Figure 1.
Immunohistochemical analysis of Pokemon expression in HCC tissue (Test Group) and adjacent noncancerous liver tissue (Control Group).
(A). Typical representative immunohistochemical results from one pair of HCC tissue (HCCT) and adjacent noncancerous liver tissue (ANLT) using an anti-Pokemon antibody. Pokemon was stained brown in granules. (B). A bar graph representing the relative expression level of Pokemon in the HCCT and ANLT groups, as evaluated by positive staining points. (A paired-samples t-test was used to compare the data from the two groups, *P<0.01).
Table 1.
The expression of Pokemon protein in HCC.
Figure 2.
Pokemon protein expressions in human HepG2 and Huh-7 cell lines and the siRNA silencing effects of Pokemon.
(A) Western blot analysis of Pokemon protein expression in the HepG2 and Huh-7 cell lines. The Pokemon expression in HL-7702 cells was used as a control, and the expression of β-actin was used as an internal control; (B) RT–PCR analysis of the Pokemon RNA level in HepG2 and Huh-7 cells that were stably transfected with Pokemon siRNA or the Pu6 vector control. The Pokemon expression levels in wild type HepG2 and Huh-7 cells were used as positive controls. (C) Western blot analysis to confirm the siRNA silencing of Pokemon, with β-actin used as a loading control.
Figure 3.
Functional changes in HepG2 and Huh-7 cells induced by Pokemon silencing.
(A). MTT assay analysis of cell proliferation in HepG2-siPokemon, Huh-7-siPokemon and control cells (*P<0.01). (B). BrdU assay analysis of cell proliferation in HepG2-siPokemon, Huh-7-siPokemon and control cells after 24 hours of incubation (*P<0.01). (C). Representative photographs from the migration assay and a histogram of the quantification of HepG2-siPokemon, Huh-7-siPokemon and control cells (*P<0.01). (D). Representative photographs from the wound healing assay in HepG2-siPokemon and control cells.
Figure 4.
The effect of Pokemon on Akt and ERK signals as well as critical members of the cell cycle progression pathway.
(A). Western blot analysis for the phosphorylation levels of Akt, Erk, PTEN, GSK-3β and c-Raf in HepG2-siPokemon and control cells. (B). Western blot analysis for cell cycle-related proteins in HepG2-siPokemon and control cells.
Figure 5.
Pokemon silencing inhibits the proliferation of HepG2 cells in vivo.
Nude mice were subcutaneously injected with HepG2-siPokemon or HepG2-Pu6 cells, and the tumors were excised after 16 days. (A, B). Representative images of tumors in nude mice. (C). The tumor volumes were recorded every three days for 16 days. The figure shows the mean ± SD (8 mice per group, *P<0.01). (D). Confirmation that Pokemon knockdown and decreased P-Akt levels are maintained in vivo.