Fig. 3

(+)-JQ1 attenuates NFκB phosphorylation, translocation and transcription induced by LPS. a BV2 cells were transfected with either pGL3-NFκB or pGL3 along with pRL-TK. After 24 h, the cells were treated with either DMSO or (+)-JQ1 (400 nM) followed by LPS (100 ng/ml) stimulation. NFκB-mediated promoter activity was assessed by a luciferase assay (n = 3). b p65 nuclear translocation upon LPS (100 ng/ml) stimulation in either the presence or absence of (+)-JQ1 (400 nM) was detected by immunofluorescence staining and confocal microscopy. p65 immunoreactivity is shown in red, and nuclei were stained with DAPI (blue). (Scale bar = 20 μm). c BV2 cells preincubated with either (+)-JQ1 (400 nM) or DMSO were stimulated with LPS (100 ng/ml) for the indicated periods. Phosphorylation of IKKα/β (p-IKKα/β), IκBα (p-IκBα) and p65 (p-p65), as well as the total protein content, were analyzed by immnoblot. Representative results from three independent experiments are depicted here. The relative intensity of a given band was normalized (n = 6). d BV2 cells were treated with LPS (100 ng/ml) or JQ1 (400 nM), together with or without PDTC (p65 inhibitor). 24 h later, cell culture medium were collected and IL-1β, IL-6 and TNFα were tested by ELISA (n = 3). The data are presented as the mean ± SEM (* p ≤ 0.05, ** p ≤ 0.01)