RESUMO
The bone marrow is a specialized niche responsible for the maintenance of hematopoietic stem and progenitor cells during homeostasis and inflammation. Recent studies, however, have extended this essential role to the extramedullary and extravascular lung microenvironment. Here, we provide further evidence for a reservoir of hematopoietic stem and progenitor cells within the lung from Embryonic Day 18.5 until adulthood. These lung progenitors display distinct microenvironment-specific developmental kinetics compared with their bone marrow counterparts, exemplified by a rapid shift from a common myeloid to a megakaryocyte-erythrocyte progenitor-dominated niche with increasing age. In adult mice, influenza A viral infection results in a transient reduction in multipotent progenitors within the lungs, with a parallel increase in downstream granulocyte-monocyte progenitors and dendritic cell populations associated with acute viral infections. Our findings suggest that lung hematopoietic progenitors play a role in reestablishing immunological homeostasis in the respiratory mucosa, which may have significant clinical implications for maintaining pulmonary health after inflammatory perturbation.
Assuntos
Células-Tronco Hematopoéticas , Vírus da Influenza A , Pulmão , Camundongos Endogâmicos C57BL , Infecções por Orthomyxoviridae , Animais , Células-Tronco Hematopoéticas/virologia , Células-Tronco Hematopoéticas/metabolismo , Pulmão/virologia , Pulmão/patologia , Infecções por Orthomyxoviridae/virologia , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/patologia , Camundongos , CinéticaRESUMO
Persistent regional and systemic inflammation may promote pain and hyperalgesia in complex regional pain syndrome (CRPS). In this study, we investigated whether stimulation of α1-adrenoceptors (α1-AR) on peripheral blood mononuclear cells (PBMC) might contribute to this inflammatory state. PBMC were isolated from venous blood collected from 21 CRPS patients and 21 sex and age-matched controls. Lipopolysaccharide (LPS), a bacterial toxin, was administered to cultured PBMC for 24 h to trigger inflammation. Exposure to LPS resulted in heightened gene expression of α1-AR subtype B (α1B-AR) in PBMC of CRPS patients relative to controls. Interleukin (IL)-1ß and IL-6 levels did not change when the α1-AR agonist phenylephrine was administered to naïve PBMC. However, α1-AR stimulation following LPS treatment increased IL-6 mRNA and protein levels in PBMC of patients and controls. To investigate the possible consequence of heightened IL-6 levels on immunoglobulin G antibody production, PBMC were stimulated with CD40 ligand and IL-21 to generate plasmablasts (B cells that secrete antibodies). This response was similar in patients and controls. Adding IL-6 to the cell culture medium increased plasmablast differentiation in controls and antibody production both in patients and controls. These findings suggest that the inflammatory cascade associated with elevated levels of IL-6 may generate α1B-AR expression in CRPS PBMC. A reciprocal interaction between heightened α1-AR expression in PBMC and IL-6 secretion may contribute to systemic inflammation and antibody production in CRPS.
Assuntos
Síndromes da Dor Regional Complexa , Leucócitos Mononucleares , Humanos , Interleucina-6 , Lipopolissacarídeos/farmacologia , Inflamação , Interleucina-1beta , Receptores AdrenérgicosRESUMO
BACKGROUND: Influenza A virus (IAV) is the only influenza virus causing flu pandemics (i.e., global epidemics of flu disease). Influenza (the flu) is a highly contagious disease that can be deadly, especially in high-risk groups. Worldwide, these annual epidemics are estimated to result in about 3 to 5 million cases of severe illness and in about 290,000 to 650,000 respiratory deaths. We intend to reveal the effect of IAV infection on the host's metabolism, immune response, and neurotoxicity by using a mouse IAV infection model. METHODS: 51 metabolites of murine blood plasma (33 amino acids/amino acid derivatives (AADs) and 18 metabolites of the tryptophan pathway) were analyzed by using Ultra-High-Performance Liquid Chromatography-Mass Spectrometry with Electrospray Ionization at the acute (7 days post-infection (dpi)), resolution (14 dpi), and recovery (21 dpi) stages of the virus infection in comparison with controls. RESULTS: Among the 33 biogenic amino acids/AADs, the levels of five amino acids/AADs (1-methylhistidine, 5-oxoproline, α-aminobutyric acid, glutamine, and taurine) increased by 7 dpi, whereas the levels of ten amino acids/AADs (4-hydroxyproline, alanine, arginine, asparagine, cysteine, citrulline, glycine, methionine, proline, and tyrosine) decreased. By 14 dpi, the levels of one AAD (3-methylhistidine) increased, whereas the levels of five amino acids/AADs (α-aminobutyric acid, aminoadipic acid, methionine, threonine, valine) decreased. Among the 18 metabolites from the tryptophan pathway, the levels of kynurenine, quinolinic acid, hydroxykynurenine increased by 7 dpi, whereas the levels of indole-3-acetic acid and nicotinamide riboside decreased. CONCLUSIONS: Our data may facilitate understanding the molecular mechanisms of host responses to IAV infection and provide a basis for discovering potential new mechanistic, diagnostic, and prognostic biomarkers and therapeutic targets for IAV infection.
Assuntos
Vírus da Influenza A , Influenza Humana , Animais , Camundongos , Humanos , Triptofano , Aminoácidos/metabolismo , Metionina , Vírus da Influenza A/metabolismoRESUMO
A positive feedback loop between inflammatory cytokines and alpha1-adrenoceptors (α1-AR) (a target of the sympathetic nervous system neurotransmitter norepinephrine) influences inflammatory responses in immune cells. This cross-talk between the sympathetic nervous system and immune system may play a role in promoting chronic inflammation. Emerging evidence shows that α1-AR interact with inflammatory cytokines in keratinocytes, and this epidermal adrenergic signalling may contribute to skin inflammatory responses following injury, disease or stress. In this study, utilizing an in vitro approach, we hypothesized that α1-AR interact in a positive feedback loop with inflammatory mediators in keratinocytes. The pro-inflammatory cytokine tumor necrosis factor α (TNFα) was used to induce an inflammatory state in cultured keratinocytes. TNFα increased interleukin (IL)-1ß, IL-6, IL-8 and nerve growth factor (NGF) production and gene expression levels of α1-AR subtype B (α1B-AR). Additional stimulation of α1-AR further increased IL-6 levels, while maintaining high levels of IL-8 and decreasing levels of IL-1ß and NGF. Our results suggest that reciprocal influences between α1-ARs and inflammatory cytokines may play a role in normal inflammatory responses. However, if unchecked, this cycle could contribute to pathology (e.g. chronic inflammatory diseases, chronic pain conditions, and stress-induced cancer progression).
Assuntos
Citocinas/metabolismo , Retroalimentação , Mediadores da Inflamação/metabolismo , Inflamação/imunologia , Queratinócitos/metabolismo , Receptores Adrenérgicos alfa 1/química , Agonistas de Receptores Adrenérgicos alfa 1/farmacologia , Células Cultivadas , Humanos , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Inflamação/patologia , Queratinócitos/citologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/imunologia , Fenilefrina/farmacologia , Receptores Adrenérgicos alfa 1/metabolismo , Transdução de Sinais , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
Keratinocytes produce cytokines and nerve growth factor (NGF) as part of a repair response to injury, disease or stress, and express alpha1-adrenoceptors (α1-AR). The expression of these receptors is elevated in some inflammatory diseases and chronic pain conditions. In this study, we investigated whether inflammatory signalling affects α1-AR expression in keratinocytes in vitro. Tumor necrosis factor α (TNFα) was administered to human keratinocytes, after which the levels of other key pro-inflammatory cytokines and NGF were measured. The production of these cytokines and NGF increased in cells treated with TNFα compared to untreated cells. Furthermore, exposure to TNFα increased gene expression of the α1-AR subtype B in keratinocytes. Our results suggest that inflammatory cytokines released during injury stimulate α1-AR expression in keratinocytes. The up-regulation of α1-AR may amplify the adrenergic sensitivity of these cells to catecholamines released during sympathetic nervous system activation after injury which, in turn, could heighten the inflammatory response.
Assuntos
Citocinas/metabolismo , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Fatores de Crescimento Neural/metabolismo , Receptores Adrenérgicos alfa 1/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Adulto , Linhagem Celular , Citocinas/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Humanos , Inflamação/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Interleucina-8/genética , Interleucina-8/metabolismo , Pessoa de Meia-Idade , Fator de Crescimento Neural/genética , Fator de Crescimento Neural/metabolismo , Receptores Adrenérgicos alfa 1/genéticaRESUMO
BACKGROUND: Low vitamin D levels have been associated with allergic diseases. Vitamin D has potent immunomodulatory properties, but the mechanisms remain unclear. We have investigated the effect of oral vitamin D supplementation on circulating immune cell phenotypes in infants. METHOD: A double-blinded randomised controlled trial was conducted to investigate the effect of oral vitamin D supplementation (400 IU/d) on eczema and immune development. A subset of 78 infants was included in this analysis. Phenotypic analysis of immune cell subsets was performed using flow cytometry. RESULTS: Vitamin D supplementation resulted in median 25(OH)D levels of 80.5 vs 59.5 nmol/L in the placebo group at 3 months of age (P = .002) and 87.5 vs 77 nmol/L at 6 months of age (P = .08). We observed significant changes in immune cell composition from birth (cord blood) to 6 months of age. Vitamin D supplementation did not impact these changes, nor did immune cell composition correlate with plasma 25(OH)D levels. Through exploratory analysis, we identified possible associations with eczema development and increased abundance of naïve CD4- T cells at birth, as well as associations with basophils, iNKT and central memory CD4+ T cells, and altered expression patterns of IgE receptor (FcεR1) on monocytes and dendritic cells with eczema at 6 months. CONCLUSIONS: Vitamin D supplementation in infants who were vitamin D sufficient at birth did not affect developmental changes in immune cells during the first 6 months of life. However, immune cell profiles at birth and at 6 months of age were associated with early life eczema.
Assuntos
Eczema , Deficiência de Vitamina D , Colecalciferol , Suplementos Nutricionais , Método Duplo-Cego , Feminino , Humanos , Lactente , Recém-Nascido , Vitamina D , Deficiência de Vitamina D/tratamento farmacológico , VitaminasRESUMO
BACKGROUND: Influenza virus infection during pregnancy is associated with enhanced disease severity. However, the underlying mechanisms are still not fully understood. We hypothesized that normal alveolar macrophage (AM) functions, which are central to maintaining lung immune homeostasis, are altered during pregnancy and that this dysregulation contributes to the increased inflammatory response to influenza virus infection. METHODS: Time-mated BALB/c mice were infected with a low dose of H1N1 influenza A virus at gestation day 9.5. Inflammatory cells in bronchoalveolar lavage (BAL) fluid were assessed by flow cytometry. RESULTS: Our findings confirm previous reports of increased severity of influenza virus infection in pregnant mice. The heightened inflammatory response detected in BAL fluid from infected pregnant mice was characterized by neutrophil-rich inflammation with concomitantly reduced numbers of AM, which were slower to return to baseline counts, compared with nonpregnant infected mice. The increased infection severity and inflammatory responses to influenza during pregnancy were associated with a pregnancy-induced shift in AM phenotype at homeostatic baseline, from the M1 (ie, classical activation) state toward the M2 (ie, alternative activation) state, as evidence by increased expression of CD301 and reduced levels of CCR7. CONCLUSION: These results show that pregnancy is associated with an alternatively activated phenotype of AM before infection, which may contribute to heightened disease severity.
Assuntos
Vírus da Influenza A Subtipo H1N1/imunologia , Influenza Humana/virologia , Animais , Líquido da Lavagem Broncoalveolar/virologia , Modelos Animais de Doenças , Feminino , Humanos , Influenza Humana/imunologia , Pulmão/imunologia , Pulmão/virologia , Macrófagos Alveolares/imunologia , Macrófagos Alveolares/virologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fenótipo , GravidezRESUMO
Respiratory IgE-sensitization to innocuous antigens increases the risk for developing diseases such as allergic asthma. Dendritic cells (DC) residing in the airways orchestrate the immune response following antigen exposure and their ability to sample and present antigens to naïve T cells in airway draining lymph nodes contributes to allergen-specific IgE-sensitization. In order to characterize inhaled antigen capture and presentation by DC subtypes in vivo, we used an adjuvant-free respiratory sensitization model using two genetically distinct rat strains, one of which is naturally resistant and the other naturally susceptible to allergic sensitization. Upon multiple exposures to ovalbumin (OVA), the susceptible strain developed OVA-specific IgE and airway inflammation, whereas the resistant strain did not. Using fluorescently tagged OVA and flow cytometry, we demonstrated significant differences in antigen uptake efficiency and presentation associated with either IgE-sensitization or resistance to allergen exposures in respective strains. We further identified CD4+ conventional DC (cDC) as the subset involved in airway antigen sampling in both strains, however, CD4+ cDC in the susceptible strain were less efficient in OVA sampling and displayed increased MHC-II expression compared with the resistant strain. This was associated with generation of an exaggerated Th2 response and a deficiency of airway regulatory T cells in the susceptible strain. These data suggest that subsets of cDC are able to induce either sensitization or resistance to inhaled antigens as determined by genetic background, which may provide an underlying basis for genetically determined susceptibility to respiratory allergic sensitization and IgE production in susceptible individuals.
Assuntos
Células Dendríticas/imunologia , Imunização , Imunoglobulina E/imunologia , Pulmão/imunologia , Animais , Hipersensibilidade/imunologia , Hipersensibilidade/patologia , Inflamação/patologia , Linfonodos/patologia , Ovalbumina/imunologia , Fenótipo , Ratos Endogâmicos BN , Linfócitos T Reguladores/imunologiaRESUMO
IgE sensitisation has increased significantly over the last decades and is a crucial factor in the development of allergic diseases. IgE antibodies are produced by B cells through the process of antigen presentation by dendritic cells, subsequent differentiation of CD4⺠Th2 cells, and class switching in B cells. However, many of the factors regulating these processes remain unclear. These processes affect males and females differently, resulting in a significantly higher prevalence of IgE sensitisation in males compared to females from an early age. Before the onset of puberty, this increased prevalence of IgE sensitisation is also associated with a higher prevalence of clinical symptoms in males; however, after puberty, females experience a surge in the incidence of allergic symptoms. This is particularly apparent in allergic asthma, but also in other allergic diseases such as food and contact allergies. This has been partly attributed to the pro- versus anti-allergic effects of female versus male sex hormones; however, it remains unclear how the expression of sex hormones translates IgE sensitisation into clinical symptoms. In this review, we describe the recent epidemiological findings on IgE sensitisation in male and females and discuss recent mechanistic studies casting further light on how the expression of sex hormones may influence the innate and adaptive immune system at mucosal surfaces and how sex hormones may be involved in translating IgE sensitisation into clinical manifestations.
Assuntos
Doença , Imunoglobulina E/imunologia , Animais , Feminino , Hormônios Esteroides Gonadais/metabolismo , Humanos , Hipersensibilidade/imunologia , Hipersensibilidade/terapia , Masculino , Pesquisa Translacional BiomédicaRESUMO
The pulmonary parenchymal and mucosal microenvironments are constantly exposed to the external environment and thus require continuous surveillance to maintain steady-state immunological homeostasis. This is achieved by a mobile network of pulmonary dendritic cells (DC) and macrophages (mø) that constantly sample and process microenvironmental antigens into signals that can initiate or dampen inflammation, either locally or after onward migration to draining lymph nodes. The constant steady-state turnover of pulmonary DC and mø requires replenishment from bone marrow precursors; however, the nature of the pulmonary precursor cell (PC) remains unclear, although recent studies suggest that subsets of pulmonary DC may derive from circulating monocytic precursors. In the current study, we describe a population of cells in steady-state mouse lung tissue that has the surface phenotypic and ultrastructural characteristics of a common DC progenitor. Irradiation and reconstitution studies confirmed the bone marrow origins of this PC and showed that it had rapid depletion and reconstitution kinetics that were similar to those of DC, with a 50% repopulation by donor-derived cells by Days 7-9 after reconstitution. This was significantly faster than the rates observed for mø, which showed 50% repopulation by donor-derived cells beyond Days 16-21 after reconstitution. Purified PC gained antigen-presenting function and a cell surface phenotype similar to that of pulmonary DC after maturation in vitro, with light and electron microscopy confirming a myeloid DC morphology. To the best of our knowledge, this is the first study to describe a PC for DC in lung tissue; the findings have implications for the restoration of pulmonary immunological homeostasis after bone marrow transplant.
Assuntos
Células Dendríticas/citologia , Pulmão/citologia , Células-Tronco/citologia , Animais , Células Apresentadoras de Antígenos/efeitos dos fármacos , Células Apresentadoras de Antígenos/imunologia , Biomarcadores/metabolismo , Transplante de Medula Óssea , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/metabolismo , Epitopos/imunologia , Feminino , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Antígenos de Histocompatibilidade Classe II/metabolismo , Cinética , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Monócitos/citologia , Monócitos/efeitos dos fármacos , Fenótipo , Células-Tronco/efeitos dos fármacosRESUMO
To address how surface charge affects the fate of potential nanocarriers in the lung, gold nanoparticles (AuNPs) coated with polyvinyl alcohol containing either positively (NH2) or negatively (COOH) charged functional groups were intra-nasally instilled in mice, and their uptake by antigen presenting cell populations (APC) in broncho-alveolar lavage (BAL) fluid, trachea, and lung parenchyma, as well as trafficking to the lung draining lymph nodes (LDLNs) was assessed by flow cytometry. Furthermore, CD4+ T cell proliferation in LDLNs was investigated following instillation. All APC subpopulations preferentially captured positively-charged AuNPs compared to their negatively-charged counterparts. Uptake of AuNPs up-regulated expression of co-stimulatory molecules on all APC populations. Furthermore, positively-charged AuNPs induced enhanced OVA-specific CD4+ T cell stimulation in LDLNs compared to negatively-charged AuNPs, or polymer alone. Our findings demonstrate surface charge as a key parameter determining particle uptake by APC, and down-stream immune responses depend on the presence of particle core-bound polymer.
Assuntos
Ouro/administração & dosagem , Pulmão , Ativação Linfocitária , Nanopartículas , Animais , Linfócitos T CD4-Positivos , Proliferação de Células , CamundongosRESUMO
The respiratory tract is an attractive target organ for novel diagnostic and therapeutic applications with nano-sized carriers, but their immune effects and interactions with key resident antigen-presenting cells (APCs) such as dendritic cells (DCs) and alveolar macrophages (AMs) in different anatomical compartments remain poorly understood. Polystyrene particles ranging from 20 nm to 1,000 nm were instilled intranasally in BALB/c mice, and their interactions with APC populations in airways, lung parenchyma, and lung-draining lymph nodes (LDLNs) were examined after 2 and 24 hours by flow cytometry and confocal microscopy. In the main conducting airways and lung parenchyma, DC subpopulations preferentially captured 20-nm particles, compared with 1,000-nm particles that were transported to the LDLNs by migratory CD11blow DCs and that were observed in close proximity to CD3⺠T cells. Generally, the uptake of particles increased the expression of CD40 and CD86 in all DC populations, independent of particle size, whereas 20-nm particles induced enhanced antigen presentation to CD4⺠T cells in LDLNs in vivo. Despite measurable uptake by DCs, the majority of particles were taken up by AMs, irrespective of size. Confocal microscopy and FACS analysis showed few particles in the main conducting airways, but a homogeneous distribution of all particle sizes was evident in the lung parenchyma, mostly confined to AMs. Particulate size as a key parameter determining uptake and trafficking therefore determines the fate of inhaled particulates, and this may have important consequences in the development of novel carriers for pulmonary diagnostic or therapeutic applications.
Assuntos
Apresentação de Antígeno , Células Apresentadoras de Antígenos/imunologia , Movimento Celular , Linfonodos/imunologia , Tamanho da Partícula , Transferência Adotiva , Animais , Antígeno CD11b/imunologia , Linfócitos T CD4-Positivos/imunologia , Antígenos CD40/imunologia , Proliferação de Células , Células Cultivadas , Feminino , Citometria de Fluxo , Pulmão/imunologia , Pulmão/patologia , Macrófagos Alveolares/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Microscopia Confocal , Ovalbumina/imunologia , Poliestirenos/administração & dosagem , Poliestirenos/imunologia , Fatores de TempoRESUMO
Chronic innocuous aeroallergen exposure attenuates CD4(+) T cell-mediated airways hyperresponsiveness in mice; however, the mechanism(s) remain unclear. We examined the role of airway mucosal dendritic cell (AMDC) subsets in this process using a multi-OVA aerosol-induced tolerance model in sensitized BALB/c mice. Aeroallergen capture by both CD11b(lo) and CD11b(hi) AMDC and the delivery of OVA to airway draining lymph nodes by CD8α(-) migratory dendritic cells (DC) were decreased in vivo (but not in vitro) when compared with sensitized but nontolerant mice. This was functionally significant, because in vivo proliferation of OVA-specific CD4(+) T cells was suppressed in airway draining lymph nodes of tolerized mice and could be restored by intranasal transfer of OVA-pulsed and activated exogenous DC, indicating a deficiency in Ag presentation by endogenous DC arriving from the airway mucosa. Bone marrow-derived DC Ag-presenting function was suppressed in multi-OVA tolerized mice, and allergen availability to airway APC populations was limited after multi-OVA exposure, as indicated by reduced OVA and dextran uptake by airway interstitial macrophages, with diffusion rather than localization of OVA across the airway mucosal surface. These data indicate that inhalation tolerance limits aeroallergen capture by AMDC subsets through a mechanism of bone marrow suppression of DC precursor function coupled with reduced Ag availability in vivo at the airway mucosa, resulting in limited Ag delivery to lymph nodes and hypoproliferation of allergen-specific CD4(+) T cells.
Assuntos
Alérgenos/administração & dosagem , Linfócitos T CD4-Positivos/imunologia , Proliferação de Células , Células Dendríticas/imunologia , Epitopos de Linfócito T/imunologia , Tolerância Imunológica , Ovalbumina/administração & dosagem , Fragmentos de Peptídeos/administração & dosagem , Mucosa Respiratória/imunologia , Administração por Inalação , Alérgenos/imunologia , Alérgenos/toxicidade , Sequência de Aminoácidos , Animais , Células da Medula Óssea/imunologia , Células da Medula Óssea/metabolismo , Hiper-Reatividade Brônquica/imunologia , Hiper-Reatividade Brônquica/patologia , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/metabolismo , Células Cultivadas , Células Dendríticas/citologia , Células Dendríticas/metabolismo , Modelos Animais de Doenças , Epitopos de Linfócito T/administração & dosagem , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Dados de Sequência Molecular , Ovalbumina/imunologia , Ovalbumina/toxicidade , Fragmentos de Peptídeos/imunologia , Fragmentos de Peptídeos/toxicidade , Mucosa Respiratória/citologia , Mucosa Respiratória/metabolismo , Células-Tronco/citologia , Células-Tronco/imunologia , Células-Tronco/metabolismoRESUMO
ABSTRACT: Alpha-1 adrenoceptors are overexpressed in the epidermis of a subgroup of patients with complex regional pain syndrome (CRPS). Activating α 1 -adrenoceptors in epidermal cells increases production of the proinflammatory cytokine interleukin-6 (IL-6), a mediator of inflammation. To investigate whether this might exacerbate inflammation in CRPS, primary keratinocytes or dermal fibroblasts were cultured from skin biopsies obtained from the affected limb of 25 patients and a similar site in 28 controls. The fundamental proinflammatory cytokine, tumor necrosis factor alpha, was administered for 24 hours to initiate inflammation. After this, cells were incubated for 6 hours with the α 1 -adrenoceptor agonist phenylephrine. Exposure to tumor necrosis factor alpha induced proinflammatory cytokine mRNA production and protein secretion in keratinocytes and fibroblasts and enhanced α 1B -adrenoceptor mRNA expression in keratinocytes. Additional stimulation of α 1 adrenoceptors with phenylephrine increased the production of IL-6 mRNA and protein secretion in both cell types. Under all conditions, gene and protein α 1 -adrenoceptor levels and cytokine gene expression and protein secretion were similar, overall, in patients and controls, except for abnormally high α 1 -adrenoceptor protein levels in the keratinocytes of 3 of 17 patients. These findings suggest that persistent inflammation in CRPS is not due to dysfunction of skin cells but is a normal response to extrinsic signals. After α 1 -adrenoceptor stimulation of keratinocytes, increases in IL-6 mRNA but not protein were proportional to basal α 1 -adrenoceptor protein levels. Skin cells play an important role in persistent inflammation in CRPS. Potentially, a positive feedback loop between α 1 -adrenoceptors and IL-6 production in skin cells contributes to this inflammatory state.
Assuntos
Síndromes da Dor Regional Complexa , Interleucina-6 , Humanos , Citocinas/genética , Citocinas/metabolismo , Inflamação/metabolismo , Interleucina-6/metabolismo , Queratinócitos/metabolismo , Queratinócitos/patologia , Fenilefrina/farmacologia , Receptores Adrenérgicos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Receptores Adrenérgicos alfa 1/metabolismoRESUMO
Respiratory syncytial virus (RSV) causes annual epidemics of infections affecting the whole population. In vitro, it has been shown to infect and persist in human dendritic cells (DCs) for prolonged periods. Initially persistence is associated with low levels of replication before the virus becomes dormant. Reactivation of viral replication can be triggered many months later. Infection of DCs is likely to influence the host's ability to generate effective long-term memory responses. A well-established animal was utilized to confirm that RSV both infects and persists in pulmonary DCs in vivo. Mice were infected with a modified strain of RSV expressing red fluorescent protein (RSV-RFP) when replicating. Clinical symptoms of infection were monitored using weight change and inflammatory cell counts from bronchoalveolar lavage, which correlated with the RSV viral titer (quantitative polymerase chain reaction). Lung tissues were collected at 3, 5, 7, and 21 days postinfection (dpi) to assess leukocyte populations by flow cytometry. Clinical symptoms and RSV viral load peaked at 5 dpi. RSV-RFP was most prevalent in macrophages at 3 dpi and also observed in B cells and DCs. At 21 dpi, RSV-RFP remained evident in a subset of conventional DCs (CD103+CD11b+) even though both clinical symptoms and pulmonary inflammation had resolved. These results confirm that in this well-established mouse model, RSV persists in lung conventional DCs following resolution of the acute infection. Further work is required to explore whether the virus continues with low-level replication before becoming dormant in vivo, as has been described in vitro.
Assuntos
Infecções por Vírus Respiratório Sincicial , Vírus Sincicial Respiratório Humano , Animais , Humanos , Camundongos , Pulmão , Macrófagos , Células Dendríticas , Camundongos Endogâmicos BALB CRESUMO
An important feature of atopic asthma is the T cell-driven late phase reaction involving transient bronchoconstriction followed by development of airways hyperresponsiveness (AHR). Using a unique rat asthma model we recently showed that the onset and duration of the aeroallergen-induced airway mucosal T cell activation response in sensitized rats is determined by the kinetics of functional maturation of resident airway mucosal dendritic cells (AMDCs) mediated by cognate interactions with CD4+ T helper memory cells. The study below extends these investigations to chronic aeroallergen exposure. We demonstrate that prevention of ensuing cycles of T cell activation and resultant AHR during chronic exposure of sensitized rats to allergen aerosols is mediated by CD4+CD25+Foxp3+LAG3+ CTLA+CD45RC+ T cells which appear in the airway mucosa and regional lymph nodes within 24 h of initiation of exposure, and inhibit subsequent Th-mediated upregulation of AMDC functions. These cells exhibit potent regulatory T (T reg) cell activity in both in vivo and ex vivo assay systems. The maintenance of protective T reg activity is absolutely dependent on continuing allergen stimulation, as interruption of exposure leads to waning of T reg activity and reemergence of sensitivity to aeroallergen exposure manifesting as AMDC/T cell upregulation and resurgence of T helper 2 cytokine expression, airways eosinophilia, and AHR.
Assuntos
Asma/imunologia , Hiper-Reatividade Brônquica/imunologia , Hiper-Reatividade Brônquica/prevenção & controle , Ativação Linfocitária/imunologia , Mucosa Respiratória/imunologia , Mucosa Respiratória/patologia , Linfócitos T Reguladores/imunologia , Animais , Asma/patologia , Hiper-Reatividade Brônquica/patologia , Células Cultivadas , Técnicas de Cocultura , Modelos Animais de Doenças , Feminino , Masculino , RatosRESUMO
A significant proportion of chronic obstructive pulmonary disease exacerbations are strongly associated with rhinovirus infection (HRV). In this study, we combined long-term cigarette smoke exposure with HRV infection in a mouse model. Our aim was to better understand the effects of HRV infection on such exacerbations, using a realistic method for generating a COPD-like phenotype. After 12-weeks of cigarette smoke exposure, adult female BALB/c mice were infected with HRV-1A and three days later we assessed a range of outcomes including lung volume and function, collected lung tissue for measurement of viral titre, bronchoalveolar lavage for assessment of pulmonary inflammation and levels of key mediators, and fixed lungs for stereological structural analyses. Cigarette smoke exposure alone significantly increased total cells and macrophages, and reduced MIP-2 in bronchoalveolar lavage. HRV-1A infection alone increased neutrophilic inflammation, IP-10 and total protein in lavage and also increased specific airway resistance measured at functional residual capacity. Cigarette smoke and HRV-1A together impacted various lung structural parameters including increasing stereological lung volume. Our results show that long-term cigarette smoke exposure and HRV-1A infection both individually impact respiratory outcomes and combine to alter aspects of lung structure in a mouse model, thus providing insight into the development of future mechanistic studies and appropriate interventions in human disease.
Assuntos
Fumar Cigarros/efeitos adversos , Exposição por Inalação/efeitos adversos , Infecções por Picornaviridae/complicações , Doença Pulmonar Obstrutiva Crônica/etiologia , Doença Pulmonar Obstrutiva Crônica/patologia , Rhinovirus/patogenicidade , Exacerbação dos Sintomas , Animais , Modelos Animais de Doenças , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Doença Pulmonar Obstrutiva Crônica/imunologia , Doença Pulmonar Obstrutiva Crônica/metabolismoRESUMO
OBJECTIVES: Incomplete maturation of immune regulatory functions at birth is antecedent to the heightened risk for severe respiratory infections during infancy. Our forerunner animal model studies demonstrated that maternal treatment with the microbial-derived immune training agent OM-85 during pregnancy promotes accelerated postnatal maturation of mechanisms that regulate inflammatory processes in the offspring airways. Here, we aimed to provide proof of concept for a novel solution to reduce the burden and potential long-term sequelae of severe early-life respiratory viral infection through maternal oral treatment during pregnancy with OM-85, already in widespread human clinical use. METHODS: In this study, we performed flow cytometry and targeted gene expression (RT-qPCR) analysis on lungs from neonatal offspring whose mothers received oral OM-85 treatment during pregnancy. We next determined whether neonatal offspring from OM-85 treated mothers demonstrate enhanced protection against lethal lower respiratory infection with mouse-adapted rhinovirus (vMC0), and associated lung immune changes. RESULTS: Offspring from mothers treated with OM-85 during pregnancy display accelerated postnatal seeding of lung myeloid populations demonstrating upregulation of function-associated markers. Offspring from OM-85 mothers additionally exhibit enhanced expression of TLR4/7 and the IL-1ß/NLRP3 inflammasome complex within the lung. These treatment effects were associated with enhanced capacity to clear an otherwise lethal respiratory viral infection during the neonatal period, with concomitant regulation of viral-induced IFN response intensity. CONCLUSION: These results demonstrate that maternal OM-85 treatment protects offspring against lethal neonatal respiratory viral infection by accelerating development of innate immune mechanisms crucial for maintenance of local immune homeostasis in the face of pathogen challenge.
RESUMO
The airway mucosal response to allergen in asthma involves influx of activated T helper type 2 cells and eosinophils, transient airflow obstruction, and airways hyperresponsiveness (AHR). The mechanism(s) underlying transient T cell activation during this inflammatory response is unclear. We present evidence that this response is regulated via bidirectional interactions between airway mucosal dendritic cells (AMDC) and T memory cells. After aerosol challenge, resident AMDC acquire antigen and rapidly mature into potent antigen-presenting cells (APCs) after cognate interactions with T memory cells. This process is restricted to dendritic cells (DCs) in the mucosae of the conducting airways, and is not seen in peripheral lung. Within 24 h, antigen-bearing mature DCs disappear from the airway wall, leaving in their wake activated interleukin 2R+ T cells and AHR. Antigen-bearing activated DCs appear in regional lymph nodes at 24 h, suggesting onward migration from the airway. Transient up-regulation of CD86 on AMDC accompanies this process, which can be reproduced by coculture of resting AMDC with T memory cells plus antigen. The APC activity of AMDC can be partially inhibited by anti-CD86, suggesting that CD86 may play an active role in this process and/or is a surrogate for other relevant costimulators. These findings provide a plausible model for local T cell activation at the lesional site in asthma, and for the transient nature of this inflammatory response.
Assuntos
Asma/imunologia , Comunicação Celular , Células Dendríticas/fisiologia , Pulmão/imunologia , Sistema Respiratório/imunologia , Linfócitos T/fisiologia , Animais , Antígenos CD/análise , Antígeno B7-2 , Antígenos de Histocompatibilidade Classe II/análise , Memória Imunológica , Ativação Linfocitária , Glicoproteínas de Membrana/análise , Mucosa/imunologia , Ovalbumina/imunologia , Ratos , Linfócitos T/imunologiaRESUMO
Inhaled allergens are known for their immediate and ongoing effects in the respiratory tract (RT). In this report, we track inhaled antigen in normal mice for 7 days and find that while it is cleared from the airways, inhaled antigen persists in peripheral lung tissue and the draining lymph nodes (DLNs). The persistence of antigen led to ongoing presentation in the lymph nodes, but not the lungs, that decreased with time in direct proportion with the frequency of antigen-bearing RT dendritic cells (DCs). There was evidence of functional changes among the antigen-bearing DCs in the lymph nodes, as the expression of CD40, CD80 and CD86 were modulated over the course of 7 days. At the same time, there was a decrease in both CD4(+) T-cell proliferation in lymph nodes and the generation of recirculating CD4(+) T cells. However, early presentation of lower doses of inhaled antigen also resulted in a decrease in CD4(+) T-cell proliferation and recirculation. Thus, T-cell recirculation depends on the strength of stimulus in the DLNs and is produced by a combination of the dose of antigen delivered to the RT, DC migration and co-stimulatory molecule expression. These results provide an important insight into the fate of inhaled antigen in vivo and the influence of persistent antigen presentation on T-cell activation in the lymph nodes.