LPS-TLR4 signalling is active in both WT and conditional LysMcre-gp96flox-mice.

<p>Peritoneal macrophages were isolated from both WT and conditional gp96-KO and treated with lipopolysaccharides (LPS) (100ng/ml) for 30, 60 and 120 minutes. A and B) Total protein was isolated after LPS treatment and Western blots performed for p-IκBα, IκBα, p-NF-κB and β-actin. Graphs show the densitometry quantification of the blots (n = 3). C) After LPS treatment, macrophages from both WT and conditional gp96-KO were permeabilized and the expression of p- NF-κB was analysed by flow cytometry. The histograms are representative of three independent experiments. D) RNA was isolated after LPS treatment and the expression of IL-6, IL-8 and TNF-α mRNA was measured by real-time quantitative PCR. “Vh” represents macrophage treated with vehicle. Graphs show the fold induction of each gene (n = 4). In all cases data are expressed as mean±SEM. Statistical analysis was performed by One-way ANOVA followed by a Newman-Keuls test. * = <i>P</i><0.05, *** = <i>P</i><0.001 vs respective non-treated macrophages and ### = <i>P</i><0.001 vs LPS-treated macrophages from WT mice.