Cx32 exerts its effect through negative regulation of Snail expression. (A) Knockdown of Cx32 increased Snail but not Slug or Twist-1 expression in SMMC-7721 cells, while overexpression of Cx32 decreased Snail but not Slug or Twist-1 expression in HepG2 cells, as evidenced by western blot analysis. (B) Expression of Snail was negatively regulated by Cx32 as confirmed by immunofluorescence assay (original magnification, ×400). (C) Snail mRNA was detected by real-time PCR in different groups. (D) Cell morphological changes of SMMC-7721 and HepG2 cells in the presence of both Cx32 and Snail modulation (original magnification, ×200). (E) Silencing of Snail in SMMC-7721 cells rescued Cx32 knockdown-induced EMT-related protein changes while upregulation of Snail in HepG2 cells abrogated Cx32 overexpression-induced EMT-related proteins modulation. (F) Invasive capacity of SMMC-7721 and HepG2 cells in the presence of both Cx32 and Snail modulation (original magnification, ×200). NC, negative control; OE, overexpression; SiC, siRNA of Cx32; SiS, siRNA of Snail; OEC, overexpression of Cx32; OES, overexpression of Snail. Data represent the mean ± SEM of three to four independent experiments. *P<0.01 vs. NC (C); *P<0.01 vs. control, #P<0.01 vs. SiC+SiS (or OEC+OES) (F).