Figure 1

miR-148a-3p is frequently downregulated in bladder cancer and is regulated by DNA methylation. (a) Statistical analysis indicated that miR-148a-3p expression was significantly lower in bladder cancer tissues than in adjacent non-tumor tissues. (b) Representative images of ISH staining of TMA. miR-148a-3p localized to the cytoplasm. (c) miR-148a-3p levels in bladder cancer cell lines (J82, UM-UC-3 and T24) were detected and compared with the non-tumor urothelial cell line SV-HUC-1. (d) The demethylating agent 5-Aza-dC stimulated miR-148a-3p expression compared with DMSO-treated samples. (e) The regions analyzed by bisulfite-sequencing PCR (BSP) are indicated. (f) Methylation profile in T24 and UM-UC-3 cells. The open and filled circles represent the unmethylated and methylated CpG islands, respectively. Ten clones from each cell line were analyzed. (g) The fold change of miR-148a-3p level was similar in both bladder cancer cell lines and normal epithelial bladder cell. (h and i) Decreased DNMT1 expression was observed in miR-148a-3p-transfected T24 and UM-UC-3 cells via qRT-PCR and western blot. (j) The miR-148a-3p-targeting sites in the DNMT1 3′-UTR were mutated. (k) miR-148a-3p significantly suppressed the luciferase activity of vector that carried the DNMT1 3′-UTR but not control vector. (l) Statistical analysis indicated that DNMT1 expression in bladder cancer tissues was significantly higher than that in adjacent non-tumor tissues. (m) Representative images of IHC staining of TMA. DNMT1 localized to the nucleus. Error bars represent the S.E. obtained from three independent experiments; *P<0.05. Scale bar=100 μm