RESUMO
BACKGROUND: Allergic asthma is a chronic airway inflammatory disease involving the complementary actions of innate and adaptive immune responses. Endogenously generated cannabinoids acting via CB2 receptors play important roles in both homeostatic and inflammatory processes. However, the contribution of CB2-acting eicosanoids to the innate events preceding sensitization to the common house dust mite (HDM) allergen remains to be elucidated. We investigated the role of CB2 activation during allergen-induced pulmonary inflammation and natural killer (NK) cell effector function. METHODS: Lung mucosal responses in CB2-deficient (CB2-/- ) mice were examined and compared with wild-type (WT) littermates following intranasal exposure to HDM allergen. RESULTS: Mice lacking CB2 receptors exhibited elevated numbers of pulmonary NK cells yet were resistant to the induction of allergic inflammation exemplified by diminished airway eosinophilia, type 2 cytokine production and mucus secretion after allergen inhalation. This phenomenon was corroborated when WT mice were treated with a CB2-specific antagonist that caused a pronounced inhibition of HDM-induced airway inflammation and goblet cell hyperplasia. Unexpectedly, the preponderance of NK cells in the lungs of CB2-/- mice correlated with reduced numbers of group 2 innate lymphoid cells (ILC2s). Depletion of NK cells restored the allergen responsiveness in the lungs and was associated with elevated ILC2 numbers. CONCLUSIONS: Collectively, these results reveal that CB2 activation is crucial in regulating pulmonary NK cell function, and suggest that NK cells serve to limit ILC2 activation and subsequent allergic airway inflammation. CB2 inhibition may present an important target to modulate NK cell response during pulmonary inflammation.
Assuntos
Asma/patologia , Inflamação/imunologia , Células Matadoras Naturais/fisiologia , Receptor CB2 de Canabinoide/fisiologia , Imunidade Adaptativa , Animais , Antígenos de Dermatophagoides/farmacologia , Asma/imunologia , Imunidade Inata , Células Matadoras Naturais/imunologia , Camundongos , Camundongos Knockout , Receptor CB2 de Canabinoide/deficiência , Receptor CB2 de Canabinoide/imunologiaRESUMO
There is a growing interest in the development and use of natural emulsifiers, which provide biodegradability as well as non-toxicity along with giving better performance compared to existing emulsifying agents used in the food industry. A large variety of sources of starting material, i.e., the microorganisms, are available to be used, hence giving a diverse range of applications. The focus of this review paper is on the production of bioemulsifiers, which are said to be "green surfactants", from fungi, bacteria and yeasts; furthermore, an overview pertaining to the knowledge gained over the years in terms of characterization techniques is reported. The methods used for the characterization and isolation such as TLC, GC-MS, HPLC, NMR have also been studied. The end-application products such as cookies, muffins, and doughs along with the methods used for the incorporation of bioemulsifiers, microorganisms from which they are derived, properties imparted to the product with the use of a particular bioemulsifier and comparison with the existing food grade emulsifiers has been discussed in detail. The future prospects indicate that newer bioemulsifiers with anti-microbial, anti-oxidant and stabilization properties will prove to have a larger impact, and emphasis will be on improving the performance at an economically viable methodology.
RESUMO
Nedocromil sodium and sodium cromoglycate inhibited histamine release from rat peritoneal mast cells. Tachyphylaxis was observed with both drugs. The 2 compounds were extremely selective in their action, being less active against peritoneal mast cells from the hamster and completely ineffective against mast cells from the mouse. Human basophil leucocytes, tissue mast cells of the guinea-pig and rat intestinal mast cells were also unresponsive. Both drugs inhibited immunological histamine release from human pulmonary mast cells obtained by bronchoalveolar lavage (BAL) and, less effectively, from lung parenchyma. Nedocromil sodium was about 1 order of magnitude more potent than sodium cromoglycate in each case. Tachyphylaxis was observed with the dispersed lung, but not with the cells obtained by BAL, and the degree of inhibition varied inversely with the magnitude of the secretory response. The possible clinical significance of these observations is discussed.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Cromolina Sódica/farmacologia , Liberação de Histamina/efeitos dos fármacos , Mastócitos/metabolismo , Quinolonas/farmacologia , Animais , Cricetinae , Humanos , Técnicas In Vitro , Pulmão/citologia , Masculino , Mastócitos/efeitos dos fármacos , Mesocricetus , Camundongos , Nedocromil , Nippostrongylus/imunologia , Peritônio/citologia , Ratos , Ratos EndogâmicosRESUMO
Extracellular adenosine 5'-triphosphate (ATP) induced a characteristic, dose-dependent release of histamine and prostaglandin D2 (PGD2) from rat peritoneal mast cells. The process was relatively slow, non-cytotoxic, maximal at physiological pH and dependent on external calcium. Strontium and barium ions were able to substitute for calcium, although higher concentrations were required for maximal release. Cells stimulated in the absence of calcium progressively lost the ability to respond to subsequent reintroduction of the cation. The secretion of histamine induced by ATP was largely unaffected by the anti-asthmatic drugs disodium cromoglycate and nedocromil sodium but was inhibited by structurally related flavonoids and by cAMP-active drugs. Importantly, the non-hydrolysable guanosine 5'-triphosphate (GTP) analogue, GTP-gamma-S, elicited a dose-dependent release of histamine when introduced into mast cells permeabilized with ATP in the absence of external calcium. ATP thus appears to be a useful cell permeabilizing tool with which to study the biochemical processes involved in mast cell activation.
Assuntos
Trifosfato de Adenosina/farmacologia , Permeabilidade da Membrana Celular/efeitos dos fármacos , Liberação de Histamina/efeitos dos fármacos , Mastócitos/metabolismo , Animais , Cromonas/farmacologia , AMP Cíclico/metabolismo , Guanosina 5'-O-(3-Tiotrifosfato)/farmacologia , Técnicas In Vitro , Inositol 1,4,5-Trifosfato/farmacologia , Mastócitos/efeitos dos fármacos , Metais/farmacologia , Peso Molecular , Cavidade Peritoneal/citologia , Prostaglandina D2/metabolismo , Ratos , Ratos Sprague-DawleyRESUMO
Adenosine 5'-triphosphate (ATP) induced a non-cytotoxic, calcium-dependent release of histamine and prostaglandin D2 from rat serosal mast cells. The effect was tissue and species specific. In particular, tissue mast cells from the guinea pig and man were totally unresponsive. The release evoked by the nucleotide was unaffected by antiasthmatic chromones but was inhibited by structurally related flavonoids and by cAMP-active drugs. Secretion was blocked both by the P2Y-purinoceptor antagonist reactive blue 2 and the P2X-antagonist suramin, whereas the P2-agonists 2-methylthio ATP, alpha, beta- and beta, gamma-methylene ATP were inactive. These findings indicate that ATP acts through a novel purinoceptor or binding site which is present on certain types of mast cells.
Assuntos
Trifosfato de Adenosina/farmacologia , Liberação de Histamina/efeitos dos fármacos , Mastócitos/metabolismo , Animais , Cricetinae , Cobaias , Humanos , Técnicas In Vitro , Mastócitos/efeitos dos fármacos , Mesocricetus , Camundongos , Infecções por Nematoides/imunologia , Infecções por Nematoides/metabolismo , Ratos , Ratos Endogâmicos , Estimulação QuímicaRESUMO
Theophylline has been shown by several investigators to attenuate the late asthmatic response (LAR) to inhaled allergen, suggesting that it has anti-inflammatory or immunomodulatory properties. We have, therefore, undertaken a double-blind, placebo-controlled study to examine the effects of low-dose theophylline on bronchoalveolar lavage (BAL) and blood T-lymphocyte profile and activation in asthmatics following antigen challenge and the development of a LAR. Peripheral blood and BAL samples were obtained from 17 subjects with mild atopic asthma before and after 6 weeks of treatment with either oral theophylline or placebo. The mean serum theophylline concentration achieved was 6.6 micrograms.mL-1, which is below the currently accepted therapeutic range. Following theophylline therapy, there was a significant decrease in the number of BAL lymphocytes compared to placebo. On flow cytometric analysis of BAL cells, a significant loss of CD3+ T-lymphocytes, comprising both CD4+ and CD8+ subsets, was demonstrated. Moreover, there was a decrease in the number of BAL CD4+ T-cells expressing the activation marker very late activation antigen-1 (VLA-1), and an apparent reduction in human leucocyte antigen-DR (HLA-DR). Correspondingly, this was accompanied in the blood by an elevation in the proportion of activated CD4+ T-lymphocytes, in particular those expressing HLA-DR. These findings provide further evidence that theophylline has an anti-inflammatory action in asthma.
Assuntos
Asma/tratamento farmacológico , Testes de Provocação Brônquica , Líquido da Lavagem Broncoalveolar/citologia , Broncodilatadores/uso terapêutico , Linfócitos T/efeitos dos fármacos , Teofilina/uso terapêutico , Adulto , Asma/imunologia , Broncodilatadores/administração & dosagem , Broncoscopia , Relação CD4-CD8 , Relação Dose-Resposta a Droga , Método Duplo-Cego , Feminino , Citometria de Fluxo , Humanos , Masculino , Pico do Fluxo Expiratório/efeitos dos fármacos , Subpopulações de Linfócitos T/efeitos dos fármacos , Subpopulações de Linfócitos T/imunologia , Linfócitos T/imunologia , Teofilina/administração & dosagem , Resultado do TratamentoRESUMO
Theophylline, in addition to its bronchodilator effect, may attenuate inflammation in asthma. We did a double-blind placebo-controlled study of the effect of oral theophylline on the inflammatory response of the bronchial mucosa to inhalation of allergen in 19 atopic asthmatic subjects. Bronchoscopy and bronchial biopsy were done 24 hours after allergen inhalation before and after six weeks of treatment with oral slow-release theophylline, 200 mg 12 hourly. The mean serum concentration was 36.6 mumol/L, which is below the currently-accepted therapeutic range. After treatment with theophylline there was a significant reduction in the number of EG2-positive activated eosinophils (5.9 before and 2.1 after treatment, Wilcoxon signed rank p < 0.05) and total eosinophils (16.7 before and 7.6 after treatment, p < 0.05) beneath the epithelial basement membrane. We conclude that low-dose oral theophylline attenuates airway inflammatory response to allergen inhalation in atopic asthma.
Assuntos
Asma/prevenção & controle , Teofilina/uso terapêutico , Administração Oral , Adulto , Alérgenos/efeitos adversos , Brônquios/efeitos dos fármacos , Brônquios/patologia , Broncoscopia , Relação Dose-Resposta a Droga , Método Duplo-Cego , Eosinófilos/efeitos dos fármacos , Feminino , Humanos , Fluxo Expiratório Máximo , Teofilina/administração & dosagem , Teofilina/sangueRESUMO
DO11.10 transgenic mice, expressing an OVA-specific TCR, were used to study pulmonary T cell responses to inhaled Ags. Before OVA inhalation, the activation of lung parenchymal T cells elicited both strong proliferative responses and IL-2 production. However, following Ag inhalation the proliferative responses of the lung T cells, when restimulated in vitro with OVA323-339 peptide or immobilized anti-CD3, were severely attenuated and associated with a decrease in the level of production of IL-2 but not IFN-gamma. Such immune regulation was tissue-specific, because T cell responses in the lymph nodes and spleens were normal. This dramatic aerosol-induced attenuation of parenchymal T cell proliferation was also observed in BALB/c mice immunized with OVA and in BALB/c mice following adoptive transfer of DO11.10 T cells bearing either a Th1 or Th2 phenotype. In mice that had received Th2 cells, the reduced proliferative responses were associated with a decrease in IL-2 expression but augmented IL-4 and IL-5 production. Invariably, the inhibition of proliferation was a consequence of the action of F4/80+ interstitial macrophages and did not involve alveolar macrophages or their products. These observations demonstrate that clonal expansion of T cells in the lung compartment is prevented following the onset of either Th1- or Th2-mediated inflammation. This form of immune regulation, which appears as a selective defect in IL-2-driven proliferation, may serve to prevent the development of chronic pulmonary lymphoproliferative responses.
Assuntos
Antígenos/imunologia , Pulmão/imunologia , Pulmão/patologia , Células Th1/imunologia , Células Th2/imunologia , Administração por Inalação , Administração Intranasal , Aerossóis , Animais , Antígenos/administração & dosagem , Antígenos de Diferenciação/análise , Adesão Celular/imunologia , Comunicação Celular/imunologia , Contagem de Células , Citocinas/biossíntese , Citocinas/genética , Inflamação/imunologia , Inflamação/metabolismo , Interleucina-2/antagonistas & inibidores , Interleucina-2/biossíntese , Interleucina-4/biossíntese , Interleucina-5/biossíntese , Pulmão/metabolismo , Ativação Linfocitária/imunologia , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Ovalbumina/administração & dosagem , Ovalbumina/imunologia , RNA Mensageiro/biossíntese , Células Th1/metabolismo , Células Th2/metabolismoRESUMO
CD80 and CD86 interact with CD28 and deliver costimulatory signals required for T cell activation. We demonstrate that ex vivo allergen stimulation of bronchial biopsy tissue from mild atopic asthmatic, but not atopic nonasthmatic, subjects induced production of IL-5, IL-4, and IL-13. Explants from both study groups did not produce IFN-gamma, but secreted the chemokine RANTES without any overt stimulation. In addition to allergen, stimulation of asthmatic explants with mAbs to CD3 and TCR-alphabeta but not TCR-gammadelta induced IL-5 secretion. Allergen-induced IL-5 and IL-13 production by the asthmatic tissue was inhibited by anti-CD80 and, to a lesser extent, by anti-CD86 mAbs. In contrast, the production of these cytokines by PBMCs was not affected by mAbs to CD80, was inhibited by anti-CD86, and was strongly attenuated in the presence of both Abs. FACS analysis revealed that stimulated asthmatic bronchial tissue was comprised of CD4+ T cells that expressed surface CD28 (75. 3%) but little CTLA-4 (4.0%). Neutralizing mAbs to CD40 ligand had no effect on the cytokine levels produced by asthmatic tissue or PBMCs. Collectively, these findings suggest that allergen-specific alphabeta T cells are resident in asthmatic bronchial tissue and demonstrate that costimulation by both CD80 and CD86 is essential for allergen-induced cytokine production. In contrast, CD86 appears to be the principal costimulatory molecule required in PBMC responses. Attenuation of type 2 alphabeta T cell responses in the bronchial mucosa by blocking these costimulatory molecules may be of therapeutic potential in asthma.
Assuntos
Alérgenos/imunologia , Antígenos CD , Asma/imunologia , Brônquios/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta , Subpopulações de Linfócitos T/imunologia , Adulto , Antígeno B7-1 , Antígeno B7-2 , Antígenos CD40 , Citocinas , Feminino , Humanos , Ativação Linfocitária , Masculino , Glicoproteínas de Membrana , Receptores de Antígenos de Linfócitos T gama-delta , Transdução de Sinais , Células Th2RESUMO
Asthma is a complex disorder characterized by airway hyperreactivity and inflammation. To analyze cellular interactions required for the secretion of cytokines by the bronchial mucosa, we have evaluated the ex vivo response of tissue explants to allergen. Endobronchial mucosal biopsy tissue from mild atopic asthmatic subjects and normal control subjects were maintained in culture for 24 h. To detect reactivity to allergen, the explants were stimulated with dust mite extract Dermatophagoides pteronyssinus (Der p). Our analysis revealed that without any overt stimulation, mRNA transcripts for interleukin (IL)-5 and IL-13 were expressed by asthmatic but not normal bronchial tissue. In contrast, the expression of interferon-gamma was observed in a higher proportion of cultured bronchial biopsies from the normal control subjects than in those from asthmatic subjects. Addition of Der p allergen did not change the cytokine profile of the explants from control volunteers but augmented the expression of IL-5 mRNA and induced secretion of the protein by the asthmatic bronchial tissue. In most cases, allergen also increased the production of IL-13 by bronchial tissue from asthmatic subjects. The allergen-induced secretion of IL-5 and IL-13 was inhibited by the fusion protein CTLA-4Ig, reflecting a requirement for CD80 (B7-1) and/or CD86 (B7-2) costimulation for the expression of the Th2 cytokines. This requirement for B7/CD28 costimulation is consistent with the hypothesis that IL-5 and IL-13 are produced by allergen-specific T cells resident in the asthmatic bronchial mucosa.
Assuntos
Alérgenos/imunologia , Asma/imunologia , Antígeno B7-1/imunologia , Brônquios/metabolismo , Interleucina-13/metabolismo , Interleucina-5/metabolismo , Adulto , Antígenos CD/imunologia , Antígeno B7-2 , Biópsia , Antígenos CD28/imunologia , Técnicas de Cultura , Citocinas/genética , Citocinas/metabolismo , Feminino , Expressão Gênica , Humanos , Interleucina-13/genética , Interleucina-5/genética , Masculino , Glicoproteínas de Membrana/imunologia , Pessoa de Meia-Idade , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T/imunologiaRESUMO
BACKGROUND: T cells play an important role in airway inflammation in asthma through the release of T(H)2 cytokines. Optimal T-cell activation by antigen-presenting cells requires co-stimulatory signaling, such as the interaction of CD80, CD86, or both with CD28. In patients with mild allergic asthma, the fusion protein cytotoxic T-lymphocyte antigen 4Ig (CTLA-4Ig), which inhibits CD28-mediated signaling, blocks the release of IL-5 and IL-13 from bronchial explant cultures exposed to the allergen Dermatophagoides pteronyssinus. OBJECTIVES: To assess costimulation in more severe forms of atopic asthma, we have compared the ability of CTLA-4Ig to block allergen-induced cytokine responses of bronchial explants and PBMCs from patients with moderately severe asthma. METHODS: Bronchial explants and PBMCs were cultured in vitro, and cytokine expression was measured by means of quantitative RT-PCR and ELISA. RESULTS: Constitutive mRNA transcripts for IL-5, IL-13, and GM-CSF were detected in the tissue explants, but only IL-5 mRNA increased significantly with allergen stimulation. Consistent with increased transcription, allergen-stimulated IL-5 protein release into explant supernatants, but this was not blocked by CTLA-4Ig. Allergen did not induce GM-CSF release, and IL-13 protein could not be detected in the explant supernatants under any condition. In contrast, allergen enhanced production of IL-5 and IL-13 by PBMC cultures from the same subjects, and this was inhibited effectively by CTLA-4Ig. CONCLUSIONS: These data suggest that IL-5 production in the airways of subjects with moderately severe asthma is largely independent of CD28-mediated costimulation. The different requirements for CD28-mediated costimulation in PBMC cultures and bronchial tissue cultures emphasizes the importance of the tissue microenvironment in pulmonary inflammatory responses in severe asthma.
Assuntos
Alérgenos/imunologia , Asma/imunologia , Antígeno B7-1/fisiologia , Brônquios/imunologia , Antígenos CD28/fisiologia , Citocinas/biossíntese , Imunoconjugados , Abatacepte , Adulto , Antígenos CD/fisiologia , Antígenos de Diferenciação/uso terapêutico , Asma/terapia , Antígeno B7-2 , Antígeno CTLA-4 , Células Cultivadas , Feminino , Humanos , Masculino , Glicoproteínas de Membrana/fisiologia , Mucosa/imunologiaRESUMO
Interleukin (IL)-5 is a pleiotropic cytokine that exhibits biologic activity on cells of diverse hemopoieitic lineages. IL-5 enhances mediator release from human basophils and plays a pivotal role in the chemoattraction, proliferation, differentiation, survival, and activation of eosinophils. Th2- and Tc2-like T cells, mast cells, basophils, and eosinophils are the known cellular sources of this cytokine. Using a sensitive and novel reverse transcription-polymerase chain reaction enzyme-linked immunosorbent assay system, we found that IL-5 messenger RNA (mRNA) was constitutively expressed in bronchial biopsies obtained from healthy individuals, and that the levels of IL-5 mRNA expression decreased 1. 5 h after exposure to 0.12 ppm ozone for 2 h. Because the oxidative effects of ozone are confined to the epithelial cell surface and it is known that ozone induces epithelial damage and shedding, we hypothesized that epithelial cells might be a source of IL-5 mRNA. We demonstrate here that both transformed human bronchial epithelial cell lines (A549 and 16HBE14o-) and primary human bronchial and nasal epithelial cells grown in culture constitutively express IL-5 mRNA, which is upregulated on stimulation with tumor necrosis factor (TNF)-alpha. Culture supernatants derived from A549 cells exposed to TNF-alpha and interferon-gamma demonstrated detectable levels of IL-5 protein, and immunostaining of bronchial biopsies obtained from healthy human airways revealed the presence of IL-5 protein localized to the bronchial epithelium. To our knowledge, this is the first report demonstrating IL-5 production by human airway epithelial cells. This observation provides further evidence for the role of airway epithelium in regulating airway immune responses, in particular enhancing chemotaxis, activation, and survival of eosinophils, which could play an important role in the pathogenesis of bronchial asthma.