RESUMEN
Nitric oxide (NO) is one of the most important interneural signaling molecules mediating hippocampal functions for learning and memory. The diurnal expression of neuronal NO synthase (nNOS) has been well studied. However, the temporal profile and underlying mechanisms of inducible NOS (iNOS) expression in a normal photoperiod or in altered light-dark cycles remain unclear. We examined the temporal profile of iNOS expression in adult male Sprague-Dawley rats maintained in a 12h-light/ 12h-dark photoperiod (12L/12D), which were pre-synchronized for 7 days before the experiment. The protein and mRNA levels of iNOS in the cortex and hippocampus were measured to examine the photic influences on iNOS expression. The results showed rhythmically changes of the levels of iNOS mRNA and protein in the hippocampus, but not in the cerebral cortex. The iNOS mRNA levels peaked at Zeitgeber time (ZT) 6 and ZT22, and the protein levels peaked at ZT8 and ZT18. Notably, the peaks in iNOS mRNA and protein levels in the 12L/12D group were 10 to 12 h apart, and the rhythmic pattern was absent in the 24-h period of the darkness group. In addition, the level of the phosphorylated cAMP response element binding protein (phospho-CREB) was the highest at ZT18, prior to a peak in iNOS mRNA expression at ZT22. A phospho-CREB-iNOS signaling pathway was further confirmed by the interaction of phospho-CREB and the iNOS DNA in histone complexes isolated by chromatin immunoprecipitation at ZT18. In conclusion, the photoperiod affects the diurnal expression of iNOS and activation of CREB in the hippocampus, which provide clues for the possible impact of circadian changes in hippocampal functions.
Asunto(s)
Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Hipocampo/enzimología , Óxido Nítrico Sintasa de Tipo II/genética , Fotoperiodo , Animales , Hipocampo/fisiología , Masculino , Fosforilación , ARN Mensajero/análisis , Ratas , Ratas Sprague-DawleyRESUMEN
OBJECTIVE: Toll-like receptors (TLRs) are important molecules for detecting both pathogen invasion and tissue damage. The expression of TLR4 is upregulated in ischemic stroke, at least in the subacute stage. However, the TLR downstream pathways in the context of stroke have not been well studied in previous research. The purpose of this study is to elucidate the TLR4 downstream pathways following ischemic stroke. DESIGN AND METHODS: In this study, 12 ischemic stroke patients and 12 controls were selected from among 89 ischemic stroke patients and 166 controls. The chosen subjects had the highest levels of TLR4 mRNA in the peripheral blood. The differences in the TLR downstream signaling pathways, which were studied by using an RT2 Profiler TM PCR array system (Qiagen), were analyzed. The differentially expressed genes were analyzed by using GeneSpring GX and visualized based on the TLR pathways in the Kyoto Encyclopedia of Genes and Genomes (KEGG). RESULTS: The genes upregulated in stroke patients were found to be involved in the MyD88-independent pathway and in UBE2V1-TRAF6 ubiquitin-mediated proteolysis. The genes were more expressed in extracellular space, receptor binding, and cytokine receptor binding by use of gene ontology (GO) terms than in control patients. CONCLUSIONS: We found that the MyD88-independent pathway and the ubiquitin-mediated proteolysis pathway, especially TRAF6, may be the most vital molecules among TLR downstream pathways in incidences of ischemic stroke.
Asunto(s)
Transducción de Señal/genética , Accidente Cerebrovascular/genética , Receptores Toll-Like/genética , Receptores Toll-Like/metabolismo , Isquemia Encefálica/genética , Isquemia Encefálica/patología , Estudios de Casos y Controles , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos , Proteolisis , Reproducibilidad de los Resultados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Accidente Cerebrovascular/patología , Ubiquitina/metabolismoRESUMEN
OBJECTIVES: Toll-like receptors (TLRs) are molecules conserved in evolution for detecting pathogen invasions and tissue damage and are involved in atherogenesis. This study explores the mRNA expression of TLRs and their probable role in further disease occurrence among ischemic stroke patients. DESIGN AND METHODS: A total of 89 ischemic stroke patients and 166 controls were recruited for this study. Total RNA was extracted and mRNA was reverse-transcribed to cDNA and was analyzed for TLRs and interleukin 8 (IL8). RESULTS: The TLR4 mRNA expression level is significantly higher in the stroke group. Conversely, IL-8 mRNA levels decreased significantly in the patient group. CONCLUSION: Our results suggest that TLR4 overexpression in mRNA levels is observed in stroke patients, which might account for the probable inflammatory injury before or after stroke. A reduction of IL-8 expression could result from the downregulatory effects of aspirin.