Propofol Ameliorates Microglia Activation by Targeting MicroRNA-221/222-IRF2 Axis

Figure 4

miR-221/222 directly targets IRF2. (a) Predictive targets of miR-221/222 from three independent databases (TargetScan, miRDB, and miRBase). (b) IRF2 protein expression in BV2 microglia after transfection of miR-221 and miR-222 mimics was analyzed by Western blotting; β-actin was loaded as a control. (c) Modeling miR-221 target sequences in 3-UTR of IRF2; a mutant 3-UTR with substitutions in the complementary site for the seed region of IRF2 was used. (d) BV2 cells were transfected with luciferase reporter vector (IRF2-WT or IRF2-Mut) and miR-221 mimics, and luciferase reporter activity was measured after transfection for 48 h. (e) Modeling miR-222 target sequences in 3-UTR of IRF2; a mutant 3-UTR with substitutions in the complementary site for the seed region of IRF2 was used. (f) BV2 cells were transfected with luciferase reporter vector (IRF2-WT or IRF2-Mut) and miR-222 mimics, and luciferase reporter activity was measured after transfection for 48 h. (g) IRF2 protein expression in BV2 microglia after LPS stimulation and/or transfection with pcDNA3.1-IRF2 was analyzed by Western blotting; β-actin was loaded as a control. (h) The effects of IRF2 overexpression on the mRNA expression of inflammatory genes (Il1b, IL6, Tnf, Ptgs2, and Nos2) in LPS-primed BV2 cells were determined by real-time qPCR analysis (). (i) The effects of IRF2 overexpression on the level of inflammatory cytokines (IL-1β, IL-6, and TNF-α) in conditioned medium from LPS-primed BV2 cells were analyzed by ELISA (). , , and .

(a)

(b)

(c)

(d)

(e)

(f)

(g)

(h)

(i)