Fig. 5

CREB1 interacts with miR-181a-5p through a feedback loop in FD BMSCs. After si-CREB1 treatment for 48 h, CREB1 (a) and miR-181a-5p (b) in FD BMSCs were detected by quantitative real-time PCR. c Three primers were designed to cover the miR-181a promoter region and were used to identify CREB1 binding sites in the ChIP assay. All three putative CREB1 binding sites in the miR-181a promoter region were identified. d CREB1 mRNA expression was detected by real-time PCR in FD BMSCs transfected with miR-181a-5p mimics. e Protein expression of CREB1 and p-CREB1 was detected by western blot. f Immunofluorescence analysis of p-CREB1 expression in FD BMSCs transfected with miR-181a-5p mimics. g Effect of miR-181a-5p on a dual-luciferase reporter plasmid bearing wild-type (WT)/mutated (Mut) CREB1 binding sites was analyzed. Cells were cotransfected with either WT-CREB1 or Mut-CREB1 and miR-181a-5p mimics or miR NC. Firefly and Renilla luciferase activities were measured in cell lysates. Scale bar, 100 μm. The data are presented as the mean ± S.E.M. values (n = 3). *P < 0.05; **P < 0.01; ^#P > 0.05