FIGURE 4.

ERK/CREB signalling regulated the expression of miR‐212‐3p in HBeAg‐induced macrophages. After pre‐treated with DMSO or inhibitor of ERK (PD98059 20 μmol/L), JNK (SP600125 20 μmol/L) and p38 (SB203580 20 μmol/L) for 1 h, RAW264.7 macrophages were stimulated with HBeAg for 24 h, and then, the level of miR‐212‐3p was evaluated with q‐PCR (A). The expression of non‐ phosphorylation and phosphorylation of CREB in RAW264.7 macrophages was analysed with Western blot after stimulated by HBeAg for different time (0, 30, 60, 120 min) (B). RAW264.7 macrophages were pre‐treated with DMSO or inhibitor of ERK (PD98059, 10 μmol/L), JNK (SP600125, 10 μmol/L) and p38 (SB203580, 10 μmol/L) for 1 h and then were treated with HBeAg for 30 min, the level of non‐ phosphorylation and phosphorylation of CREB was measured by Western blot (C). After pre‐treated with DMSO or inhibitor of CREB (KG‐501, 10 μmol/L) for 1 h, RAW264.7 macrophages were stimulated with HBeAg for 24 h. q‐PCR analysed the level of miR‐212‐3p (D). RAW264.7 macrophages nuclear lysates were immunoprecipitated by anti‐CREB antibody or non‐specific IgG antibody. The antibody‐bound DNA sequences were assessed by standard end‐point PCR and agarose gel electrophoresis. This graph shows the binding of CREB with the miR‐212‐3p proximal promoter with or without stimulation by HBeAg (E). Similar results were obtained in three independent experiments (mean ± SD of triplicates in A, D). *P < 0.05, **P < 0.01, ***P < 0.001