Fig. 4

MiR-335 was down-regulated by DNA methylation in GC cell lines and tissues. a SGC-7901 and MKN-45 cells were cultured in the absence or presence of the demethylating agent 5-aza for 72 h. MiR-335 levels in treated cells were significantly higher than those in control cells. Error bars represent the SD from five replicates. **P < 0.01 between groups. b SGC-7901 and MKN-45 cells were cultured in the absence or presence of 5-aza for 72 h. CRKL mRNA levels were similar in treated and negative control cells (P > 0.05). Error bars represent the SD from five replicates. c CRKL expression levels were significantly reduced after 5-aza treatment, indicated by western blot analysis. d DNA methylation BSP in GC cell lines. Five single clones are represented for each sample. CpG islands are depicted and each CpG site is indicated by a vertical bar. Black and white circles represent hypermethylation and hypomethylation, respectively