Yuan et al. BMC Cancer 2011, 11:57 http://www.biomedcentral.com/1471-2407/11/57 Page 8 of 10 Figure 4 Re-expressing ΔNp63 could significantly attenuate the effect of miR-203 on the inhibition of ESCC cell proliferation. (A, B) The expression level of ΔNp63 protein was detected by Western Blot at 48 hr posttransfection and normalized to that of b-actin. Results showed that the expression level of ΔNp63 protein was significantly higher in cells cotransfected with miR-203 and pcDNA-ΔNp63 as compared to the cells transfected with miR-203 and empty pcDNA. (C-F) Cell proliferation was evaluated by cell cycle analysis (C), annexin V-FITC/propidium iodide double staining (D), population doubling time determination (E) and clonogenic assay (F) as described in materials and methods. Results showed that the proliferative capacity of cells cotransfected with miR-203 and pcDNA-ΔNp63 was significantly higher than that of cells cotransfected with miR-203 and empty pcDNA. Transfection condition: 1 × 106 cells (Eca109 or TE-1) were cotransfected with 50 pmol of miR- 203 and pcDNA-ΔNp63 (or empty pcDNA). Data represent mean ± SEM from 4 independent experiments; *, P < 0.05 by t test. **, P < 0.01 by t test.