Figure 3.

Effects of NFκB and MAPK inhibitors on TNF-α-mediated MMP-1 and MMP-13 expression in dental pulp (A, left) and PDL (A, right) cells. Cells were pre-treated with BMS345541 (IKK phosphorylation inhibitor; 5 µM; 1.27 mg/mL), U0126 (MEK-1/2 inhibitor; 10 µM; 4.03 mg/mL), SB203580 (p38 inhibitor; 10 µM; 3.77 mg/mL), and SP600125 (JNK inhibitor; 10 µM; 2.02 mg/mL) for 1 hr and stimulated with 10 ng/mL of TNF-α for 24 hrs. Western immunoblots show basal levels of MMP-1 and MMP-13, after stimulation with the inhibitors plus TNF-α or TNF-α alone. Coomassie blue staining shows equal loading for all lanes. (B) Time-course experiments showing the effects of TNF-α on p38 (43 kDa) phosphorylation levels after treatment of dental pulp cells with TNF-α (10 ng/mL) for the indicated periods. Ratios of phosphorylated to total p38 levels were calculated by densitometric analysis and are shown above the panels. The effects of p38 MAPK inhibition on TNF-α-mediated DPP, DSP (C), and MMP-1 (D) expression in dental pulp cells were evaluated by Western immunoblotting. Cells were pre-treated with 10 µM of p38 inhibitor (SB203580) for 1 hr and then treated with TNF-α (10 ng/mL) plus p38 inhibitor (10 µM) for 6, 24, and 48 hrs. (E) Effects of p38 inhibition on mineralized nodule formation were assessed as in Fig. 1. Graphs depict mean and standard deviation from 3 different cell donors; *p < 0.05.