Figure 6

Basal and NECA-induced leukemia inhibitory factor (LIF) expression and secretion levels in primary astrocytes are dependent on ERK1/2- and p38- but not on JNK-MAPK activation. Primary cortical astrocytes were treated without or with NECA (1 μM) for 2 hours (for real-time PCR) or 4 hours (for ELISA). Where indicated, astrocytes were pre-treated for 2 hours with specific inhibitors of ERK1/2 (U 0126, 5 μM), p38 (SB 203580, 10 μM) or JNK (SP 600125, 10 μM) pathways, prior to NECA stimulation. (A) shows real-time PCR analysis of LIF gene expression (relative to GAPDH). Data are normalized to the control and presented as Mean ± SEM of three independent experiments. P < 0.05. (B) shows a representation of sandwich ELISA experiment performed to detect LIF content in supernatants of cultured astrocytes. Each bar corresponds to the mean concentration of LIF in triplicate samples; error bars indicate SEM. Observations were confirmed by repeating the experiments two additional times. P < 0.05. ERK, extracellular signal-regulated kinase; JNK, c-Jun N-terminal kinase; MAPK, mitogen-activated protein kinase; NECA, 5′-N-ethylcarboxamide.