Effect of LPS and conditioned medium of LPS-treated glial or neuronal cultures on MAPKs activation in endothelial cells and neutrophil transmigration. Endothelial primary cultures were left untreated (UT) or were treated with LPS (10 ng/ml) for 15, 30 or 60 min. Cell lysates were assayed for JNK and ERK1/2 activation by Western blot analysis. Levels of phosphorylated MAPKs were analyzed semi-quantitatively, normalized relative to total MAPKs, and represented as fold increase compared to untreated cultures (a). Endothelial primary cultures were left untreated (UT), were treated with LPS, or were treated with conditioned medium of untreated glial (C Glia CM) or neuronal cultures (C Neu CM), or conditioned medium of LPS (10 ng/ml for 24 hours)-treated glial (LPS Glia CM) or LPS (10 ng/ml for 24 hours)-treated neurons (LPS Neu CM), and neutrophil trans-endothelial migration was assayed (b). Endothelial primary cultures were treated with conditioned medium of neuronal cultures treated with VIPER or CP7 (2 μM) in the absence or the presence of LPS (10 ng/ml) (LPS Neu CM), and neutrophil trans-endothelial migration was assayed (c). Data are presented as mean ± SD of three independent experiments carried out on separate cultures. *P <0.05, **P <0.01, ***P <0.001 LPS-treated versus untreated cultures, ##P <0.01 LPS + VIPER versus LPS + CP7, using one-way ANOVA and Tukey’s multiple comparison post-hoc test. ANOVA, analysis of variance; ERK1/2, extracellular-signal regulated kinase1/2; JNK, c-Jun-N-terminal kinase; LPS, lipopolysaccharide; MAPK, mitogen-activated protein kinase.