RESUMO
BACKGROUND: There is a growing body of evidence to suggest that CD8+ T lymphocytes contribute to local allergen-induced eosinophilic inflammation. Since bone marrow (BM) responses are intricately involved in the induction of airway eosinophilia, we hypothesized that CD8+ T lymphocytes, as well as CD4+ T lymphocytes, may be involved in this process. METHODS: Several approaches were utilized. Firstly, mice overexpressing interleukin-5 (IL-5) in CD3+ T lymphocytes (NJ.1638; CD3IL-5+ mice) were bred with gene knockout mice lacking either CD4+ T lymphocytes (CD4-/-) or CD8+ T lymphocytes (CD8-/-) to produce CD3IL-5+ knockout mice deficient in CD4+ T lymphocytes (CD3IL-5+/CD4-/-) and CD8+ T lymphocytes (CD3IL-5+/CD8-/-), respectively. Secondly, CD3+, CD4+ and CD8+ T lymphocytes from naïve CD3IL-5+ and C57BL/6 mice were adoptively transferred to immunodeficient SCID-bg mice to determine their effect on BM eosinophilia. Thirdly, CD3IL-5+, CD3IL-5+/CD8-/- and CD3IL-5+/CD4-/- mice were sensitized and allergen challenged. Bone marrow and blood samples were collected in all experiments. RESULTS: The number of BM eosinophils was significantly reduced in CD3IL-5+/CD8-/- mice compared to CD3IL-5+ mice and CD3IL-5+/CD4-/- mice. Serum IL-5 was significantly higher in CD3IL-5+/CD4-/- mice compared to CD3IL-5+ mice but there was no difference in serum IL-5 between CD3IL-5+/CD4-/- and CD3IL-5+/CD8-/- mice. Adoptive transfer of CD8+, but not CD4+ T lymphocytes from naïve CD3IL-5+ and C57BL/6 mice restored BM eosinophilia in immunodeficient SCID-bg mice. Additionally, allergen challenged CD3IL-5+/CD8-/- mice developed lower numbers of BM eosinophils compared to CD3IL-5+ mice and CD3IL-5+/CD4-/- mice. CONCLUSION: This study shows that CD8+ T lymphocytes are intricately involved in the regulation of BM eosinophilopoiesis, both in non-sensitized as well as sensitized and allergen challenged mice.
Assuntos
Células da Medula Óssea/imunologia , Linfócitos T CD8-Positivos/imunologia , Eosinófilos/imunologia , Leucopoese , Eosinofilia Pulmonar/imunologia , Hipersensibilidade Respiratória/imunologia , Subpopulações de Linfócitos T/imunologia , Transferência Adotiva , Animais , Líquido da Lavagem Broncoalveolar/citologia , Complexo CD3/análise , Complexo CD3/genética , Antígenos CD4/análise , Antígenos CD4/genética , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/transplante , Antígenos CD8/análise , Antígenos CD8/genética , Linfócitos T CD8-Positivos/transplante , Proteína Básica Maior de Eosinófilos/biossíntese , Interleucina-5/biossíntese , Interleucina-5/sangue , Interleucina-5/genética , Contagem de Leucócitos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos SCID , Ovalbumina , Eosinofilia Pulmonar/sangue , Hipersensibilidade Respiratória/sangue , Subpopulações de Linfócitos T/transplanteRESUMO
BACKGROUND: IL-17E is a new TH2 cytokine that promotes airway eosinophilia in mice. IL-17E proinflammatory activity has been proposed to involve induction of cytokine and chemokine production. Recruitment of inflammatory cells may be mediated by tissue-resident cells. OBJECTIVE: This study aimed to evaluate whether fibroblasts represent a target of IL-17E for the production of eosinophil active mediators in the lung. METHODS: Expression of IL-17B receptor (IL-17BR), a receptor for IL-17E, was evaluated by immunofluorescent staining, Western blot, and real-time PCR in human primary lung fibroblasts. Mediator production was analyzed by using real-time PCR and ELISA after stimulation of fibroblasts with IL-17E alone or in combination with TNF-alpha and TGF-beta1. Expression of IL-17E and of eosinophil major basic protein was evaluated by immunohistochemistry in bronchial biopsies from subjects with asthma. RESULTS: Human primary lung fibroblasts constitutively expressed IL-17BR. IL-17BR mRNA levels were increased in cells stimulated with TNF-alpha and decreased with TGF-beta1. IL-17E slightly upregulated CC chemokine ligand (CCL)-5, CCL-11, GM-CSF, and CXC chemokine ligand (CXCL)-8 mRNA in fibroblasts. Moreover, IL-17E and TNF-alpha synergistically induced GM-CSF and CXCL-8 mRNA. IL-17E also potentiated the upregulation of CXCL-8 transcripts observed with TGF-beta1. In contrast, TGF-beta1 decreased IL-17E-induced CCL-11 mRNA. The capacity of IL-17E to enhance GM-CSF and CXCL-8 responses to TNF-alpha was accompanied by production and secretion of both proteins by lung fibroblasts. Finally, IL-17E was detected in asthma in eosinophil-infiltrated bronchial submucosa. CONCLUSION: IL-17E may contribute to the induction and maintenance of eosinophilic inflammation in the airways by acting on lung fibroblasts. This study supports a role for IL-17E in asthma pathophysiology.
Assuntos
Asma/imunologia , Citocinas/imunologia , Eosinófilos/imunologia , Fibroblastos/imunologia , Interleucina-17/fisiologia , Asma/fisiopatologia , Biópsia , Brônquios/imunologia , Brônquios/patologia , Células Cultivadas , Quimiocina CCL11 , Quimiocina CCL5 , Quimiocinas CC/biossíntese , Quimiocinas CC/imunologia , Quimiocinas CXC/biossíntese , Quimiocinas CXC/imunologia , Fatores Quimiotáticos de Eosinófilos/biossíntese , Fatores Quimiotáticos de Eosinófilos/imunologia , Quimiotaxia de Leucócito , Citocinas/biossíntese , Citocinas/fisiologia , Proteína Básica Maior de Eosinófilos/biossíntese , Proteína Básica Maior de Eosinófilos/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/biossíntese , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Humanos , Inflamação , Interleucina-17/biossíntese , Pulmão/imunologia , Receptores de Interleucina/biossíntese , Receptores de Interleucina/imunologia , Receptores de Interleucina-17 , Fator de Crescimento Transformador beta/fisiologia , Fator de Crescimento Transformador beta1 , Fator de Necrose Tumoral alfa/fisiologia , Regulação para CimaRESUMO
BACKGROUND: Basophils possess characteristics of both mast cells and eosinophils, and all 3 cell types often are found together, particularly during allergic reactions. A mAb (J175-7D4) generated against the recombinant pro-form of human eosinophil granule major basic protein 1 (rproMBP1) appeared to stain only basophils in tissue specimens. OBJECTIVE: We investigated J175-7D4 to characterize its specificity for basophils. METHODS: Fluid-phase immunoprecipitation, Western blotting, and immunocytochemistry and immunohistochemistry were used to establish the specificity of J175-7D4. RESULTS: First, J175-7D4 binds to various glycosylated and proteolytically processed forms of rproMBP1, but not to major basic protein. Second, cells transfected with the rproMBP1 gene and human placental tissue (known to express the pro-form of major basic protein 1 [proMBP1]) stain specifically with J175-7D4. In contrast, although mature eosinophils contain substantial major basic protein, they lack proMBP1 and do not stain. Neutrophils, lymphocytes, monocytes, and skin mast cells also are not stained. However, blood basophils are stained by J175-7D4, anti-IgE, Wright-Giemsa (metachromatically), and a previously characterized basophil-specific mAb, 2D7. Finally, formaldehyde-fixed, paraffin-embedded basophils are identically detected by J175-7D4 and 2D7, and J175-7D4 also recognizes putative basophils in formaldehyde-fixed, paraffin-embedded tissue specimens from inflammatory dermatoses, such as atopic dermatitis and delayed pressure urticaria. CONCLUSION: The J175-7D4 mAb recognizes proMBP1 as a novel marker for human basophils. J175-7D4 should prove useful for characterizing basophil involvement in human health and disease.
Assuntos
Anticorpos Monoclonais/imunologia , Basófilos/imunologia , Proteína Básica Maior de Eosinófilos/biossíntese , Imunoproteínas/biossíntese , Precursores de Proteínas/biossíntese , Basófilos/metabolismo , Biomarcadores , Proteína Básica Maior de Eosinófilos/imunologia , HumanosRESUMO
To determine whether immunostimulatory sequences of DNA (ISS) can reverse established airway remodeling, mice that had developed airway remodeling following 3 mo of repetitive OVA challenges, were treated with ISS for 1-3 mo. Systemic administration of ISS to mice that had already developed established airway remodeling significantly reduced the degree of airway collagen deposition (assessed by lung collagen content, peribronchial trichrome staining, and immunostaining with anticollagen type III and type V Abs). ISS reduced bronchoalveolar lavage and lung levels of TGF-beta1 and reduced the number of TGF-beta1-positive eosinophils and TGF-beta1-positive mononuclear cells recruited to the airway. In vitro studies demonstrated that ISS inhibited TGF-beta1 expression by macrophages (RAW 264.7 cell line and bone marrow-derived macrophages). In addition, ISS significantly reduces lung levels of expression of the chemokine thymus- and activation-regulated chemokine, as well as the number of peribronchial CD4(+) lymphocytes that express Th2 cytokines that promote peribronchial fibrosis. Overall, these studies demonstrate that ISS can reverse features of airway collagen deposition by reducing levels of lung TGF-beta1, as well as by reducing levels of the chemokine thymus- and activation-regulated chemokine and the numbers of peribronchial CD4(+) lymphocytes that drive the ongoing Th2 immune response.