as well as mRNA expression of eotaxin and RANTES. In contrast, neutralization of one of the ligands for RANTES receptors, macrophage-inflammatory protein 1alpha, reduces only slightly lung eosinophilia and BHR. Finally, MCP-1 neutralization diminishes drastically BHR and inflammation, and this correlates with a pronounced decrease in monocyte- and lymphocyte-derived inflammatory mediators. These results suggest that different chemokines activate different cellular and molecular pathways that in a coordinated fashion contribute to the complex pathophysiology of asthma, and that their individual blockage results in intervention at different levels of these processes.
Assuntos
Quimiocinas CC/fisiologia , Hipersensibilidade/imunologia , Inflamação/imunologia , Pulmão/imunologia , Animais , Anticorpos/imunologia , Asma/fisiopatologia , Quimiocina CCL11 , Quimiocina CCL4 , Quimiocina CCL5/farmacologia , Quimiocinas CC/antagonistas & inibidores , Fatores Quimiotáticos de Eosinófilos/farmacologia , Citocinas/farmacologia , Modelos Animais de Doenças , Imuno-Histoquímica , Leucócitos/efeitos dos fármacos , Leucócitos/metabolismo , Pulmão/citologia , Proteínas Inflamatórias de Macrófagos/farmacologia , Camundongos , Camundongos Endogâmicos , Proteínas Quimioatraentes de Monócitos/farmacologia , Ovalbumina/imunologia , RNA Mensageiro/metabolismoRESUMO
T1/ST2 is an orphan receptor of unknown function that is expressed on the surface of murine T helper cell type 2 (Th2), but not Th1 effector cells. In vitro blockade of T1/ST2 signaling with an immunoglobulin (Ig) fusion protein suppresses both differentiation to and activation of Th2, but not Th1 effector populations. In a nascent Th2-dominated response, anti-T1/ST2 monoclonal antibody (mAb) inhibited eosinophil infiltration, interleukin 5 secretion, and IgE production. To determine if these effects were mediated by a direct effect on Th2 cells, we next used a murine adoptive transfer model of Th1- and Th2-mediated lung mucosal immune responses. Administration of either T1/ST2 mAb or T1/ST2-Ig abrogated Th2 cytokine production in vivo and the induction of an eosinophilic inflammatory response, but failed to modify Th1-mediated inflammation. Taken together, our data demonstrate an important role of T1/ST2 in Th2-mediated inflammatory responses and suggest that T1/ST2 may prove to be a novel target for the selective suppression of Th2 immune responses.
Assuntos
Imunidade nas Mucosas , Pulmão/imunologia , Proteínas de Membrana , Proteínas/fisiologia , Receptores de Interleucina-1/fisiologia , Mucosa Respiratória/imunologia , Células Th2/imunologia , Alérgenos , Animais , Anticorpos Monoclonais/farmacologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Células COS , Diferenciação Celular , Humanos , Interferon gama/biossíntese , Proteína 1 Semelhante a Receptor de Interleucina-1 , Interleucinas/biossíntese , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Proteínas/imunologia , Receptores de Superfície Celular , Receptores de Interleucina , Receptores de Interleucina-1/imunologia , Proteínas Recombinantes de Fusão/imunologia , Mucosa Respiratória/patologia , Transdução de Sinais , Células Th1/citologia , Células Th1/imunologia , Células Th1/patologia , Células Th2/patologia , TransfecçãoRESUMO
BACKGROUND: As Th2 type lymphocytes orchestrate the cardinal features of allergic asthma, inhibiting their recruitment to the lungs could be of therapeutic benefit. Although human Th2 cells express the CCR4 chemokine receptor and increased production of CCR4 ligands has been found in asthmatic airways, studies in animals have reached contradictory conclusions on whether blocking this pathway would be beneficial. OBJECTIVE: As a lack of efficacy might be due to differences between mouse and man, we readdressed this question using a humanized severe combined immunodeficiency model of asthma. METHODS: Mice received peripheral blood mononuclear cells from house dust mite (HDM) allergic asthmatic patients and then underwent bronchial challenge with HDM. RESULTS: This resulted in marked allergic inflammation and bronchial hyper-reactivity. Administration of CCR4 blocking antibody abolished the airway eosinophilia, goblet cell hyperplasia, IgE synthesis and bronchial hyper-reactivity. In this chimeric system, human CD11c+ dendritic cells (DCs) were the predominant source of CCR4 ligands, suggesting that DC-derived chemokines attract Th2 cells. In separate experiments using human DCs, in vitro exposure to HDM of DCs from HDM allergic patients but not healthy controls caused CCL17 and CCL22 release that resulted in chemoattraction of polarized human Th2 cells in a CCR4-dependent way. CONCLUSIONS: Taken together, our data provide proof of concept that CCR4 blockade inhibits the salient features of asthma and justify further clinical development of CCR4 antagonists for this disease.
Assuntos
Asma/imunologia , Células Dendríticas/imunologia , Inflamação/imunologia , Receptores CCR4/imunologia , Células Th2/imunologia , Animais , Anticorpos/imunologia , Asma/metabolismo , Asma/patologia , Asma/prevenção & controle , Quimiocina CCL17/imunologia , Quimiocina CCL17/metabolismo , Quimiocina CCL22/imunologia , Quimiocina CCL22/metabolismo , Células Dendríticas/metabolismo , Modelos Animais de Doenças , Humanos , Inflamação/metabolismo , Inflamação/prevenção & controle , Pulmão/imunologia , Pulmão/patologia , Camundongos , Camundongos SCID , Pyroglyphidae/imunologia , Receptores CCR4/antagonistas & inibidores , Células Th2/metabolismoRESUMO
Nociceptive information is relayed through the spinal cord dorsal horn, a critical area in sensory processing. The neuronal circuits in this region that underpin sensory perception must be clarified to better understand how dysfunction can lead to pathological pain. This study used an optogenetic approach to selectively activate spinal interneurons that express the calcium-binding protein calretinin (CR). We show that these interneurons form an interconnected network that can initiate and sustain enhanced excitatory signaling, and directly relay signals to lamina I projection neurons. Photoactivation of CR interneurons in vivo resulted in a significant nocifensive behavior that was morphine sensitive, caused a conditioned place aversion, and was enhanced by spared nerve injury. Furthermore, halorhodopsin-mediated inhibition of these interneurons elevated sensory thresholds. Our results suggest that dorsal horn circuits that involve excitatory CR neurons are important for the generation and amplification of pain and identify these interneurons as a future analgesic target.
Assuntos
Calbindina 2/genética , Interneurônios/metabolismo , Neuralgia/fisiopatologia , Neurônios/metabolismo , Corno Dorsal da Medula Espinal/metabolismo , Analgésicos Opioides/farmacologia , Animais , Calbindina 2/metabolismo , Modelos Animais de Doenças , Expressão Gênica , Halorrodopsinas/genética , Halorrodopsinas/metabolismo , Interneurônios/efeitos dos fármacos , Interneurônios/patologia , Camundongos , Camundongos Transgênicos , Morfina/farmacologia , Rede Nervosa/efeitos dos fármacos , Rede Nervosa/metabolismo , Rede Nervosa/patologia , Neuralgia/tratamento farmacológico , Neuralgia/genética , Neuralgia/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/patologia , Optogenética/métodos , Limiar da Dor/efeitos dos fármacos , Técnicas de Patch-Clamp , Estimulação Luminosa , Corno Dorsal da Medula Espinal/efeitos dos fármacos , Corno Dorsal da Medula Espinal/patologia , Técnicas de Cultura de Tecidos , TransgenesRESUMO
We have previously reported that human eosinophil granule major basic protein and synthetic cationic proteins such as poly-L-arginine and poly-L-lysine, can increase airway responsiveness in vivo. In the present study, we have investigated whether activation of sensory C-fibers is important in this phenomenon. Dose-response curves to methacholine were constructed before and 1 h after intratracheal instillation of poly-L-lysine in anaesthetized spontaneously breathing rats, and the concentration of methacholine required to induce a doubling in total lung resistance was calculated. Poly-L-lysine induced a fivefold increase in airway responsiveness, which was inhibited by neonatal capsaicin treatment and potentiated by phosphoramidon (100 micrograms/ml). Furthermore, pretreatment with either CP, 96-345, or RP-67580 two selective NK-1 receptor antagonists inhibited poly-L-lysine-induced airway hyperresponsiveness and plasma protein extravasation. In vitro, cationic proteins stimulated the release of calcitonin gene-related peptide-like immunoreactivity from perfused slices of the main bronchi. Our results demonstrate that cationic proteins can activate sensory C-fibers in the airways, an effect which is important in the subsequent development of airway hyperresponsiveness and plasma protein extravasation. Cationic proteins may therefore function as a link between inflammatory cell accumulation and sensory nerve activation.
Assuntos
Brônquios/fisiologia , Neurônios Aferentes/fisiologia , Peptídeos/farmacologia , Transdução de Sinais/fisiologia , Substância P/fisiologia , Animais , Proteínas Sanguíneas/metabolismo , Brônquios/efeitos dos fármacos , Brônquios/inervação , Peptídeo Relacionado com Gene de Calcitonina/análise , Permeabilidade Capilar/fisiologia , Capsaicina/farmacologia , Cátions/farmacologia , Feminino , Glicopeptídeos/farmacologia , Masculino , Cloreto de Metacolina/farmacologia , Antagonistas dos Receptores de Neurocinina-1 , Neurônios Aferentes/efeitos dos fármacos , Polilisina/farmacologia , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacosRESUMO
In the current series of experiments we investigated the role of bradykinin in airway hyperresponsiveness induced by human eosinophil-granule major basic protein (MBP). Bronchoalveolar lavage was performed after intratracheal instillation of MBP or poly-L-lysine in anesthetized, intubated rats, and levels of immunoreactive kinins and kallikrein-like activity were determined. Both MBP and poly-L-lysine induced a three- and eightfold increase in levels of kallikrein-like activity and i-kinins, respectively. To determine whether kinin production is required for the development of airway hyperresponsiveness induced by cationic proteins, dose-response curves to methacholine were constructed before and 1 h after intratracheal instillation of either MBP or poly-L-lysine (100 micrograms). MBP and poly-L-lysine induced an increase in airway responsiveness, which was inhibited by pretreatment with a selective BK-2 receptor antagonist, NPC 17713 (250 micrograms/ml). Our results demonstrate that MBP and poly-L-lysine activate kallikrein and stimulate the generation of i-kinins in vivo, an effect that may be related to the cationic charge of these proteins. Furthermore, the ability of these proteins to increase airway responsiveness appears to be dependent on the generation of i-kinins.
Assuntos
Proteínas Sanguíneas/farmacologia , Bradicinina/fisiologia , Hipersensibilidade/imunologia , Mediadores da Inflamação/farmacologia , Sistema Respiratório/imunologia , Ribonucleases , Sequência de Aminoácidos , Animais , Bradicinina/biossíntese , Broncoconstrição/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ativação Enzimática , Proteínas Granulares de Eosinófilos , Eosinófilos/imunologia , Feminino , Humanos , Calicreínas/biossíntese , Masculino , Cloreto de Metacolina/farmacologia , Dados de Sequência Molecular , Oligopeptídeos/farmacologia , Poliaminas/farmacologia , Polieletrólitos , Polilisina/farmacologia , Ratos , Ratos Sprague-DawleyRESUMO
The aim of this study was to investigate whether dendritic cells (DCs) can induce sensitization to aeroallergen in a mouse model of allergic asthma. Ovalbumin-pulsed (OVA-pulsed) or unpulsed myeloid DCs that were injected into the airways of naive mice migrated into the mediastinal lymph nodes. When challenged 2 weeks later with an aerosol of OVA, activated CD4 and CD8 lymphocytes, eosinophils, and neutrophils were recruited to the lungs of actively immunized mice. These CD4(+) lymphocytes produced predominantly IL-4 and IL-5 but also IFN-gamma, whereas CD8(+) lymphocytes produced predominantly IFN-gamma. Histological analysis revealed perivascular and peribronchial eosinophilic infiltrates and goblet cell hyperplasia. Studies in IL-4(-/-) and CD28(-/-) mice revealed that production of IL-4 by host cells and provision of costimulation to T cells by DCs were critical for inducing the response. Lung CD4(+) T cells strongly expressed the Th2 marker T1/ST2, and signaling through this molecule via a ligand expressed on DCs was essential for the establishment of airway eosinophilia. These data demonstrate that DCs in the airways induce sensitization to inhaled antigen and that molecules expressed on the surface of these cells are critical for the development of Th2-dependent airway eosinophilia.
Assuntos
Antígenos/administração & dosagem , Asma/etiologia , Células Dendríticas/imunologia , Eosinofilia Pulmonar/etiologia , Células Th2/imunologia , Administração por Inalação , Alérgenos/administração & dosagem , Animais , Asma/imunologia , Asma/patologia , Antígenos CD28/genética , Antígenos CD28/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Interleucina-4/genética , Interleucina-4/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Mutantes , Ovalbumina/administração & dosagem , Ovalbumina/imunologia , Eosinofilia Pulmonar/imunologia , Eosinofilia Pulmonar/patologia , Transdução de SinaisRESUMO
While considerable progress has been made in understanding the events by which eosinophils accumulate in various pathophysiological conditions, the mechanisms controlling the resolution of eosinophilic inflammation are poorly understood. In the present study, we demonstrate that lung eosinophils obtained by bronchoalveolar lavage (BAL) after aerosol allergen provocation of immunized mice expressed the Fas receptor. Stimulation of purified eosinophils in vitro with a monoclonal anti-Fas mAb (1 ng-1 microg/ml) induced a dose/time dependent loss of cell viability from 24-72 h. Measurement of DNA fragmentation with propidium iodide confirmed that anti-Fas induced eosinophil death by apoptosis. While incubation with IL-3, IL-5, or GM-CSF prevented spontaneous apoptosis, these factors failed to prevent anti-Fas induced apoptosis. Administration of anti-Fas mAb to the lungs after the induction of a lung eosinophilia increased the number of peroxidase positive macrophages in BAL fluid 4-12 h later which was followed by a marked reduction in the number of eosinophils in the airways. Importantly, Fas-mediated resolution of eosinophilic inflammation occurred in the absence of any overt secondary inflammatory changes in the lungs. We speculate that defects in this pathway may at least in part explain the chronic eosinophilic inflammation often observed in the lungs of asthmatic individuals.
Assuntos
Apoptose , Eosinófilos/imunologia , Pulmão/imunologia , Receptor fas/fisiologia , Animais , Anticorpos Monoclonais/farmacologia , Separação Celular , Sobrevivência Celular , Células Cultivadas , Eosinófilos/citologia , Eosinófilos/patologia , Citometria de Fluxo , Inflamação , Cinética , Pulmão/patologia , Camundongos , Camundongos Endogâmicos , Fatores de TempoRESUMO
The objective of this study was to investigate the contribution of secondary lymphoid organs in the generation and maintenance of experimental allergic airway inflammation. We employed a previously reported murine model of respiratory mucosal allergic sensitization, induced by repeated aerosolizations of ovalbumin in the context of a GM-CSF airway environment. We executed this protocol in wild-type (WT) and lymphotoxin-alpha-deficient mice (LTalpha-KO) mice, which are devoid of lymph nodes (LNs) and possess rudimentary spleen structures. Despite the lack of pulmonary LNs draining the airway compartment, LTalpha-KO mice were fully capable of mounting a robust inflammatory response in the airways, consisting of Th2 polarized CD4+ T cells and eosinophils. This was accompanied by IL-5, IL-13, and IFN-gamma production by splenocytes and generation of ovalbumin-specific serum IgE. Exposure to the same antigen 7 weeks after complete resolution of airway inflammation once again induced a Th2 polarized infiltrate, demonstrating intact immunological memory. To investigate inherent plasticity in establishing antigen-specific immunity, mice were splenectomized before sensitization. Allergic sensitization was completely abrogated in splenectomized LTalpha-KO mice, compared with eusplenic LTalpha-KO controls. These data demonstrate that secondary lymphoid organs, either LN or spleen, are essential for the generation of allergic airway responses.
Assuntos
Modelos Animais de Doenças , Linfonodos/imunologia , Hipersensibilidade Respiratória/imunologia , Baço/imunologia , Administração Intranasal , Animais , Líquido da Lavagem Broncoalveolar , Linfócitos T CD8-Positivos/imunologia , Células Cultivadas , Eosinófilos/imunologia , Vetores Genéticos/administração & dosagem , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Fator Estimulador de Colônias de Granulócitos e Macrófagos/fisiologia , Imunoglobulina E/biossíntese , Memória Imunológica , Inflamação , Interferon gama/biossíntese , Interleucina-13/biossíntese , Interleucina-5/biossíntese , Linfonodos/anormalidades , Linfotoxina-alfa/genética , Linfotoxina-alfa/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ovalbumina/toxicidade , Eosinofilia Pulmonar/etiologia , Eosinofilia Pulmonar/imunologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/fisiologia , Hipersensibilidade Respiratória/etiologia , Organismos Livres de Patógenos Específicos , Baço/anormalidades , Baço/citologia , Esplenectomia , Células Th2/imunologiaRESUMO
Recently, considerable progress has been made in understanding the molecular basis by which cytokines released from CD4+ helper T cells contribute to allergic disease. A subset of CD4+ helper T cells, termed TH2 cells, produce interleukin 4 (IL-4) and IL-5, but not interferon gamma. IL-4 has a critical role in causing B-cell immunoglobulin-isotype switch leading to IgE synthesis, and IL-5 governs eosinophilic inflammation of airway tissue. Studies on the mechanisms whereby TH2 cells, and non-T cells such as metachromatic cells, produce a highly restricted panel of cytokines has revealed molecular mechanisms that may affect our views on the induction and treatment of asthma, and these are discussed in this review by Gary Anderson and Anthony Coyle. TH2 cytokine biology may enable pharmacologists to design better, and perhaps even preventative, therapies for the treatment of asthma and allergy. Surprisingly IL-4, rather than IL-5, is emerging as a critical drug target owing to its central role in the regulation of CD4+ helper T cell phenotype commitment.