RESUMO
IL-9-secreting Th9 cells have been considered to play a pivotal role in the pathogenesis of atopic diseases. To what extent IL-9-producing cells are induced or regulated by sensitization with naturally occurring allergens is not yet clear. Naturally occurring allergens are capable of inducing IL-6 production in dendritic cells (DCs). Whether allergen-induced IL-6 supports a Th9 subtype by increasing IL-9 production, as observed in in vitro studies, or rather favors Th17 differentiation is not finally resolved. Therefore, in the present study we have investigated the impact of IL-6 on the Th9/Th17 balance depending on the predominant cytokine milieu and, additionally, in vivo using a DC-driven murine asthma model. In vitro, IL-6 increases Th9 cells under strong IL-4 and TGF-ß activation, whereas under moderate IL-4 and TGF-ß activation the presence of IL-6 shifts naive CD4(+) cells to Th17 cells. To induce allergic airway inflammation, OVA-pulsed DCs from IL-6-deficient or wild-type donors were adoptively transferred into BALB/c mice. Recipients receiving IL-6-producing wild-type DCs showed a significant decrease of Th9- and IL-4-producing Th2 cells but an increase of Th17 cells in lung tissue in comparison with recipients sensitized with IL-6-deficient DCs. Our data suggest that the IL-6-mediated reduction of Th2-related IL-4 leads to a decline of the Th9 immune response and allows Th17 differentiation.
Assuntos
Alérgenos/imunologia , Asma/imunologia , Linfócitos T CD4-Positivos/imunologia , Inflamação/imunologia , Interleucina-17/imunologia , Interleucina-6/imunologia , Interleucina-9/imunologia , Animais , Modelos Animais de Doenças , Feminino , Interleucina-6/biossíntese , Camundongos , Camundongos Endogâmicos BALB CRESUMO
In medicine, patent blue violet (PBV) is utilized for staining lymphatic vessels in sentinel lymph node (SLN) surgery. Moreover, PBV (also called E131 ) is used as food additive. We report on a 51-year-old non-atopic female with early breast cancer, who was scheduled for SLN excision and experienced an intra-operative anaphylactic reaction. In diagnostics the skin prick test (SPT) was positive to PBV. Hypersensitivity reactions to PBV can arise after the first exposure in surgery as sensitization may arise from either PBV (E131) in foods (i.e. in sweets or blue curacao) or from other structurally closely related triarylmethane dyes in objects of everyday life like textiles, detergents, paints, cold remedies and cosmetics. This article supports the necessity of an increased awareness of the possibility of anaphylactic reactions to PBV during SLN surgery, even if the patient never had contact to PBV before.
Assuntos
Anafilaxia/induzido quimicamente , Neoplasias da Mama/cirurgia , Corantes/efeitos adversos , Complicações Intraoperatórias/induzido quimicamente , Corantes de Rosanilina/efeitos adversos , Biópsia de Linfonodo Sentinela , Neoplasias da Mama/patologia , Feminino , Humanos , Pessoa de Meia-Idade , Testes CutâneosRESUMO
Vascular endothelial (VE)-protein tyrosine phosphatase (PTP) associates with VE-cadherin, thereby supporting its adhesive activity and endothelial junction integrity. VE-PTP also associates with Tie-2, dampening the tyrosine kinase activity of this receptor that can support stabilization of endothelial junctions. Here, we have analyzed how interference with VE-PTP affects the stability of endothelial junctions in vivo. Blocking VE-PTP by antibodies, a specific pharmacological inhibitor (AKB-9778), and gene ablation counteracted vascular leak induction by inflammatory mediators. In addition, leukocyte transmigration through the endothelial barrier was attenuated. Interference with Tie-2 expression in vivo reversed junction-stabilizing effects of AKB-9778 into junction-destabilizing effects. Furthermore, lack of Tie-2 was sufficient to weaken the vessel barrier. Mechanistically, inhibition of VE-PTP stabilized endothelial junctions via Tie-2, which triggered activation of Rap1, which then caused the dissolution of radial stress fibers via Rac1 and suppression of nonmuscle myosin II. Remarkably, VE-cadherin gene ablation did not abolish the junction-stabilizing effect of the VE-PTP inhibitor. Collectively, we conclude that inhibition of VE-PTP stabilizes challenged endothelial junctions in vivo via Tie-2 by a VE-cadherin-independent mechanism. In the absence of Tie-2, however, VE-PTP inhibition destabilizes endothelial barrier integrity in agreement with the VE-cadherin-supportive effect of VE-PTP.
Assuntos
Caderinas/deficiência , Células Endoteliais/metabolismo , Receptor TIE-2/metabolismo , Proteínas Tirosina Fosfatases Classe 3 Semelhantes a Receptores/metabolismo , Compostos de Anilina/farmacologia , Animais , Antígenos CD/metabolismo , Caderinas/metabolismo , Permeabilidade Capilar/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/ultraestrutura , Deleção de Genes , Inativação Gênica/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/citologia , Neutrófilos/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Fosfotirosina/metabolismo , RNA Interferente Pequeno/metabolismo , Proteínas Recombinantes de Fusão/farmacologia , Ácidos Sulfônicos/farmacologia , Proteínas rap1 de Ligação ao GTP/metabolismoRESUMO
Since people in industrialized countries spend most of their time indoors, the effects of indoor contaminants such as volatile organic compounds become more and more relevant. Benzene and toluene are among the most abundant compounds in the highly heterogeneous group of indoor volatile organic compounds. In order to understand their effects on lung epithelial cells (A549) representing lung's first line of defense, we chose a global proteome and a targeted metabolome approach in order to detect adverse outcome pathways caused by exposure to benzene and toluene. Using a DIGE approach, 93 of 469 detected protein spots were found to be differentially expressed after exposure to benzene, and 79 of these spots were identified by MS. Pathway analysis revealed an enrichment of proteins involved in Nrf2-mediated and oxidative stress response glycolysis/gluconeogenesis. The occurrence of oxidative stress at nonacute toxic concentrations of benzene and toluene was confirmed by the upregulation of the stress related proteins NQO1 and SOD1. The changes in metabolism were validated by ion chromatography MS/MS analysis revealing significant changes of glucose-6-phosphate, fructose-6-phosphate, 3-phosphoglycerate, and NADPH. The molecular alterations identified as a result of benzene and toluene exposure demonstrate the detrimental effect of nonacute toxic concentrations on lung epithelial cells. The data provided here will allow for a targeted validation in in vivo models.
Assuntos
Benzeno/toxicidade , Células Epiteliais/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Tolueno/toxicidade , Carbono/metabolismo , Linhagem Celular , Análise por Conglomerados , Eletroforese em Gel Bidimensional , Células Epiteliais/metabolismo , Humanos , Pulmão/citologia , Pulmão/metabolismo , Proteoma/análise , Proteoma/química , Proteoma/efeitos dos fármacos , Proteoma/metabolismo , Proteômica , Mucosa Respiratória/citologia , Transdução de Sinais/efeitos dos fármacos , Testes de Toxicidade SubagudaRESUMO
Obesity is associated with multiple adverse health effects and a high risk of developing metabolic and cardiovascular diseases. Therefore, there is a great need to identify circulating parameters that link changes in body fat mass with obesity. This study combines proteomic and metabolomic approaches to identify circulating molecules that discriminate healthy lean from healthy obese individuals in an exploratory study design. To correct for variations in physical activity, study participants performed a one hour exercise bout to exhaustion. Subsequently, circulating factors differing between lean and obese individuals, independent of physical activity, were identified. The DIGE approach yielded 126 differentially abundant spots representing 39 unique proteins. Differential abundance of proteins was confirmed by ELISA for antithrombin-III, clusterin, complement C3 and complement C3b, pigment epithelium-derived factor (PEDF), retinol binding protein 4 (RBP4), serum amyloid P (SAP), and vitamin-D binding protein (VDBP). Targeted serum metabolomics of 163 metabolites identified 12 metabolites significantly related to obesity. Among those, glycine (GLY), glutamine (GLN), and glycero-phosphatidylcholine 42:0 (PCaa 42:0) serum concentrations were higher, whereas PCaa 32:0, PCaa 32:1, and PCaa 40:5 were decreased in obese compared to lean individuals. The integrated bioinformatic evaluation of proteome and metabolome data yielded an improved group separation score of 2.65 in contrast to 2.02 and 2.16 for the single-type use of proteomic or metabolomics data, respectively. The identified circulating parameters were further investigated in an extended set of 30 volunteers and in the context of two intervention studies. Those included 14 obese patients who had undergone sleeve gastrectomy and 12 patients on a hypocaloric diet. For determining the long-term adaptation process the samples were taken six months after the treatment. In multivariate regression analyses, SAP, CLU, RBP4, PEDF, GLN, and C18:2 showed the strongest correlation to changes in body fat mass. The combined serum proteomic and metabolomic profiling reveals a link between the complement system and obesity and identifies both novel (C3b, CLU, VDBP, and all metabolites) and confirms previously discovered markers (PEDF, RBP4, C3, ATIII, and SAP) of body fat mass changes.
Assuntos
Proteínas do Sistema Complemento/metabolismo , Espectrometria de Massas/métodos , Obesidade/sangue , Tecido Adiposo/metabolismo , Adulto , Cirurgia Bariátrica/métodos , Biomarcadores/sangue , Biologia Computacional/métodos , Estudos Transversais , Eletroforese em Gel Bidimensional/métodos , Ensaio de Imunoadsorção Enzimática/métodos , Humanos , Processamento de Imagem Assistida por Computador , Focalização Isoelétrica/métodos , Estilo de Vida , Masculino , Metabolômica/métodosRESUMO
In industrialized countries, people spend more time indoors and are therefore increasingly exposed to volatile organic compounds that are emitted at working places and from consumer products, paintings, and furniture, with chlorobenzene (CB) and 1,2-dichlorobenzene (DCB) being representatives of the halogenated arenes. To unravel the molecular effects of low concentrations typical for indoor and occupational exposure, we exposed human lung epithelial cells to CB and DCB and analyzed the effects on the proteome level by 2-D DIGE, where 860 protein spots were detected. A set of 25 and 30 proteins were found to be significantly altered due to exposure to environmentally relevant concentrations of 10(-2) g/m(3) of CB or 10(-3) g/m(3) of DCB (2.2 and 0.17 ppm), respectively. The most enriched pathways were cell death signaling, oxidative stress response, protein quality control, and metabolism. The involvement of oxidative stress was validated by ROS measurement. Among the regulated proteins, 28, for example, voltage-dependent anion-selective channel protein 2, PDCD6IP protein, heat shock protein beta-1, proliferating cell nuclear antigen, nucleophosmin, seryl-tRNA synthetase, prohibitin, and protein arginine N-methyltransferase 1, could be correlated with the molecular pathway of cell death signaling. Caspase 3 activation by cleavage was confirmed for both CB and DCB by immunoblotting. Treatment with CB or DCB also caused differential protein phosphorylation, for example, at the proteins HNRNP C1/C2, serine-threonine receptor associated protein, and transaldolase 1. Compared to previous results, where cells were exposed to styrene, for the chlorinated aromatic substances besides oxidative stress, apoptosis was found as the predominant cellular response mechanism.