FIGURE 1.

miR-365 is a negative regulator of IL-6. A, HEK293 cells were co-transfected with the indicated miRNA expression construct (0.4 μg), pMIR-IL6 3′-UTR reporter plasmid (0.2 μg), and pRL-TK (0.05 μg). At 24 h post-transfection, cells were collected, and luciferase activity was measured. Luciferase activity of the empty vector was regarded as 1. *, p < 0.01, as compared with the empty vector group. B, mimics (20, 40, and 60 nm) or inhibitors (60, 80, and 100 nm) of miR-365 were co-transfected with pMIR-IL-6 3′-UTR reporter plasmid into HEK293 cells. A scrambled mimic (60 nm) or inhibitor (100 nm) was used as control. Luciferase activity was measured at 24 h post-transfection. The luciferase activity of the scrambled mimic or inhibitor was regarded as 1. C and D, HeLa cells were transfected with the indicated miRNA oligonucleotides. Poly(I:C) or poly(dAT:dAT) were transfected at 24 h after the initial transfection. Culture supernatants were collected 24 h later, and IL-6 was measured using a human IL-6 ELISA kit (C). Total RNA was extracted from cells, and real-time PCR assay was performed with specific primers listed in “Experimental Procedures” (D).