Figure 6.

MEK Regulates Pim-1 via ERK/AMPK/IL-23/STAT3 Cascade

(A) IL-23 is specifically upregulated by AZD6244. HT29 cells were treated with AZD6244 (100 nmol/L) for 2 h, and mRNA levels of IL-8, IL-10, IL-16, and IL-23 were then detected by real-time PCR.

(B) HT29 cells were treated with recombinant IL-23 (10 ng/mL) for 2 h or IL-23 siRNA (50 nmol/L) for 48 h. STAT3 phosphorylation and Pim-1 expression were then detected by western blot analysis.

(C) HT29 cells were treated with AZD6244 (100 nmol/L) for 2 h with or without 48-h pretreatment with a scramble or IL-23 siRNA (50 nmol/L), and protein expression or phosphorylation levels were then detected by specific antibodies as indicated.

(D) Same experiment as (C) with AZD6244 replaced by an AMPK inhibitor, compound C.

(E) HT29 cells were treated either with AZD6244 (100 nmol/L) for 2 h, and then IL-23 expression was detected by real-time PCR.

(F) HT29 cells were treated either with compound C (40 μmol/L) for 2 h, and then IL-23 expression was detected by real-time PCR.

(G) Mechanistic model for MEK inhibition-induced Pim-1 upregulation. MEK inhibition increases IL-23 expression through ERK/AMPK signaling, which then activates STAT3, thereby promoting Pim-1 transactivation.

Data are represented as mean ± SD. Comparisons of two groups are performed with Student's t test. ∗p < 0.05, ∗∗∗∗p < 0.0001.