TF cytoplasmic domain regulates TLR4-induced ERK1/2 phosphorylation and TF expression via modulation of the p38 MAPK pathway. (A) LPS-induced ERK1/2 phosphorylation in BMMs is dependent on PI3-kinase and negatively regulated by p38 in wild-type, but not TF^ΔCT, BMMs. Cells were pretreated for 30 minutes with the p38 inhibitor SB203580 (10 μM), the PI-3 kinase inhibitor LY292004 (30 μM), or the MEK inhibitor U0126 (10 μM) followed by stimulation with LPS (1 μg/mL) for 10 minutes. A representative blot (left panel) and densitometric analysis of 4 experiments (right panel) are shown. (B) BMMs obtained by culturing of bone marrow cells in medium containing (20 ng/mL) recombinant mouse M-CSF. Cells were pretreated for 30 minutes with the p38 inhibitor SB203580 (10 μM) or DMSO (vehicle) followed by stimulation with LPS (1 μg/mL) for 10 minutes; phosphorylated ERK1/2 was determined by Western blotting. (C) LPS-induced TF induction in BMMs is negatively regulated by p38 in wild-type, but not TF^ΔCT, BMMs. BMMs were stimulated for 6 hours in the presence or absence of the indicated inhibitors at concentrations as described for panel A. TF mRNA induction was quantified by real-time PCR and TF activity was quantified by one stage clotting assay. *denotes statistically significant differences relative to wild-type control.