RESUMEN
Allergic conjunctivitis (AC) is one of the most common eye disorders in ophthalmology. In mice models, it has been suggested that control of allergic conjunctivitis is a delicate balance between Tregs and inflammatory migrating effector cells. Our aim was to evaluate the frequency of Tregs and the frequency of homing receptors expressing cells in peripheral blood mononuclear cells (PBMC) from patients with perennial allergic conjunctivitis (PAC). The analyses of phenotypic markers on CD4+ T cells and both soluble or intracellular cytokines were performed by flow cytometry. CD4+CD25+ cells were 15 times more frequent in PBMC from patients than HC; the vast majority of these CD4+CD25+ cells were FOXP3-, and most of CD4+ T cells were CCR4+ and CCR9+ cells. Upon allergen-stimulation, no significant changes were observed in frequency of Treg; however, an increased frequency of CD4+CCR4+CCR9+ cells, CD4+CD103+ cells and CD4+CD108+ cells with increased IL-5, IL-6, and IL-8 production was observed. These findings suggest an immune dysregulation in PAC, characterized by diminished frequency of Tregs and increased frequency of circulating activated CD4+ T cells; upon allergen-stimulation, these cells were expressing cell-surface molecules related to mucosa homing and were able to trigger an inflammatory microenvironment.
Asunto(s)
Conjuntivitis Alérgica/inmunología , Linfocitos T Colaboradores-Inductores/inmunología , Linfocitos T Reguladores/inmunología , Adolescente , Antígenos Dermatofagoides/inmunología , Niño , Preescolar , Conjuntivitis Alérgica/metabolismo , Citocinas/metabolismo , Femenino , Factores de Transcripción Forkhead , Humanos , Inmunofenotipificación , Subunidad alfa del Receptor de Interleucina-2/metabolismo , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/metabolismo , Recuento de Linfocitos , Masculino , Fenotipo , Receptores CCR/metabolismo , Receptores CCR4/metabolismo , Receptores de Quimiocina/metabolismo , Linfocitos T Colaboradores-Inductores/metabolismo , Linfocitos T Reguladores/metabolismoRESUMEN
Pterygium is one of the most frequent pathologies in ophthalmology, and is a benign, fibrovascular lesion originating from the bulbar conjunctiva. It is composed of an epithelium and highly vascular, subepithelial, loose connective tissue. The etiology of pterygium is not clearly understood; the most widely recognized originating factor is ultraviolet radiation. It has been proposed that pterygium and neoplasia have common features, raising the possibility that pterygium is a neoplastic-like growth disorder. In this study, proteomic analysis was performed to show that peroxiredoxin 2 is overexpressed in pterygia compared to healthy conjunctivas. Twelve pterygium specimens were obtained together with healthy conjunctival tissue from the same eyes. Total proteins of pterygia and healthy conjunctivas were analyzed in SDS-PAGE. This analysis showed protein bands expressed exclusively in pterygium samples at the range of 20-25 kDa. After this, 2D electrophoresis was performed for the separation of total proteins; differential spots expressed in pterygium were excised and sequenced. Mass spectrometry (MS) data were searched in the NCBInr and EST databases using the MASCOT program. The spot was identified as peroxiredoxin 2. Real-time PCR, western blot and immunohistochemistry showed that peroxiredoxin 2 was increased in pterygium compared to healthy conjunctiva. Although, these results suggest that overexpression of peroxiredoxin 2 in pterygium could protect the cell against oxidative stress-induced apoptosis, further studies are required to establish the functional role of peroxiredoxin 2 in pterygium to determine its role in peroxidation and apoptosis in this pathology.
Asunto(s)
Proteínas del Ojo/metabolismo , Peroxirredoxinas/metabolismo , Pterigion/enzimología , Adulto , Secuencia de Aminoácidos , Western Blotting , Conjuntiva/enzimología , Electroforesis en Gel Bidimensional , Electroforesis en Gel de Poliacrilamida , Proteínas del Ojo/química , Proteínas del Ojo/genética , Femenino , Humanos , Inmunohistoquímica , Focalización Isoeléctrica , Masculino , Espectrometría de Masas , Datos de Secuencia Molecular , Peso Molecular , Oxidación-Reducción , Peroxirredoxinas/química , Peroxirredoxinas/genética , Proteómica , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
BACKGROUND: Airway allergic diseases are regulated by interleukin (IL)-5, which causes infiltration of eosinophils into the bronchial epithelium, and by IL-4 which increases serum immunoglobulin E (IgE) production and promotes CD30 expression on Th cells. CD30 generates a costimulatory signal involved in apoptosis or cell proliferation, depending on the microenvironment. Our aims were: (i) to analyze if CD4+ CD30+ T cells from allergic patients proliferate in response to Dermatophagoides pteronyssinus, and (ii) if upon stimulation this cell population produces IL-4 and IL-5. METHODS: Peripheral blood mononuclear cell (PBMC) from 17 allergic rhinitis and mild allergic asthma patients and 12 healthy nonallergic individuals were stimulated with allergen in the presence or absence of anti-IL-4, anti-IL-5 or anti-IL-4Ralpha monoclonal antibodies (mAbs). TdT-mediated dUTP nick end-labeling (TUNEL) assay, 7-aminoactinomycin-D (7-AAD) intercalation, and flow cytometry were used to determine the CD4+ CD30+ blasts percentage, cell proliferation, apoptosis, and intracellular cytokines after 7 culture days. RESULTS: Cell proliferation induced with allergen showed that 90% of the allergen-stimulated blasts were CD4+, 50% of which were CD30+. Allergen-stimulated PBMC showed a progressive increase (mean: from 7% to 23%) of CD4+ CD30+IFN-gamma+ and CD4+ CD30+IL-4+ blasts which diminished (mean: 6%) after 5 culture days. In contrast, CD4+ CD30+IL-5+ blasts showed a continuous progression (from 12% to 24%) that maintained after 7 culture days. The vast majority of CD4+ CD30+ blasts were negative to 7-AAD or TUNEL. Additionally, a significant decrease (34%) was observed in the number of CD4+ CD30+ blasts when IL-4 was neutralized. CONCLUSIONS: These data suggest that specific allergen stimulation of PBMC isolated from allergic patients generates a nonapoptotic CD4+ CD30+ blast subset that produces IL-5.
Asunto(s)
Alérgenos/farmacología , Antígenos Dermatofagoides/inmunología , Linfocitos T CD4-Positivos/inmunología , Dermatophagoides pteronyssinus , Interleucina-5/biosíntesis , Adolescente , Adulto , Apoptosis/inmunología , Asma/sangre , Estudios de Casos y Controles , Proliferación Celular , Células Cultivadas , Femenino , Citometría de Flujo , Humanos , Antígeno Ki-1/inmunología , Activación de Linfocitos , Masculino , Probabilidad , Rinitis Alérgica Estacional/sangre , Muestreo , Sensibilidad y Especificidad , Estadísticas no ParamétricasRESUMEN
The atopic diseases are mediated by type I hypersensitivity. These disorders are multifactorial, although, the inherency is probably the most important cause. The disregulation of the Th1/Th2 immune response is one of the predominant factors on the generation and maintenance on the atopic process. We analyze the mechanisms of the regulation of the response Th1/Th2, the recent theories of the disregulation on this response as an etiology, the soluble factors as cytokines, chemokines and their receptors, surface cell markers and transcriptional factors recently mentioned to be important on this disregulation mechanism. Otherwise, we propose some general considerations and perspectives.
Asunto(s)
Hipersensibilidad Inmediata/inmunología , Membrana Celular/inmunología , Quimiocinas/inmunología , Citocinas/inmunología , Humanos , Células TH1/inmunología , Células Th2/inmunologíaRESUMEN
The porcine immune system is unique in the expression of CD4+CD8+ (double-positive, DP) lymphocytes. These cells have been associated with immunological memory due to their gradual increase with age, the expression of memory phenotype and their ability to respond to recall viral antigen. This work analyzes the biological function of CD4+CD8- and CD4+CD8+ lymphocytes in the immune response to porcine rubulavirus (PRv). CD4+CD8- cells isolated from pigs 3 weeks after infection with porcine rubulavirus proliferated in response to homologous virus and generated lymphoblasts which were predominantly of the CD4+CD8+ phenotype, whereas stimulation with mitogen induced proliferation but did not switch the phenotype. CD4+CD8- lymphocytes isolated after 10 weeks of infection proliferated in response to phytohemagglutinin (PHA) but did not proliferate in response to homologous virus and did not change their phenotype, whereas CD4+CD8+ lymphocytes proliferated in response to PHA and to viral antigen. The cytokine profile of both lymphocyte populations showed the presence of IL-2 and IL-10 transcripts, quantitation demonstrated that CD4+CD8+ cells expressed mainly IL-10, whereas CD4+CD8- lymphocytes expressed primarily IL-2. Our results show that CD4+CD8- lymphocytes in the early phase of porcine rubulavirus infection can be converted to double-positive cells expressing IL-10 in an antigen-dependent manner, and that CD4+CD8- T-cells late in infection do not acquire CD8.