Objective: To construct a recombinant adenovirus expression vector containing the anti-oncogene PTEN and to investigate the effects of the PTEN gene on the proliferation of prostate cancer PC-3 cells and the expressions of cyclin D1 and p21 in the PC-3 cells.
Methods: The PTEN gene was amplified from the rat hippocampus by RT-PCR and cloned into the shuttle plasmid pEN-TR2A. The plasmids were constructed and amplified in 293A cells. Prostate cancer PC-3 cells were cultured in vitro and infected with the adenoviral vector carrying the PTEN gene (Ad-PTEN). The up-regulation of the PTEN protein was measured by indirect immuno-fluorescence assay; the expressions of PTEN, cyclin D1 and p21 in the cells infected with Ad-PTEN and Ad-LacZ were determined by
Results: The Western blot; and the effect of PTEN on the cell proliferation was detected by MTT assay and plate colony formation. recombinant adenoviral vector Ad-PTEN was successfully constructed. Western blot showed a significantly increased expression of the PTEN protein in the PC-3 cells infected with Ad-PTIEN (0.215 +/-0.065) as compared with that in the control ([0.052 +/-0.009], t = 4. 30, P <0.05) and the Ad-LacZ group ( [0. 056 +/- 0.008 ] , t =4.21, P <0.05). The expression of cyclin D1 was significantly lower in the Ad-PTEN-infected PC-3 cells (0. 256 +/- 0. 072) than in the control ( [0. 502 +/- 0. 087 ], t = 3.77, P < 0.05) and the Ad-LacZ group ([0.498 +/-0.081] , t =3.87, P <0.05), while the expression of p21 remarkably higher in the Ad-PTEN-infected PC-3 cells (0.589 +/-0. 076) than in the control ([0. 146 +/-0.026] , t = 9.55, P<0. 01) and the Ad-LacZ group ([0. 163 +/-0. 024] , t = 9.26, P <0.01). Ad-PTEN significantly inhibited the growth of the PC-3 cells (21.98%) at 48 h (t = 6.80, P <0.01). The colony formation rate of the PC-3 cells was (37.4 +/-4. 18)% in the Ad-PTEN group, significantly lower than (54.9 +/-4.81)% in the control (t =4.76, P<0.01) and (56.5 +/- 5.42)% in the Ad-LacZ group (t=4.83, P<0.01).
Conclusion: The expression of PTEN induced by Ad-PTEN can significantly inhibit the proliferation of PC-3 cells, down-regulate the expression of cyclin D1, and up-regulate the expression of p21.