RESUMEN
The role of the aryl hydrocarbon receptor (AhR) in hemostasis has recently gained increased attention. Here, we demonstrate, by qRT-PCR and western blot, that human platelets express both AhR mRNA and AhR protein. AhR protein levels increase in a dose dependent manner when incubated with either 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or omeprazole. Treatment of platelets with puromycin blocks increased AhR protein synthesis in the presence of AhR activators. Additionally, treatment of platelets with either activator results in phosphorylation of p38MAPK and cPLA2, two key signaling molecules in platelet activation pathways. Using the AhR competitive inhibitors alpha naphthoflavone and CH-223191, we show that phosphorylation of p38MAPK is AhR dependent. Further, inhibition of p38MAPK blocks downstream cPLA2 phosphorylation induced by TCDD or omeprazole. Treatment with AhR activators results in platelet priming, as demonstrated by increased platelet aggregation, which is inhibited by AhR antagonists. Our data support a model of the platelet AhR non-genomic pathway in which treatment with AhR activators results in increased expression of the AhR, phosphorylation of p38MAPK and cPLA2, leading to platelet priming in response to agonist.
Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/agonistas , Plaquetas/efectos de los fármacos , Omeprazol/toxicidad , Activación Plaquetaria/efectos de los fármacos , Dibenzodioxinas Policloradas/toxicidad , Inhibidores de la Bomba de Protones/toxicidad , Receptores de Hidrocarburo de Aril/agonistas , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/antagonistas & inhibidores , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Plaquetas/metabolismo , Relación Dosis-Respuesta a Droga , Activación Enzimática , Fosfolipasas A2 Grupo IV/metabolismo , Humanos , Ligandos , Fragmentos de Péptidos/farmacología , Fosforilación , Agregación Plaquetaria/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Puromicina/farmacología , ARN Mensajero/metabolismo , Receptores de Hidrocarburo de Aril/antagonistas & inhibidores , Receptores de Hidrocarburo de Aril/genética , Receptores de Hidrocarburo de Aril/metabolismo , Receptores de Trombina/agonistas , Receptores de Trombina/metabolismo , Transducción de Señal/efectos de los fármacos , Factores de Tiempo , Regulación hacia Arriba , Proteínas Quinasas p38 Activadas por Mitógenos/antagonistas & inhibidores , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Increasing evidence suggests a role for a systemic pro-coagulant state in the pathogenesis of cardiac dysfunction subsequent to inhalation of airborne particulate matter (PM). We evaluated platelet activation, systemic cytokines and pulmonary gene expression in mice exposed to concentrated ambient particulate matter (CAPs) in the summer of 2008 (S08) and winter of 2009 (W09) from the San Joaquin Valley of California, a region with severe PM pollution episodes. Additionally, we characterized the PM from both exposures including organic compounds, metals, and polycyclic aromatic hydrocarbons. Mice were exposed to an average of 39.01 µg/m(3) of CAPs in the winter and 21.7 µg/m3 CAPs in the summer, in a size range less than 2.5 µm for 6 h/day for 5 days per week for 2 weeks. Platelets were analyzed by flow cytometry for relative size, shape, CD41, P-selectin and lysosomal associated membrane protein-1 (LAMP-1) expression. Platelets from W09 CAPs-exposed animals had a greater response to thrombin stimulation than platelets from S08 CAPs-exposed animals. Serum cytokines were analyzed by bead based immunologic assays. W09 CAPs-exposed mice had elevations in IL-2, MIP-1α, and TNFα. Laser capture microdissection (LCM) of pulmonary vasculature, parenchyma and airways all showed increases in CYP1a1 gene expression. Pulmonary vasculature showed increased expression of ICAM-1 and Nox-2. Our findings demonstrate that W09 CAPs exposure generated a greater systemic pro-inflammatory and pro-coagulant response to inhalation of environmentally derived fine and ultrafine PM. Changes in platelet responsiveness to agonists, seen in both exposures, strongly suggests a role for platelet activation in the cardiovascular and respiratory effects of particulate air pollution.
Asunto(s)
Contaminantes Atmosféricos/toxicidad , Citocinas/sangre , Regulación de la Expresión Génica/efectos de los fármacos , Material Particulado/toxicidad , Activación Plaquetaria/efectos de los fármacos , Estaciones del Año , Animales , California , Monitoreo del Ambiente , Perfilación de la Expresión Génica , Exposición por Inhalación , Pulmón/irrigación sanguínea , Pulmón/efectos de los fármacos , Pulmón/inmunología , Pulmón/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Tamaño de la PartículaRESUMEN
Mixed connective tissue disease (MCTD), undifferentiated connective tissue disease and overlap syndromes are autoimmune systemic diseases that must be differentiated. Antibodies against the U1-ribonucleoprotein complex --spliceosome-- allows the diagnosis of mixed connective tissue disease. Links between the immunologic and clinical phenomena are emerging. Longitudinal studies of patients with MCTD highlight the impact of pulmonary hypertension and contribute to define the disease. Immunogenetic studies hold MCTD as an independent disease.