Figure 6
From: Micheliolide inhibits LPS-induced inflammatory response and protects mice from LPS challenge
MCL inhibits PI3K/Akt/p70S6K activation after LPS stimulation, accounting for the decreased IL-10 expression.
Raw264.7 cells (1 × 106/well) were plated in 6-well plates overnight and stimulated with LPS (100 ng/mL) with or without MCL (10 μM) for the indicated time periods. (a) Phospho-Akt at Ser473, total Akt, phospho-p70S6K and β-Actin were detected by Western blot. (b) Raw264.7 cells (1 × 106/well) were seeded overnight and pretreated with MCL (10 μM) or LY294002 (10 μM) for 30 min. Subsequently, cells were stimulated with LPS (100 ng/mL) or MCL (10 μM) as indicated for 15 min or 30 min. Western blot analyses of Akt (Ser473) phosphorylation, total Akt and β-Actin were carried out. (c,d) Raw264.7 cells were transfected with Akt-expressing plasmid (Myr-Akt-HA) and empty plasmid (mock), respectively. Phosphorylation of Akt (Ser473) was confirmed by Western blot. Forty-eight hours after transfection, cells were stimulated as indicated and IL-10 secretion was tested using ELISA. Similar results were obtained in three independent experiments. Data are shown as mean ± SD of three independent experiments; *p < 0.05.
