RESUMO
BACKGROUND: Seasonal allergic rhinitis is a chronic inflammation in the nasal mucosa triggered by inhaled aeroallergens. The inflammatory reaction is controlled by allergen-specific T cells, but where and how these T cells become activated is not fully understood. OBJECTIVES: We wanted to determine whether allergen-specific T-helper (Th) 2 cells are residing in the nasal mucosa under steady-state conditions outside of the pollen season and, if so, whether these cells are activated locally in response to allergen challenge. METHODS: Mucosal biopsies from the lower turbinate were obtained out of season from patients with either birch- or grass-pollen-allergic rhinitis and from healthy controls. Cultured explant samples were challenged with relevant pollen extract or with a mix of overlapping 20-mer peptides derived from the sequence of the major birch allergen, Betula verrucosa (Bet v) 1. After 24 h, culture medium was harvested for multiplex cytokine and tryptase analysis. RESULTS: Significant amounts of interleukin (IL)-5 were secreted from resident cells in response to ex vivo allergen challenge in the allergic group only. No increase was observed for the other cytokines measured. Production of IL-5 in response to both extract and the Bet v1-derived peptide mix strongly suggested that T cells were a major source of IL-5. CONCLUSION: Our explant model indicated that local presentation of antigen to resident allergen-specific Th2 cells is the early event in the pathogenesis of allergic rhinitis. These findings identify possible cellular targets for anti-inflammatory treatment.
Assuntos
Antígenos de Plantas/imunologia , Interleucina-5/imunologia , Modelos Imunológicos , Mucosa Nasal/imunologia , Rinite Alérgica/imunologia , Células Th2/imunologia , Adolescente , Adulto , Criança , Feminino , Humanos , Masculino , Mucosa Nasal/patologia , Rinite Alérgica/patologia , Células Th2/patologia , Técnicas de Cultura de TecidosRESUMO
Lymphocyte recruitment to peripheral tissues is fundamental for immune surveillance and homeostasis, but the chemokines and chemokine receptors responsible for tissue-specific homing of T cells to the upper airway mucosa have not been determined. To address this, we analyzed the chemokines expressed in the normal human nasal mucosa and found that CCL28 is preferentially expressed at a high level on the lumenal face of vascular endothelial cells in the mucosa. Analysis of the cognate chemokine receptors revealed that close to 50% of the CD4(+) T cells in the human nasal mucosa expressed the CCL28 receptor CCR3, whereas CCR3 was hardly detectable on T cells in the small intestine and skin. In the circulation, CCR3(+) T cells comprised a small subset that did not express homing receptors to the intestine or skin. Moreover, depletion of CCR3(+)CD4(+) T cells abrogated the proliferative response of human blood CD4(+) T cells against the opportunistic nasopharyngeal pathogen Haemophilus influenzae, indicating that the CCR3(+)CD4(+) T-cell subset in the circulation contains antigen specificities relevant for the upper airways. Together, these findings indicate that CCL28-CCR3 interactions are involved in the homeostatic trafficking of CD4(+) T cells to the upper airways.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Quimiocinas CC/metabolismo , Endotélio Vascular/imunologia , Haemophilus influenzae/imunologia , Mucosa Nasal/imunologia , Receptores CCR3/metabolismo , Receptores de Retorno de Linfócitos/metabolismo , Adulto , Idoso , Antígenos de Bactérias/imunologia , Movimento Celular , Proliferação de Células , Células Cultivadas , Feminino , Humanos , Ativação Linfocitária , Depleção Linfocítica , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
Secretory IgA (SIgA) antibodies represent the first line of antigen-specific immune defense protecting the mucosal surfaces against environmental pathogens and antigens, and maintaining homeostasis with the commensal microbiota. The polymeric immunoglobulin receptor (pIgR) has the dual role of transporting locally produced dimeric IgA across mucosal epithelia, and serving as the precursor of secretory component, a glycoprotein that enhances the immune functions of SIgA. The complex regulation of pIgR expression and transcytosis by host and microbial factors is finely tuned to optimize the role of SIgA in mucosal immunity. Disruption of this regulatory network in disease states similar to inflammatory bowel disease can result in profound consequences for mucosal homeostasis and systemic sequelae. Future research into the function and regulation of pIgR and SIgA may offer new insights into the prevention and treatment of infectious and inflammatory diseases that originate at mucosal surfaces.
Assuntos
Imunidade nas Mucosas , Imunoglobulina A/imunologia , Doenças Inflamatórias Intestinais/imunologia , Mucosa Intestinal/imunologia , Receptores Fc/imunologia , Animais , Exposição Ambiental/efeitos adversos , Regulação da Expressão Gênica/imunologia , Humanos , Depuração Mucociliar/imunologia , Transcitose/imunologiaRESUMO
The polymeric immunoglobulin receptor (pIgR) transports IgA antibodies across intestinal epithelial cells (IECs). Expression of pIgR is upregulated by proinflammatory signaling pathways via activation of nuclear factor-κB (NF-κB). Here, we examined the contributions of the RelA-dependent classical and RelB-dependent alternative pathways of NF-κB to pIgR regulation in the HT-29 human IEC line following stimulation with tumor necrosis factor (TNF), lipopolysaccharide (LPS; Toll-like receptor 4 (TLR4) ligand), and polyinosinic: polycytidylic acid (pIC; TLR3 ligand). Whereas induction of proinflammatory genes such as interleukin-8 (IL-8) required only RelA, pIgR expression was regulated by complex mechanisms that involved both RelA and RelB. Upregulation of pIgR expression by ligation of the lymphotoxin-ß receptor suggested a direct role for the alternative NF-κB pathway. Inhibition of mitogen-activated protein kinases reduced the induction of IL-8, but enhanced the induction of pIgR by TNF and TLR signaling. Regulation of pIgR through unique signaling pathways could allow IECs to sustain high levels of IgA transport while limiting the proinflammatory responses.
Assuntos
Células Epiteliais , Mucosa Intestinal , NF-kappa B/metabolismo , Receptores de Imunoglobulina Polimérica/imunologia , Transdução de Sinais/imunologia , Células Epiteliais/imunologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/imunologia , Técnicas de Silenciamento de Genes , Células HT29 , Humanos , Fatores Imunológicos/metabolismo , Fatores Imunológicos/farmacologia , Interleucina-8/genética , Interleucina-8/imunologia , Mucosa Intestinal/imunologia , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , RNA Mensageiro/genética , Receptores de Imunoglobulina Polimérica/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Receptores Toll-Like/metabolismo , Fator de Transcrição RelA/genética , Fatores de Necrose Tumoral/metabolismo , Fatores de Necrose Tumoral/farmacologiaRESUMO
BACKGROUND: It has been suggested that Foxp3(+) regulatory T (Treg) cells inhibit allergic inflammation in humans by suppressing the activation of allergen-specific effector T cells. Whether this occurs at the site of allergen exposure has not been determined. OBJECTIVE: To determine the occurrence of Foxp3(+) Treg cells in the nasal mucosa of allergic rhinitis (AR) patients and non-allergic controls after a nasal allergen challenge. METHODS: Pollen-allergic patients (n=18) and non-allergic volunteers (n=7) were challenged locally with pollen extract or placebo for 7 days outside the pollen season. Mucosal biopsies were obtained from the inferior turbinate on days 0, 1 and 7 and subjected to multi-colour immunofluorescence and blood was drawn for eosinophil counts on days 0, 2, 5 and 7. RESULTS: Only AR patients receiving pollen extract experienced typical allergic symptoms and demonstrated increased levels of eosinophils in peripheral blood and nasal mucosa. In allergic patients, a transient early increase (day 1) in CD3(+) T cells was observed in the nasal mucosa, followed by a significant increase of Foxp3(high) T cells at day 7. No changes were found in the control group. The majority of Foxp3(high) cells co-expressed CTLA-4, CD25 and CD4, and a substantial fraction expressed the proliferation marker Ki67. CONCLUSION AND CLINICAL RELEVANCE: Experimentally induced inflammation in AR patients leads to an early inflammatory response followed by accumulation of Foxp3(high) T cells in the nasal mucosa. Our findings are similar to that observed in allergic airways of experimental mice, which suggest that Treg cells are operative in allergic upper airway inflammation. It should be explored whether Treg cells accumulating in the nasal mucosa could be targets for therapeutic intervention.
Assuntos
Fatores de Transcrição Forkhead/metabolismo , Rinite Alérgica Sazonal/imunologia , Linfócitos T Reguladores/imunologia , Adulto , Alérgenos/imunologia , Animais , Gatos , Cães , Eosinófilos/imunologia , Humanos , Masculino , Mucosa Nasal/citologia , Mucosa Nasal/imunologia , Testes de Provocação Nasal , Pólen/imunologia , Rinite Alérgica Sazonal/metabolismo , Rinite Alérgica Sazonal/fisiopatologia , Testes Cutâneos , Linfócitos T Reguladores/metabolismo , Adulto JovemRESUMO
BACKGROUND: Ultraviolet (UV) radiation has immunosuppressive effects and heliotherapy is a well-described treatment modality for psoriasis. OBJECTIVES: To characterize early sun-induced immunological changes both local and systemic in patients with psoriasis. METHODS: Twenty patients with moderate to severe psoriasis were subjected to controlled sun exposure on Gran Canaria, Canary Islands, Spain. Psoriasis Area and Severity Index (PASI) scores were evaluated. Skin biopsies were obtained from lesional and nonlesional skin in 10 patients at baseline and on day 16 and from five additional patients on day 2. Specimens were examined with immunohistochemistry and polymerase chain reaction. Blood samples were obtained from all patients at the same time points and were examined for T-cell subsets and cytokine production. RESULTS: Significant clinical improvement was achieved during the study period. CD4+ and CD8+ T cells in lesional skin were significantly reduced in both the epidermis and dermis. In contrast, dermal FOXP3+ T cells were relatively increased. In the peripheral blood skin homing cutaneous lymphocyte-associated antigen (CLA)+ T cells were significantly decreased after only 1 day in the sun and in vitro stimulated peripheral blood mononuclear cells demonstrated reduced capacity to secrete cytokines after 16 days. CONCLUSIONS: Our data show that clinical improvement of psoriasis following sun exposure is preceded by a rapid reduction in local and systemic inflammatory markers, strongly suggesting that immune modulation mediated the observed clinical effect. We cannot completely rule out that other mechanisms, such as stress reduction, may contribute, but it is extensively documented that UV irradiation is a potent inducer of immunosuppression and we therefore conclude that the observed effect was primarily due to sun exposure.
Assuntos
Citocinas/análise , Helioterapia , Psoríase/imunologia , Psoríase/radioterapia , Pele/imunologia , Pele/efeitos da radiação , Adulto , Idoso , Biópsia , Feminino , Humanos , Imuno-Histoquímica , Células de Langerhans/patologia , Leucócitos Mononucleares/imunologia , Masculino , Pessoa de Meia-Idade , Psoríase/patologia , Índice de Gravidade de Doença , Subpopulações de Linfócitos T/imunologia , Adulto JovemRESUMO
BACKGROUND: Mucosal immunity protects the epithelial barrier by immune exclusion of foreign antigens and by anti-inflammatory tolerance mechanisms, but there is a continuing debate about the role of secretory immunoglobulins (SIgs), particularly SIgA, in the protection against allergy and other inflammatory diseases. Lack of secretory antibodies may cause immune dysfunction and affect mucosally induced (oral) tolerance against food antigens. METHODS: We used polymeric Ig receptor (pIgR) knockout (KO) mice, which cannot export SIgA or SIgM, to study oral tolerance induction by ovalbumin (OVA) feeding and for parenteral antigen sensitization in the same animal. RESULTS: Remarkable systemic hyperreactivity was observed in pIgR KO mice, as 50% died after intradermal OVA challenge, which was not seen in similarly sensitized and challenged wild-type (WT) mice. Oral tolerance induced by OVA completely protected the sensitized pIgR KO mice against anaphylaxis and suppressed antibody levels (particularly IgG1) as well as delayed-type hypersensitivity (DTH) to OVA. Delayed-type hypersensitivity to a bystander antigen, human serum albumin, was also suppressed and T-cell proliferation against OVA in vitro was reduced in tolerized compared with non-tolerized pIgR KO mice. This effect was largely mediated by CD25+ T cells. Adoptive transfer of splenic putative regulatory T cells (CD4+ CD25+) obtained from OVA-fed pIgR KO mice to naïve WT mice mediated suppression of DTH against OVA after sensitization of the recipients. CONCLUSION: Compensatory regulatory T-cell function becomes critical in pIgR-deficient mice to avoid the potentially catastrophic effects of systemic immune hyperreactivity, presumably resulting from defective secretory antibody-mediated immune exclusion of microbial components.
Assuntos
Hipersensibilidade Tardia/imunologia , Tolerância Imunológica/imunologia , Imunidade nas Mucosas/imunologia , Receptores de Imunoglobulina Polimérica/deficiência , Transferência Adotiva , Animais , Ensaio de Imunoadsorção Enzimática , Feminino , Hipersensibilidade Alimentar/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ovalbumina/imunologia , Receptores de Imunoglobulina Polimérica/genética , Receptores de Imunoglobulina Polimérica/imunologia , Linfócitos T Reguladores/imunologiaRESUMO
In healthy individuals, humoral immune responses to allergens consist of serum IgA and IgG4, whereas cellular immune responses are controlled by regulatory T (Treg) cells. In search of new compounds that might prevent the onset of allergies by stimulating this type of immune response, we have focused on the mucosal adjuvant, cholera toxin B (CTB), as it induces the formation of Treg cells and production of IgA. Here, we have found that CTB suppresses the potential of dendritic cells to prime for Th2 responses to inhaled allergen. When we administered CTB to the airways of naïve and allergic mice, it strongly suppressed the salient features of asthma, such as airway eosinophilia, Th2 cytokine synthesis, and bronchial hyperreactivity. This beneficial effect was only transferable to other mice by transfer of B but not of T lymphocytes. CTB caused a transforming growth factor-beta-dependent rise in antigen-specific IgA in the airway luminal secretions, which was necessary for its preventive and curative effect, as all effects of CTB were abrogated in mice lacking the luminal IgA transporting polymeric Ig receptor. Not only do these findings show a novel therapeutic avenue for allergy, they also help to explain the complex relationship between IgA levels and risk of developing allergy in humans.
Assuntos
Adjuvantes Imunológicos/uso terapêutico , Toxina da Cólera/uso terapêutico , Células Dendríticas/imunologia , Hipersensibilidade/terapia , Imunoglobulina A Secretora/imunologia , Transferência Adotiva , Alérgenos/imunologia , Animais , Linfócitos B/imunologia , Citocinas/biossíntese , Citocinas/imunologia , Modelos Animais de Doenças , Feminino , Hipersensibilidade/imunologia , Imunoglobulina A Secretora/biossíntese , Camundongos , Camundongos Endogâmicos BALB C , Ovalbumina/imunologia , Linfócitos T Reguladores/imunologia , Células Th2/imunologia , Fator de Crescimento Transformador beta/imunologiaRESUMO
Most tumour-associated antigens (TAA) are non-mutated self-antigens. The peripheral T cell repertoire is devoid of high-avidity TAA-specific cytotoxic T lymphocytes (CTL) due to self-tolerance. As tolerance is major histocompatibility complex-restricted, T cells may be immunized against TAA presented by a non-self human leucocyte antigen (HLA) molecule and transferred to cancer patients expressing that HLA molecule. Obtaining allo-restricted CTL of high-avidity and low cross-reactivity has, however, proven difficult. Here, we show that dendritic cells transfected with mRNA encoding HLA-A*0201, efficiently present externally loaded peptides from the antigen, Melan-A/MART-1 to T cells from HLA-A*0201-negative donors. CD8(+) T cells binding HLA-A*0201/MART-1 pentamers were detected already after 12 days of co-culture in 11/11 donors. The majority of cells from pentamer(+) cell lines were CTL and efficiently killed HLA-A*0201(+) melanoma cells, whilst sparing HLA-A*0201(+) B-cells. Allo-restricted CTL specific for peptides from the leukaemia-associated antigens CD33 and CD19 were obtained with comparable efficiency. Collectively, the results show that dendritic cells engineered to express defined allo-HLA peptide complexes are highly efficient in generating CTL specifically reacting with tumour-associated antigens.
Assuntos
Antígenos de Neoplasias/imunologia , Células Dendríticas/imunologia , Antígenos HLA-A/imunologia , Isoantígenos/imunologia , Linfócitos T Citotóxicos/imunologia , Apresentação de Antígeno/imunologia , Linhagem Celular Tumoral , Citotoxicidade Imunológica , Citometria de Fluxo , Antígenos HLA-A/genética , Antígeno HLA-A2 , Humanos , Imunoterapia/métodos , Ativação Linfocitária/imunologia , Peptídeos/imunologia , Reação em Cadeia da Polimerase , TransfecçãoRESUMO
The production of immunoglobulin A (IgA) in mammals exceeds all other isotypes, and it is mostly exported across mucous membranes. The discovery of IgA and the realization that it dominates humoral mucosal immunity, in contrast to the IgG dominance of the systemic immune system, was early evidence for the distinct nature of mucosal immunology. It is now clear that IgA can function in high-affinity modes for neutralization of toxins and pathogenic microbes, and as a low-affinity system to contain the dense commensal microbiota within the intestinal lumen. The basic map of induction of IgA B cells in the Peyer's patches, which then circulate through the lymph and bloodstream to seed the mucosa with precursors of plasma cells that produce dimeric IgA for export through the intestinal epithelium, has been known for more than 30 years. In this review, we discuss the mechanisms underlying selective IgA induction of mucosal B cells for IgA production and the immune geography of their homing characteristics. We also review the functionality of secretory IgA directed against both commensal organisms and pathogens.
Assuntos
Formação de Anticorpos/imunologia , Imunidade nas Mucosas/imunologia , Imunoglobulina A/imunologia , Nódulos Linfáticos Agregados/imunologia , Plasmócitos/imunologia , Células Precursoras de Linfócitos B/imunologia , Animais , Humanos , Mucosa Intestinal/imunologia , Mucosa Intestinal/microbiologiaRESUMO
BACKGROUND: It has been suggested that allergic diseases are caused by defective suppression of allergen-specific Th2 cells by CD4(+)CD25(+) regulatory T cells. However, such studies have been hampered by the difficulty in distinguishing regulatory T cells from CD25-expressing activated T cells. Recently, it was shown that conventional T cells expressed high levels of CD127, whereas regulatory T cells were CD127(lo), allowing discrimination between these distinct T cell subpopulations. OBJECTIVE: The aim of this study was to study whether the putative regulatory subset defined as CD4(+)CD25(+)CD127(lo) was involved in grass pollen-reactive T cell responses. METHODS: Peripheral blood mononuclear cells (PBMCs) were obtained from allergic donors and non-atopic controls out of season. Grass pollen-induced cytokine production and proliferation were compared in cultures of undepleted cells and cells depleted of CD4(+)CD25(+), CD4(+)CD25(+)CD127(hi) or CD4(+)CD25(+)CD127(lo) T cells. RESULTS: Undepleted cell cultures from allergic patients showed significantly increased proliferation and Th2 cytokine production compared with non-atopic controls. Depletion of all CD25(+) T cells did not increase cytokine production or proliferation, and more importantly, no increase in Th2 cytokine production or proliferation was observed in cell cultures depleted of CD4(+)CD25(+)CD127(lo) cells (putative regulatory T cells) compared with undepleted PBMCs in both the allergic and the non-atopic group. CONCLUSION: Our study showed that T cells from grass pollen-allergic patients and non-atopic controls responded very differently to grass pollen extract, but this difference could not be explained by differences in regulatory T cell function. Further studies are needed to understand the importance of regulatory T cells in allergy.
Assuntos
Antígenos de Plantas/imunologia , Subunidade alfa de Receptor de Interleucina-7/análise , Ativação Linfocitária/imunologia , Rinite Alérgica Sazonal/imunologia , Linfócitos T Reguladores/imunologia , Linfócitos T/imunologia , Adulto , Proliferação de Células , Feminino , Humanos , Interferon gama/metabolismo , Interleucina-13/metabolismo , Subunidade alfa de Receptor de Interleucina-2/análise , Interleucina-5/metabolismo , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Teste de Cultura Mista de Linfócitos , Masculino , Pessoa de Meia-Idade , Phleum/imunologia , Pólen/imunologia , Subpopulações de Linfócitos T/química , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Linfócitos T/metabolismo , Linfócitos T Reguladores/química , Linfócitos T Reguladores/metabolismo , Adulto JovemRESUMO
BACKGROUND: Microarray Comparative Genomic Hybridization (array CGH) provides a means to examine DNA copy number aberrations. Various platforms, brands and underlying technologies are available, facing the user with many choices regarding platform sensitivity and number, localization, and density distribution of probes. RESULTS: We evaluate three different platforms presenting different nature and arrangement of the probes: The Agilent Human Genome CGH Microarray 44 k, the ROMA/NimbleGen Representational Oligonucleotide Microarray 82 k, and the Illumina Human-1 Genotyping 109 k BeadChip, with Agilent being gene oriented, ROMA/NimbleGen being genome oriented, and Illumina being genotyping oriented. We investigated copy number changes in 20 human breast tumor samples representing different gene expression subclasses, using a suite of graphical and statistical methods designed to work across platforms. Despite substantial differences in the composition and spatial distribution of probes, the comparison revealed high overall concordance. Notably however, some short amplifications and deletions of potential biological importance were not detected by all platforms. Both correlation and cluster analysis indicate a somewhat higher similarity between ROMA/NimbleGen and Illumina than between Agilent and the other two platforms. The programs developed for the analysis are available from http://www.ifi.uio.no/bioinf/Projects/. CONCLUSION: We conclude that platforms based on different technology principles reveal similar aberration patterns, although we observed some unique amplification or deletion peaks at various locations, only detected by one of the platforms. The correct platform choice for a particular study is dependent on whether the appointed research intention is gene, genome, or genotype oriented.
Assuntos
Neoplasias da Mama/genética , Dosagem de Genes , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Algoritmos , Aberrações Cromossômicas , Análise por Conglomerados , Bases de Dados Genéticas , Feminino , Perfilação da Expressão Gênica , Genoma Humano , Humanos , Masculino , Sondas de Oligonucleotídeos , Curva ROC , Sensibilidade e EspecificidadeRESUMO
beta-Glucans are glucose polymers with a variety of stimulatory effects on the immune system. The objective of this study was to determine the effect of prophylactic oral administration of soluble Saccharomyces cerevisiae-derived beta-1,3/1,6-glucan (SBG) on the outcome of experimental endotoxaemia and shock-associated organ injury. Male Wistar rats were pretreated with SBG orally (SBGpo, 20 mg/kg/day) for 14 days, subcutaneously (SBGsc, 2 mg/kg/day) for 3 days, or vehicle (placebo). Rats were anaesthetized and subjected to endotoxaemia by intravenous infusion of Escherichia coli lipopolysaccharide (LPS) (6 mg/kg) or saline infusion (sham). We observed significant levels of plasma beta-glucan in the SBGpo group (P<0 x 5), although the SBGsc group had levels approximately 40-fold higher despite a 10-fold lower dose. SBG prophylaxis caused enhanced blood pressure recovery following LPS-induced blood pressure collapse. Oral treatment with SBG attenuated the LPS-induced rise in plasma creatinine levels (P<0 x 05), indicating protection against renal injury. SBG also attenuated the plasma levels of aspartate aminotransferase and alanine aminotransferase (SBGpo, P<0 x 01; SBGsc, P<0 x 01), indicating protection against LPS-induced hepatic injury. A moderate increase in baseline interleukin (IL)-1beta levels was observed in the SBGsc group (P< 0 x 05). In the LPS-challenged rats, plasma levels of proinflammatory cytokines was moderately reduced in both SBG-treated groups compared to placebo. SBG treatment, particularly oral administration, had a striking effect on the haemodynamics of LPS-treated rats, although only a minute fraction of the orally administered beta-glucan translocated to the circulation. Enhanced organ perfusion may thus be responsible for the attenuated levels of indicators of kidney and liver injury seen in SBG-treated rats.
Assuntos
Endotoxemia/prevenção & controle , Insuficiência de Múltiplos Órgãos/prevenção & controle , Choque Séptico/prevenção & controle , beta-Glucanas/administração & dosagem , Administração Oral , Animais , Pressão Sanguínea/efeitos dos fármacos , Citocinas/sangue , Endotoxemia/induzido quimicamente , Endotoxemia/fisiopatologia , Injeções Subcutâneas , Lipopolissacarídeos , Masculino , Insuficiência de Múltiplos Órgãos/induzido quimicamente , Insuficiência de Múltiplos Órgãos/fisiopatologia , Ratos , Ratos Wistar , Saccharomyces cerevisiae , Choque Séptico/induzido quimicamente , Choque Séptico/fisiopatologia , beta-Glucanas/sangue , beta-Glucanas/uso terapêuticoRESUMO
Increased levels of interferon-gamma (IFN-gamma) transcripts have previously been found in duodenal biopsy specimens from patients with untreated coeliac disease (CD). Such samples and duodenal control mucosa were therefore studied to locate and phenotype cells spontaneously secreting IFN-gamma. Specimens were collected from consecutively recruited patients with untreated (seven), treated (four) or refractory (three) CD and from five histologically normal controls. Morphological and immunohistochemical examinations were performed, and epithelial and lamina propria cell suspensions were prepared from parallel samples. Unstimulated viable cells secreting IFN-gamma were identified and phenotyped with a new fluorescence-activated cell sorter-based assay, and IFN-gamma messenger RNA (mRNA) was analysed in snap-frozen aliquots of the same suspensions. Untreated CD cases had the highest fraction of IFN-gamma+ cells in the epithelial compartment (median 2.6%, range 1.6-6.2%) and, less strikingly, in the lamina propria compartment (1.6%, range 0.3-3.6%), followed by refractory (1.4%, 1.0-1.9%; and 0.3%, 0.0-1.2%) and treated (0.8%, 0.5-0.9%; and 0.7%, 0.2-1.1%) disease and finally the controls (0.5%, 0.3-0.9%; and 0.2%, 0.1-0.7%). IFN-gamma mRNA data supported these findings. IFN-gamma+ intraepithelial lymphocytes were mostly CD3+ and CD8+, whereas many positive lamina propria cells were CD8-. We conclude that isolated T cells spontaneously secreting IFN-gamma localize preferentially in the epithelium of patients with classical and refractory CD.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Doença Celíaca/imunologia , Interferon gama/biossíntese , Mucosa Intestinal/imunologia , Adolescente , Adulto , Idoso , Antígenos CD/análise , Antígenos CD/imunologia , Doença Celíaca/classificação , Doença Celíaca/diagnóstico , Células Cultivadas , Epitélio/imunologia , Feminino , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Imunofenotipagem , Interferon gama/genética , Interferon gama/metabolismo , Mucosa Intestinal/citologia , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/análise , Subpopulações de Linfócitos T/classificação , Subpopulações de Linfócitos T/imunologiaRESUMO
BACKGROUND AND AIMS: In mice, the B lymphocyte chemoattractant (BLC) CXC chemokine ligand 13 (CXCL13) is sufficient to induce a series of events leading to the formation of organised lymphoid tissue. Its receptor, CXCR5, is required for normal development of secondary lymphoid tissue. However, the human counterpart, B cell attracting chemokine 1 (BCA-1) has only been detected in the stomach and appendix and not in other parts of normal or diseased gut. Hence to elucidate the potential role of this chemokine and its receptor in human gut associated lymphoid tissue (GALT), we analysed their expression in normal intestine and ulcerative colitis (UC). METHODS: Frozen sections of surgical specimens were studied by multicolour immunofluorescence staining, in situ mRNA hybridisation, and reverse transcription-polymerase chain reaction. RESULTS: BCA-1 mRNA was detected in all normal colonic and UC specimens. BCA-1 was produced and accumulated in relation to peripheral dendritic elements of lymphoid follicles in Peyer's patches and normal colon, as well as in irregular lymphoid aggregates in UC lesions. BCA-1 was partially associated with the traditional follicular dendritic cell phenotype but also with extracellular fibrils in GALT structures. CXCR5 protein was expressed by mantle zone B cells and appeared at a high level on scattered germinal centre T cells. CONCLUSIONS: BCA-1 and CXCR5 are expressed in normal GALT structures as well as in irregular lymphoid aggregates in UC. This strongly suggests that BCA-1 plays an important role not only in the formation of normal GALT but also in the generation of aberrant lymphoid tissue in inflammatory bowel disease.
Assuntos
Quimiocinas CXC/metabolismo , Colite Ulcerativa/metabolismo , Nódulos Linfáticos Agregados/metabolismo , Receptores de Citocinas/metabolismo , Adulto , Linfócitos B/metabolismo , Quimiocina CXCL13 , Criança , Colite Ulcerativa/imunologia , Colo/metabolismo , Feminino , Imunofluorescência , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo , Receptores CXCR5 , Receptores de Quimiocinas , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T/metabolismoRESUMO
BACKGROUND: Refractory sprue is defined as primary or secondary failure to respond to a gluten free diet in patients with coeliac disease-like enteropathy and may signify cryptic or overt enteropathy associated T cell lymphoma. AIMS: To study in detail jejunal morphology and immunophenotypes in patients with refractory sprue in the search for features that might be useful to predict prognosis. PATIENTS: Seven patients are described, representing all such cases identified in our hospital over a 13 year period. METHODS: Biopsy and/or surgical resection specimens were examined by morphology, immunohistochemistry, including enzymatic and immunofluorescent detection, and molecular biology. RESULTS: All patients had phenotypically abnormal intraepithelial lymphocytes (IELs) that lacked CD8, T cell receptor alpha beta (or gamma delta), and/or expressed CD30 in addition to variable expression of the natural killer cell receptor CD94. A monoclonal T cell population was present in six cases, data from the seventh being inconclusive. Three patients had overt lymphoma with CD30+ tumour tissue intervening between intact mucosa that contained neoplastic IELs. Intriguingly, CD30+ IELs were observed both a long way away from, and in direct continuity with, the tumours in these patients. Such CD30+ cells were hardly detected in patients without tumours, two of which are in good health several years after the initial diagnosis. CONCLUSIONS: Our data suggest that abnormal IELs in patients with refractory sprue are phenotypically heterogeneous. CD30 expression by these cells may indicate a worse prognosis, including the occurrence of overt lymphoma.
Assuntos
Doença Celíaca/genética , Doença Celíaca/patologia , Jejuno/patologia , Antígeno Ki-1/imunologia , Linfócitos T/patologia , Adulto , Idoso , Antígenos CD/genética , Antígenos CD/imunologia , Doença Celíaca/imunologia , Epitélio/imunologia , Epitélio/patologia , Evolução Fatal , Feminino , Expressão Gênica , Humanos , Jejuno/imunologia , Antígeno Ki-1/genética , Linfoma/epidemiologia , Linfoma/genética , Masculino , Pessoa de Meia-Idade , Fenótipo , Reação em Cadeia da Polimerase , Linfócitos T/imunologiaRESUMO
Secretory Abs constitute the first line of specific immune defense at mucosal surfaces. Such Abs are generated by the active transport of polymeric Ig (pIg) across secretory epithelia mediated by the pIgR, also known as transmembrane secretory component (SC). The proinflammatory cytokine TNF-alpha is a key mediator of host responses to infections, and it can stimulate protein synthesis-dependent transcriptional up-regulation of pIgR/SC in the HT-29 intestinal adenocarcinoma cell line. By reporter gene assay we identified a novel TNF-alpha-responsive region located within a 748-bp fragment in intron 1 of the human pIgR/SC gene which depended on an NF-kappaB/Rel site for full responsiveness. EMSAs demonstrated preferential binding of the NF-kappaB/Rel family member p65 (RelA) to this DNA element after TNF-alpha stimulation, with weaker and more delayed binding of p50. Furthermore, the TNF-alpha-responsive region in intron 1 required cooperation with DNA elements located in the proximal promoter region of the gene. Mutational analysis demonstrated that an IFN-stimulated response element near the transcriptional start site in exon 1 was involved in the TNF-alpha responsiveness. Thus, DNA elements located >4 kb apart were found to cooperate in TNF-alpha-induced pIgR/SC up-regulation. The intronic TNF-alpha-responsive enhancer overlapped with a recently identified IL-4-responsive enhancer. Several intronic DNA elements found to be functionally important in the human gene are highly conserved between the human and mouse pIgR/SC genes, suggesting the presence of a conserved cytokine-responsive enhancer region.
Assuntos
Íntrons/imunologia , NF-kappa B/genética , Regiões Promotoras Genéticas/imunologia , Proteínas Proto-Oncogênicas c-rel/genética , Receptores de Imunoglobulina Polimérica/genética , Transcrição Gênica/imunologia , Fator de Necrose Tumoral alfa/fisiologia , Composição de Bases , Sítios de Ligação/genética , Sítios de Ligação/imunologia , Sequência Consenso , Regulação da Expressão Gênica/imunologia , Células HT29 , Humanos , Interferon gama/genética , Íntrons/fisiologia , NF-kappa B/metabolismo , NF-kappa B/fisiologia , Subunidade p50 de NF-kappa B , Regiões Promotoras Genéticas/fisiologia , Proteínas Proto-Oncogênicas c-rel/metabolismo , Proteínas Proto-Oncogênicas c-rel/fisiologia , Receptores de Imunoglobulina Polimérica/metabolismo , Elementos de Resposta/imunologia , Componente Secretório/genética , Deleção de Sequência , Fator de Transcrição RelA , TransfecçãoAssuntos
Imunoglobulina A/imunologia , Imunoglobulina M/imunologia , Receptores de Imunoglobulina Polimérica/imunologia , Componente Secretório/imunologia , Animais , Sítios de Ligação , Humanos , Região de Junção de Imunoglobulinas/imunologia , Transdução de Sinais/imunologia , Especificidade da EspécieRESUMO
Local production of secretory (S)IgA provides adaptive immunologic protection of mucosal surfaces, but SIgA is also protective when administered passively, such as in breast milk. Therefore, SIgA is a potential candidate for therapeutic administration, but its complex structure with four different polypeptide chains produced by two distinct cell types complicates recombinant production. The J chain is critical in the structure of SIgA because it is required for efficient polymerization of IgA and for the affinity of such polymers to the secretory component (SC)/polymeric (p)IgR. To better understand the role of the J chain in SIgA production, we have generated various mutant forms of the human J chain and analyzed the function of these mutants when coexpressed with IgA. We found that the C terminus of the J chain was not required for the formation of IgA polymers, but was essential for the binding of pIgA to SC. Likewise, we found that two of the intrachain disulfide bridges (Cys(13):Cys(101) and Cys(109):Cys(134)) were also required for the binding of pIgA to SC but, interestingly, not for IgA polymerization. Conversely, the last intrachain disulfide bridge (Cys(72):Cys(92)) was not essential for either of these two J chain functions. Finally, we demonstrated that the presence of only Cys(15) or Cys(69) was sufficient to support polymerization of IgA, but that these polymers were mostly noncovalently stabilized. Nevertheless, these polymers bound free SC with nearly the same affinity as pIgA containing wild-type J chain, but were transcytosed by pIgR-expressing polarized epithelial cells at a reduced efficiency.