Figure 3.

SIC phosphorylates p38 MAP kinase in THP1 cells and activates NF-κB

(A and C) Representative images of Western blot analysis of cell extracts from THP1 and Detroit 562 cells incubated with 5 μg/mL SIC +/− 2.5% plasma (P). Membranes were incubated with anti-p38 MAPK (p38) and anti-phospho-p38 MAPK (p-p38) antibodies.

(B and D) Band intensity analysis of Western blots displayed in A and C. The band intensity of phosphorylated p38 was normalized to the band intensity of p38 MAPK.

(E–H) Cells were incubated with 5 μg/mL SIC alone or in combination with E: 2.5% plasma F: 200 ng/mL HRG G: 5 μg/mL clusterin H: 5 μg/mL lysozyme for 18 hr. Cells were also incubated with plasma and the plasma proteins alone.

(I) THP1 cells were incubated with 5 μg/mL, 2.5 μg/mL or 1.25 μg/mL SIC in the presence or absence of 2.5% plasma (P) for 18 hr.

(J) THP1 cells were differentiated into macrophages with 100 nM PMA and the activation of NF-κB was analyzed after incubation with 5 μg/mL SIC +/− 2.5% plasma (P).

NF-κB activation was detected as color changes of QuantiBlue solution, absorbance was measured at 655 nm. All data represent mean ± SEM of 3-5 independent experiments, one-way ANOVA, Dunnett's multiple comparison test, with single pooled variance. ∗p < 0.05, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. ####p < 0.0001 compared to untreated control.