Fig 6.

Overexpression of miR-146a in endotoxin-responsive cells inhibits p38 MAPK activation, prevents RBM4 phosphorylation, and downregulates proinflammatory cytokine synthesis in response to TLR4 stimulation. Endotoxin-responsive THP-1 cells were transfected with an miRNA negative control or miR-146a mimics. After 36 h, cells were stimulated for 0 to 4 h with 1 μg/ml of LPS. (A) Levels of miR-146a were measured after 1 h by real-time PCR. Data are means ± SD of three experiments. *, P < 0.05. (B) Levels of MKP-1, p38, and phospho p38 proteins were measured in whole-cell extract by Western blot analysis. (C) Levels of RBM4 and p-RBM4 (phospho-serine-309) were measured in the nuclear and cytoplasmic fractions by Western blot analysis. (D) After 4 h in LPS, culture supernatants and cells were harvested and lysed, and then TNF-α protein levels were measured by ELISA. Data are means ± SD of three experiments. *, P < 0.05. The results in panels B and C are representative of three experiments.