RESUMO
Idiopathic pulmonary fibrosis (IPF) is an interstitial lung disease associated with chronic inflammation and tissue remodelling leading to fibrosis, reduced pulmonary function, respiratory failure and death. Bleomycin (Blm)-induced lung fibrosis in mice replicates several clinical features of human IPF, including prominent lymphoid aggregates of predominantly B-cells that accumulate in the lung adjacent to areas of active fibrosis. We have shown previously a requirement for B-cells in the development of Blm-induced lung fibrosis in mice. To determine the therapeutic potential of inhibiting B-cell function in pulmonary fibrosis, we examined the effects of anti-CD20 B-cell ablation therapy to selectively remove mature B-cells from the immune system and inhibit Blm-induced lung fibrosis. Anti-CD20 B-cell ablation did not reduce fibrosis in this model; however, immune phenotyping of peripheral blood and lung resident cells revealed that anti-CD20-treated mice retained a high frequency of CD19+ CD138+ plasma cells. Interestingly, high levels of CD138+ cells were also identified in the lung tissue of patients with IPF, consistent with the mouse model. Treatment of mice with bortezomib, which depletes plasma cells, reduced the level of Blm-induced lung fibrosis, implicating plasma cells as important effector cells in the development and progression of pulmonary fibrosis.
Assuntos
Fibrose Pulmonar Idiopática , Doenças Pulmonares Intersticiais , Humanos , Camundongos , Animais , Bleomicina/farmacologia , Plasmócitos , Fibrose Pulmonar Idiopática/tratamento farmacológico , Pulmão/metabolismo , Doenças Pulmonares Intersticiais/induzido quimicamenteRESUMO
BACKGROUND: Hundreds of genetic variants are thought to contribute to variation in asthma risk by modulating gene expression. Methods that increase the power of genome-wide association studies (GWASs) to identify risk-associated variants are needed. OBJECTIVE: We sought to develop a method that aggregates the evidence for association with disease risk across expression quantitative trait loci (eQTLs) of a gene and use this approach to identify asthma risk genes. METHODS: We developed a gene-based test and software package called EUGENE that (1) is applicable to GWAS summary statistics; (2) considers both cis- and trans-eQTLs; (3) incorporates eQTLs identified in different tissues; and (4) uses simulations to account for multiple testing. We applied this approach to 2 published asthma GWASs (combined n = 46,044) and used mouse studies to provide initial functional insights into 2 genes with novel genetic associations. RESULTS: We tested the association between asthma and 17,190 genes that were found to have cis- and/or trans-eQTLs across 16 published eQTL studies. At an empirical FDR of 5%, 48 genes were associated with asthma risk. Of these, for 37, the association was driven by eQTLs located in established risk loci for allergic disease, including 6 genes not previously implicated in disease cause (eg, LIMS1, TINF2, and SAFB). The remaining 11 significant genes represent potential novel genetic associations with asthma. The association with 4 of these replicated in an independent GWAS: B4GALT3, USMG5, P2RY13, and P2RY14, which are genes involved in nucleotide synthesis or nucleotide-dependent cell activation. In mouse studies, P2ry13 and P2ry14-purinergic receptors activated by adenosine 5-diphosphate and UDP-sugars, respectively-were upregulated after allergen challenge, notably in airway epithelial cells, eosinophils, and neutrophils. Intranasal exposure with receptor agonists induced the release of IL-33 and subsequent eosinophil infiltration into the lungs. CONCLUSION: We identified novel associations between asthma and eQTLs for 4 genes related to nucleotide synthesis/signaling and demonstrated the power of gene-based analyses of GWASs.
Assuntos
Asma/genética , Predisposição Genética para Doença/genética , Estudo de Associação Genômica Ampla/métodos , Nucleotídeos/genética , Software , Animais , Variação Genética/genética , Humanos , Camundongos , Camundongos Endogâmicos C57BL , ATPases Mitocondriais Próton-Translocadoras/genética , Nucleotídeos/biossíntese , Locos de Características Quantitativas/genética , Receptores Purinérgicos P2/genética , Receptores Purinérgicos P2Y/genéticaRESUMO
Despite widespread exposure to potentially pathogenic mycobacteria present in the soil and in domestic water supplies, it is not clear why only a small proportion of individuals contract pulmonary nontuberculous mycobacterial (NTM) infections. Here, we explore the impact of polymorphisms within three genes: P2X ligand gated ion channel 7 (P2X7R), P2X ligand gated ion channel 4 (P2X4R) and calcium/calmodulin-dependent protein kinase kinase 2 beta (CAMKK2) on susceptibility. Thirty single nucleotide polymorphisms (SNPs) were genotyped in NTM patients (n = 124) and healthy controls (n = 229). Weak associations were found between individual alleles in P2X7R and disease but were not significant in multivariate analyses adjusted to account for gender. Haplotypes spanning the three genes were derived using the fastPHASE algorithm. This yielded 27 haplotypes with frequencies >1% and accounting for 63.3% of the combined cohort. In univariate analyses, seven of these haplotypes displayed associations with NTM disease above our preliminary cut-off (p ≤ 0.20). When these were carried forward in a logistic regression model, gender and one haplotype (SH95) were independently associated with the disease (model p < 0.0001; R 2 = 0.05). Examination of individual alleles within these haplotypes implicated P2X7R and CAMKK2 in pathways affecting pulmonary NTM disease.
Assuntos
Quinase da Proteína Quinase Dependente de Cálcio-Calmodulina/genética , Pneumopatias/genética , Infecções por Mycobacterium não Tuberculosas/genética , Mycobacterium , Polimorfismo de Nucleotídeo Único , Receptores Purinérgicos P2X4/genética , Receptores Purinérgicos P2X7/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Casos e Controles , Feminino , Predisposição Genética para Doença , Haplótipos , Humanos , Pneumopatias/epidemiologia , Pneumopatias/microbiologia , Masculino , Pessoa de Meia-Idade , Mycobacterium/patogenicidade , Infecções por Mycobacterium não Tuberculosas/epidemiologia , Infecções por Mycobacterium não Tuberculosas/microbiologiaRESUMO
Complex lung diseases, such as asthma, are influenced by both genetic predisposition and environmental stimuli. The epigenetic landscape of such diseases is attracting increasing interest and research. Epigenetics broadly covers the transient and the inheritable changes to gene expression that are not directly due to changes in nucleotide sequences. Epigenetic mechanisms could have significant impact on asthma-related allergic, immune, and regulatory pathways, as well as on the generation of biomarkers and the heritable transmission of asthma phenotypes. Recent technological advances have allowed mapping of the epigenome and analysis of genome-wide epigenetic contributors to disease. As a result, ground-breaking observations regarding histone post-translational modifications in a number of immunological diseases have emerged. In this review, we look beyond the biological information coded by DNA and review the epigenetic modifications made to histones, with evidence suggesting a role for their modification in asthma.
Assuntos
Asma/genética , Montagem e Desmontagem da Cromatina , Epigênese Genética , Histonas/metabolismo , Animais , Asma/imunologia , Asma/metabolismo , Asma/fisiopatologia , Regulação da Expressão Gênica , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Pulmão/imunologia , Pulmão/fisiopatologia , Fenótipo , Prognóstico , Fatores de RiscoRESUMO
BACKGROUND: To date, no genome-wide association study (GWAS) has considered the combined phenotype of asthma with hay fever. Previous analyses of family data from the Tasmanian Longitudinal Health Study provide evidence that this phenotype has a stronger genetic cause than asthma without hay fever. OBJECTIVE: We sought to perform a GWAS of asthma with hay fever to identify variants associated with having both diseases. METHODS: We performed a meta-analysis of GWASs comparing persons with both physician-diagnosed asthma and hay fever (n = 6,685) with persons with neither disease (n = 14,091). RESULTS: At genome-wide significance, we identified 11 independent variants associated with the risk of having asthma with hay fever, including 2 associations reaching this level of significance with allergic disease for the first time: ZBTB10 (rs7009110; odds ratio [OR], 1.14; P = 4 × 10(-9)) and CLEC16A (rs62026376; OR, 1.17; P = 1 × 10(-8)). The rs62026376:C allele associated with increased asthma with hay fever risk has been found to be associated also with decreased expression of the nearby DEXI gene in monocytes. The 11 variants were associated with the risk of asthma and hay fever separately, but the estimated associations with the individual phenotypes were weaker than with the combined asthma with hay fever phenotype. A variant near LRRC32 was a stronger risk factor for hay fever than for asthma, whereas the reverse was observed for variants in/near GSDMA and TSLP. Single nucleotide polymorphisms with suggestive evidence for association with asthma with hay fever risk included rs41295115 near IL2RA (OR, 1.28; P = 5 × 10(-7)) and rs76043829 in TNS1 (OR, 1.23; P = 2 × 10(-6)). CONCLUSION: By focusing on the combined phenotype of asthma with hay fever, variants associated with the risk of allergic disease can be identified with greater efficiency.
Assuntos
Asma/diagnóstico , Asma/genética , Variação Genética , Estudo de Associação Genômica Ampla , Fenótipo , Locos de Características Quantitativas , Rinite Alérgica Sazonal/diagnóstico , Rinite Alérgica Sazonal/genética , Adulto , Alelos , Asma/complicações , Feminino , Frequência do Gene , Humanos , Lectinas Tipo C/genética , Desequilíbrio de Ligação , Masculino , Pessoa de Meia-Idade , Proteínas de Transporte de Monossacarídeos/genética , Razão de Chances , Polimorfismo de Nucleotídeo Único , Proteínas Repressoras/genética , Rinite Alérgica Sazonal/complicações , Adulto JovemAssuntos
Proteína Antagonista do Receptor de Interleucina 1/uso terapêutico , Síndrome de Schnitzler/complicações , Urticária/tratamento farmacológico , Urticária/etiologia , Biópsia por Agulha , Doença Crônica , Esquema de Medicação , Feminino , Humanos , Imuno-Histoquímica , Injeções Subcutâneas , Pessoa de Meia-Idade , Prognóstico , Doenças Raras , Síndrome de Schnitzler/diagnóstico , Índice de Gravidade de Doença , Resultado do Tratamento , Urticária/patologiaRESUMO
BACKGROUND: We aimed to identify novel genetic variants affecting asthma risk, since these might provide novel insights into molecular mechanisms underlying the disease. METHODS: We did a genome-wide association study (GWAS) in 2669 physician-diagnosed asthmatics and 4528 controls from Australia. Seven loci were prioritised for replication after combining our results with those from the GABRIEL consortium (n=26,475), and these were tested in an additional 25,358 independent samples from four in-silico cohorts. Quantitative multi-marker scores of genetic load were constructed on the basis of results from the GABRIEL study and tested for association with asthma in our Australian GWAS dataset. FINDINGS: Two loci were confirmed to associate with asthma risk in the replication cohorts and reached genome-wide significance in the combined analysis of all available studies (n=57,800): rs4129267 (OR 1·09, combined p=2·4×10(-8)) in the interleukin-6 receptor (IL6R) gene and rs7130588 (OR 1·09, p=1·8×10(-8)) on chromosome 11q13.5 near the leucine-rich repeat containing 32 gene (LRRC32, also known as GARP). The 11q13.5 locus was significantly associated with atopic status among asthmatics (OR 1·33, p=7×10(-4)), suggesting that it is a risk factor for allergic but not non-allergic asthma. Multi-marker association results are consistent with a highly polygenic contribution to asthma risk, including loci with weak effects that might be shared with other immune-related diseases, such as NDFIP1, HLA-B, LPP, and BACH2. INTERPRETATION: The IL6R association further supports the hypothesis that cytokine signalling dysregulation affects asthma risk, and raises the possibility that an IL6R antagonist (tocilizumab) may be effective to treat the disease, perhaps in a genotype-dependent manner. Results for the 11q13.5 locus suggest that it directly increases the risk of allergic sensitisation which, in turn, increases the risk of subsequent development of asthma. Larger or more functionally focused studies are needed to characterise the many loci with modest effects that remain to be identified for asthma. FUNDING: National Health and Medical Research Council of Australia. A full list of funding sources is provided in the webappendix.
Assuntos
Asma/genética , Cromossomos Humanos Par 11/genética , Loci Gênicos/genética , Proteínas de Membrana/genética , Polimorfismo de Nucleotídeo Único , Receptores de Interleucina-6/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Asma/imunologia , Criança , Pré-Escolar , Feminino , Predisposição Genética para Doença/genética , Estudo de Associação Genômica Ampla , Humanos , Hipersensibilidade Imediata/genética , Desequilíbrio de Ligação , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
This study tested the hypothesis that proinflammatory kinin peptides are involved in modulating human dendritic cell (DC) function. Inflammation is accompanied by an increased maturation of DCs and the generation of kinins, particularly Lys-des[Arg(9)]-bradykinin (Lda-BK). We assessed the role of Lda-BK in the activation and migration of human monocyte-derived DCs (hMo-DCs) matured through the use of LPS, TNF-α + IL-1ß, or CD40 ligand. Kinin B(1) and B(2) receptor mRNA and protein expression were assessed by confocal microscopy, flow cytometry, and RT-PCR. The effects of Lda-BK on the migration of mature hMo-DCs were assessed in Transwell chambers, whereas the expression of costimulatory molecules and the secretion of IL-12 were assessed by flow cytometry and ELISA, respectively. The expression of the kinin B(1) receptor (B(1)R) was down-regulated during the maturation of hMo-DCs, whereas the expression of B(2)R was unchanged. The B(1)R agonist Lda-BK was not chemotactic for hMo-DCs matured using LPS, TNF-α + IL-1ß, or CD40 ligand, but Lda-BK enhanced the secretion of IL-12p70 and inhibited the secretion of IL-12p40 by mature hMo-DCs. However, the exposure of hMo-DCs matured with TNF-α + IL-1ß to Lda-BK for 6 hours decreased subsequent migration in response to Lda-BK, the chemokine CCL19, or Lda-BK combined with CCL19. The expression of B(1)R was increased in hMo-DCs from subjects with asthma compared with subjects without asthma, in keeping with a tendency toward increased in vitro migration of asthmatic hMo-DCs in response to Lda-BK. The increased formation of Lda-BK and the enhanced expression of B(1)R as a consequence of inflammation may alter the migration of mature, antigen-laden DCs to regional lymph nodes in response to CCL19, may modulate the secretion of cytokines by these DCs, and may contribute to the accumulation of mature DCs in the lungs of patients with asthma.
Assuntos
Bradicinina/farmacologia , Células Dendríticas/citologia , Interleucina-12/metabolismo , Calidina/farmacologia , Monócitos/citologia , Adulto , Asma/metabolismo , Ligante de CD40/química , Estudos de Casos e Controles , Movimento Celular , Fatores Quimiotáticos/metabolismo , Citocinas/metabolismo , Humanos , Ligantes , RNA Mensageiro/metabolismo , Receptor B1 da Bradicinina/metabolismo , Receptores CCR7/metabolismoRESUMO
The kinins, bradykinin (BK) and Lys-des[Arg(9)]-BK, are important inflammatory mediators that act via two specific G protein-coupled kinins, B(1) and B(2) receptors (B(2)R). Kinins influence the activity of immune cells by stimulating the synthesis of cytokines, eicosanoids, and chemotactic factors. Whether human dendritic cells (DC) express kinin receptors and whether kinins influence DC function are unknown. Fluorescence immunocytochemistry and RT-PCR were used to demonstrate that immature human monocyte-derived DC (hMo-DC) constitutively expressed kinins B(1)R and B(2)R. Kinin receptor expression was induced on the 3rd and 4th days of culture during differentiation of hMo-DC from monocytes and was not dependent on the presence of IL-4 or GM-CSF. Although monocytes also expressed B(2)R mRNA, the protein was not detected. The kinin agonists BK and Lys-des[Arg(9)]-BK up-regulated the expression of their respective receptors. BK, acting via the B(2)R, increased intracellular Ca(2+), as visualized by confocal microscopy using the fluorescent Ca(2+) dye, Fluor-4 AM. Evaluation of migration in Trans-well chambers demonstrated significant enhancement by BK of migration of immature hMo-DC, which was B(2)R-dependent. However, kinins did not induce maturation of hMo-DC. The novel finding that kinin receptors are constitutively expressed in immature hMo-DC suggests that these receptors may be expressed in the absence of proinflammatory stimuli. BK, which increases the migration of immature hMo-DC in vitro, may play an important role in the migration of immature DC in noninflammatory conditions and may also be involved in the recruitment of immature DC to sites of inflammation.
Assuntos
Bradicinina/farmacologia , Cálcio/metabolismo , Células Dendríticas/metabolismo , Monócitos/metabolismo , Receptor B1 da Bradicinina/biossíntese , Receptor B2 da Bradicinina/biossíntese , Diferenciação Celular , Movimento Celular , Células Cultivadas , Células Dendríticas/citologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Humanos , Interleucina-4/farmacologia , Líquido Intracelular/metabolismo , Calidina/análogos & derivados , Calidina/farmacologia , Monócitos/citologia , Receptor B1 da Bradicinina/agonistas , Receptor B2 da Bradicinina/agonistasRESUMO
Polymorphisms in P2X4R and CAMKK2 associate with susceptibility to HIV-associated sensory neuropathy (HIV-SN) - a condition likely mediated by TNFα. As single nucleotide polymorphisms (SNPs) and haplotypes of CAMKK2, and a neighbouring gene P2X4R, mark susceptibility to HIV-SN in South Africans living with HIV, we examined the relationship between P2X4R and CAMKK2 genotypes and TNFα production. Peripheral blood mononuclear cells from 129 healthy donors were stimulated with killed Escherichia coli, and concentrations of soluble TNFα were assessed. Their DNA was genotyped for 22 SNPs in P2X4R and CAMKK2. Three SNPs within P2X4R and two SNPs within CAMKK2 influenced concentrations of TNFα, but these SNP did not associate with risk for HIV-SN. This incongruence may reflect differences in P2X4R haplotypes present in Africans and Europeans. However some CAMKK2 haplotypes were found in both populations, so CAMKK2 polymorphisms may impact upon HIV-SN via effects of the protein on pathways other than TNFα.
Assuntos
Quinase da Proteína Quinase Dependente de Cálcio-Calmodulina/genética , Infecções por HIV/complicações , Doenças do Sistema Nervoso Periférico/genética , Receptores Purinérgicos P2X4/genética , Fator de Necrose Tumoral alfa/metabolismo , Adulto , População Negra/genética , Doadores de Sangue , Escherichia coli/imunologia , Infecções por HIV/imunologia , Haplótipos , Voluntários Saudáveis , Humanos , Leucócitos Mononucleares/patologia , Doenças do Sistema Nervoso Periférico/sangue , Doenças do Sistema Nervoso Periférico/etiologia , Polimorfismo de Nucleotídeo Único , Fator de Necrose Tumoral alfa/imunologia , População Branca/genéticaRESUMO
BACKGROUND: Several animal studies, and one inoculation study in adult asthmatics have shown that interleukin-33 (IL-33) is a major contributor to type-2 inflammation in acute asthma. However, the link between IL-33 and type-2 inflammation has not been shown in naturally occurring asthma exacerbations. OBJECTIVES: To determine if airway IL-33 is associated with type-2 inflammation measured by type-2 cytokines, FeNO and sputum eosinophils in patients presenting to the Emergency Department with an asthma exacerbations. METHODS: Adult patients hospitalized due to acute asthma were enrolled. Upper airways were sampled with nasal swabs and lower airways with induced sputum. Cytokines were measured at protein level using a Luminex® assay and mRNA expression level using droplet-digital-PCR. Airway sampling was repeated four weeks after exacerbation. RESULTS: At the time of exacerbation, upper airway IL-33 correlated with upper airway IL-5 and IL-13 (Râ¯=â¯0.84, pâ¯<â¯0.01 and Râ¯=â¯0.76, pâ¯<â¯0.01, respectively) and with lower airway IL-13 (Râ¯=â¯0.49, pâ¯=â¯0.03). Similar associations were observed for mRNA expression. Lower airway IL-33 positively correlated with lower airway IL-13 (Râ¯=â¯0.84, pâ¯<â¯0.01). IL-13 and IL-33 were positively correlated with FeNO, and IL-5 with eosinophils. The association between IL-33 and type-2 cytokines were still present four weeks after exacerbation. CONCLUSION: This is the first study to demonstrate that airway IL-33 is associated with type-2 cytokines in naturally occurring asthma exacerbations in adults, providing in vivo evidence supporting that IL-33 may be driving type-2 inflammation in acute asthma. Thus supporting IL-33 as a potential future drug target due to its role, upstream in the immunological cascade.
Assuntos
Asma/imunologia , Citocinas/metabolismo , Mediadores da Inflamação/metabolismo , Doença Aguda , Adulto , Citocinas/genética , Eosinófilos/imunologia , Feminino , Seguimentos , Expressão Gênica/imunologia , Humanos , Interleucina-13/genética , Interleucina-13/metabolismo , Interleucina-33/genética , Interleucina-33/metabolismo , Interleucina-5/genética , Interleucina-5/metabolismo , Masculino , Pessoa de Meia-Idade , Mucosa Nasal/imunologia , RNA Mensageiro/genética , Índice de Gravidade de Doença , Escarro/imunologia , Adulto JovemRESUMO
Reporter assays are widely used in applications that require measurement of changes in gene expression over time (e.g. drug screening). With standard reporter vectors, the measurable effect of a treatment or compound (altered reporter activity) is substantially diluted and delayed, compared with its true effect (altered transcriptional activity). This problem is caused by the relatively long half-lives of both the reporter protein and its mRNA. As a result, the activities of compounds, ligands or treatments that have a relatively minor effect, or a substantial but transient effect, often remain undetected. To circumvent this problem, we introduced modular protein- and mRNA-destabilizing elements into a range of commonly used reporters. Our data show that both elements are required for maximal responses to both increases and decreases in transcriptional activity. The double-destabilized reporter vectors showed markedly improved performance in drug screening, kinetic assays and dose-response titrations.
Assuntos
Motivos de Aminoácidos , Genes Reporter , Sequências Reguladoras de Ácido Ribonucleico , Transcrição Gênica , Animais , Células CHO , Cricetinae , Cricetulus , Avaliação Pré-Clínica de Medicamentos , Vetores Genéticos , Meia-Vida , Células HeLa , Humanos , Proteínas/metabolismo , Estabilidade de RNA , RNA Mensageiro/metabolismo , Proteínas Recombinantes de Fusão/análise , Proteínas Recombinantes de Fusão/genética , Terminologia como Assunto , TransfecçãoRESUMO
Although exposure to potentially pathogenic nontuberculous mycobacteria (NTM) via soil and domestic water supplies is common, pulmonary infection and disease are confined to a small proportion of older individuals. Previously, alleles of a polymorphism in IL10 (rs1800896) were associated with NTM disease and we demonstrated elevated production of IL-10 by blood leukocytes from patients with pulmonary NTM. Here seven additional polymorphisms in IL10 were investigated in a larger cohort of Caucasian controls and patients with pulmonary NTM disease. This demonstrated a significant association between pulmonary NTM disease and one polymorphism (rs1518111) in strong linkage disequilibrium with rs1800896.
Assuntos
Interleucina-10/genética , Infecções por Mycobacterium não Tuberculosas/genética , Micobactérias não Tuberculosas/fisiologia , Pneumonia/genética , População Branca , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Feminino , Frequência do Gene , Predisposição Genética para Doença , Genótipo , Humanos , Desequilíbrio de Ligação , Masculino , Pessoa de Meia-Idade , Pneumonia/microbiologia , Polimorfismo de Nucleotídeo ÚnicoRESUMO
BACKGROUND: In experimental studies viral infections have been shown to induce type 2 inflammation in asthmatics, but whether this is a feature of naturally occurring virus-induced asthma exacerbations is unknown. Thymic stromal lymphopoietin (TSLP) released from the airway epithelium in response to damage, has been suggested as a link between viral infection and type 2 inflammation, but the role of TSLP in asthma exacerbations is unknown. OBJECTIVE: To assess whether type 2 inflammation, as measured by sputum eosinophils and fractional exhaled nitric oxide (FeNO), is a feature of naturally occurring virus-induced exacerbations of asthma and whether TSLP is associated with this type 2 inflammation. METHODS: Patients presenting to hospital with acute asthma were examined during the exacerbation, and after 4 weeks recovery. The assessments included spirometry, FeNO and induced sputum for differential counts and TSLP mRNA levels. Nasal swabs were collected for viral detection. RESULTS: Sputum eosinophils and FeNO were similar between virus-positive (n = 44) and negative patients (n = 44). In virus-positive patients, TSLP expression was lower at exacerbation than follow-up (p = 0.03). High TSLP at exacerbation was associated with lower sputum eosinophils (p = 0.01) and higher FEV1 (p = 0.03). In virus-positive patients, %-predicted FEV1 negatively correlated with both FeNO and sputum eosinophils (p = 0.02 and p = 0.05, respectively). CONCLUSION: Our findings support that type 2 inflammation is present in patients during virus-induced asthma exacerbations, to the same degree as non-viral exacerbations, and correlate negatively with FEV1. However, in virus-positive patients, high TSLP expression during exacerbation was associated with low sputum eosinophils, suggesting that the effect of TSLP in vivo, in the setting of an asthma exacerbation, might be different than the type 2 inducing effects observed in experimental studies.
Assuntos
Asma/virologia , Infecções Respiratórias/complicações , Viroses/complicações , Doença Aguda , Adulto , Asma/metabolismo , Asma/fisiopatologia , Citocinas/biossíntese , Citocinas/genética , Eosinofilia/virologia , Feminino , Volume Expiratório Forçado/fisiologia , Expressão Gênica/fisiologia , Humanos , Inflamação/metabolismo , Inflamação/fisiopatologia , Inflamação/virologia , Masculino , Pessoa de Meia-Idade , Óxido Nítrico/metabolismo , Fenótipo , Estudos Prospectivos , RNA Mensageiro/genética , Infecções Respiratórias/metabolismo , Infecções Respiratórias/fisiopatologia , Índice de Gravidade de Doença , Espirometria , Escarro/citologia , Escarro/metabolismo , Viroses/metabolismo , Viroses/fisiopatologia , Adulto Jovem , Linfopoietina do Estroma do TimoRESUMO
Cystic Fibrosis (CF) is often accompanied by diabetes leading to worsening lung function, the reason for which is unclear. The receptor for advanced-glycation-end-products (RAGE) regulates immune responses and inflammation and has been linked to diabetes and possibly CF. We performed a pilot study to determine if CF and CF-related diabetes (CFRD) are associated with enhanced RAGE expression. Full length (fl)RAGE, soluble (s)RAGE, endogenous soluble (es)RAGE, S100A12 (enRAGE) and advanced-glycation-end-products (AGE) expression was assessed in serum, white blood cells and sputum of patients with CF; diabetes; CFRD and healthy subjects. Sputum enRAGE/sRAGE ratios were high in CF but particularly in CFRD which negatively correlated with % predicted FEV1. Serum AGE and AGE/sRAGE ratios were high in diabetics but not in CF. A complex, multifaceted approach was used to assess the role of RAGE and its ligands which is fundamental to determining their impact on airway inflammation. There is a clear association between RAGE activity in the airways of CF and CFRD patients that is not evident in the vascular compartment and correlates with lung function, in contrast to diabetes. This strongly suggests a role for RAGE in contributing to the inflammatory overdrive seen in CF and to a greater extent in CFRD.
Assuntos
Fibrose Cística/genética , Diabetes Mellitus/genética , Inflamação/genética , Receptor para Produtos Finais de Glicação Avançada/biossíntese , Adulto , Fibrose Cística/sangue , Fibrose Cística/complicações , Fibrose Cística/patologia , Diabetes Mellitus/sangue , Diabetes Mellitus/etiologia , Diabetes Mellitus/patologia , Feminino , Regulação da Expressão Gênica , Produtos Finais de Glicação Avançada/sangue , Produtos Finais de Glicação Avançada/genética , Humanos , Imunidade Inata/genética , Inflamação/sangue , Inflamação/patologia , Masculino , Pessoa de Meia-Idade , Receptor para Produtos Finais de Glicação Avançada/sangue , Escarro/metabolismoRESUMO
Gli transcription factors of the Hedgehog (Hh) pathway have been reported to be drivers of malignant mesothelioma (MMe) cell survival. The Gli inhibitor GANT61 induces apoptosis in various cancer cell models, and has been associated directly with Gli inhibition. However various chemotherapeutics can induce cell death through generation of reactive oxygen species (ROS) but whether ROS mediates GANT61-induced apoptosis is unknown. In this study human MMe cells were treated with GANT61 and the mechanisms regulating cell death investigated. Exposure of MMe cells to GANT61 led to G1 phase arrest and apoptosis, which involved ROS but not its purported targets, GLI1 or GLI2. GANT61 triggered ROS generation and quenching of ROS protected MMe cells from GANT61-induced apoptosis. Furthermore, we demonstrated that mitochondria are important in mediating GANT61 effects: (1) ROS production and apoptosis were blocked by mitochondrial inhibitor rotenone; (2) GANT61 promoted superoxide formation in mitochondria; and (3) mitochondrial DNA-deficient LO68 cells failed to induce superoxide, and were more resistant to apoptosis induced by GANT61 than wild-type cells. Our data demonstrate for the first time that GANT61 induces apoptosis by promoting mitochondrial superoxide generation independent of Gli inhibition, and highlights the therapeutic potential of mitochondrial ROS-mediated anticancer drugs in MMe.
Assuntos
Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Mesotelioma/tratamento farmacológico , Mesotelioma/metabolismo , Mitocôndrias/metabolismo , Piridinas/farmacologia , Pirimidinas/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Fase G1/efeitos dos fármacos , Células HCT116 , Células HT29 , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Mesotelioma/genética , Mesotelioma/patologia , Mesotelioma Maligno , Mitocôndrias/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Fatores de Transcrição/metabolismo , Proteína GLI1 em Dedos de ZincoRESUMO
Eczema often precedes the development of asthma in a disease course called the 'atopic march'. To unravel the genes underlying this characteristic pattern of allergic disease, we conduct a multi-stage genome-wide association study on infantile eczema followed by childhood asthma in 12 populations including 2,428 cases and 17,034 controls. Here we report two novel loci specific for the combined eczema plus asthma phenotype, which are associated with allergic disease for the first time; rs9357733 located in EFHC1 on chromosome 6p12.3 (OR 1.27; P=2.1 × 10(-8)) and rs993226 between TMTC2 and SLC6A15 on chromosome 12q21.3 (OR 1.58; P=5.3 × 10(-9)). Additional susceptibility loci identified at genome-wide significance are FLG (1q21.3), IL4/KIF3A (5q31.1), AP5B1/OVOL1 (11q13.1), C11orf30/LRRC32 (11q13.5) and IKZF3 (17q21). We show that predominantly eczema loci increase the risk for the atopic march. Our findings suggest that eczema may play an important role in the development of asthma after eczema.
Assuntos
Asma/genética , Dermatite Atópica/genética , Proteínas Adaptadoras de Transporte Vesicular/genética , Adolescente , Adulto , Sistemas de Transporte de Aminoácidos Neutros/genética , Proteínas de Ligação ao Cálcio/genética , Proteínas de Transporte/genética , Criança , Pré-Escolar , Proteínas de Ligação a DNA/genética , Progressão da Doença , Feminino , Proteínas Filagrinas , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Fator de Transcrição Ikaros/genética , Interleucina-4/genética , Cinesinas/genética , Modelos Logísticos , Masculino , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Proteínas de Neoplasias/genética , Proteínas do Tecido Nervoso/genética , Proteínas Nucleares/genética , Fenótipo , Polimorfismo de Nucleotídeo Único , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Proteínas Repressoras/genética , Fatores de Transcrição/genética , Adulto JovemRESUMO
The region spanning the tumor necrosis factor (TNF) cluster in the human major histocompatibility complex (MHC) has been implicated in susceptibility to numerous immunopathological diseases, including type 1 diabetes mellitus and rheumatoid arthritis. However, strong linkage disequilibrium across the MHC has hampered the identification of the precise genes involved. In addition, the observation of "blocks" of DNA in the MHC within which recombination is very rare, limits the resolution that may be obtained by genotyping individual SNPs. Hence a greater understanding of the haplotypes of the block spanning the TNF cluster is necessary. To this end, we genotyped 32 human leukocyte antigen (HLA)-homozygous workshop cell lines and 300 healthy control samples for 19 coding and promoter region SNPs spanning 45 kb in the central MHC near the TNF genes. The workshop cell lines defined 11 SNP haplotypes that account for approximately 80% of the haplotypes observed in the 300 control individuals. Using the control individuals, we defined a further six haplotypes that account for an additional 10% of donors. We show that the 17 haplotypes of the "TNF block" can be identified using 15 SNPs.
Assuntos
Complexo Principal de Histocompatibilidade , Polimorfismo de Nucleotídeo Único , Fatores de Necrose Tumoral/genética , População Branca/genética , Linhagem Celular Transformada , RNA Helicases DEAD-box , Genótipo , Haplótipos , Humanos , Regiões Promotoras Genéticas , RNA Helicases/genéticaRESUMO
BACKGROUND: Novel vascular-independent conduits have been observed in some cancers. These have been variously described as vasculogenic mimicry, mosaic vessel formation, vascular co-option and intratumour embryonic-like vasculogenesis. Despite lung cancer being the most common cancer worldwide, there is little information on its neovascularisation or the pathways involved. METHODS: An in vitro model involving co-cultures of microvascular lung endothelial cells and squamous or adenocarcinoma lung cancer cells was developed to assess their angiogenic interaction. Cells were incubated and examined by phase contrast microscopy and by immunocytochemistry in both mono- and co-cultures. Cultured cells and lung cancer tissue sections were assessed for new tumour vessel formation, expression of the endothelial marker CD31 and morphology. RESULTS: Lung tumour cells and endothelial cells interacted morphologically via pseudopodia and used alternative pathways to generate new vessels. Co-culturing microvascular endothelial and squamous carcinoma cells led to endothelial cells surrounding tumour cells and the tumour cells being incorporated into vessel walls. Co-culturing endothelial and adenocarcinoma cells resulted in cellular contact and the formation of tumour cell bridges around clusters of endothelial cells. These adencocarcinoma cells became strongly positive for CD31. Tumour tissue section studies supported the in vitro findings. CONCLUSION: Lung carcinoma cells when co-cultured with lung endothelial cells modify their cellular and molecular features that encourage alternative means of providing blood supply. The mechanisms underpinning these non-angiogenic processes need to be further investigated and should be considered when anti-tumour therapeutic interventions are being considered.