Construction of GLI1 shRNA expression vector and effects on the proliferation of ovarian cancer cells

摘要

Ovarian cancer SKOV3 cells were transfected for 48 hours by the mediation of LipofectamineTM 2000. CCK-8 kit and real time RT-PCR were employed to evaluate the effects of GLI1 mRNA transfection on the proliferation of SKOV3 cells, the apoptosis rates induced by paclitaxel and the cell cycle of SKOV3 cells following the treatments were measured with flow cytometry; and cyclinD1 protein expression was detected by Western blot. In the blank ontrol, negative-control, GLI1 shRNA-1 GLI1 shRNA-2, GLI1 shRNA-3, GLI1 shRNA-4 groups, of SKOV3 cells, the expressions of GLI1 mRNA relative to GAPDH were 1±0.00, 1.03±0.02, 0.73±0.07, 0.98±0.08, 0.53±0.08, 0.68±0.04, respectively. Contrast to blank control, GLI1 mRNA expression in GLI1 shRNA-1 GLI1 shRNA-3, GLI1 shRNA-4 groups declined (P<0.05). GLI1 mRNA expression in GLI1 shRNA-3 group was obviously lower than other 3 groups (P<0.05). Meanwhile the expression of GLI1 protein decreased. SKOV3 cell proliferation and CyclinD1 protein were significantly inhibited under the conditions transfected by GLI1 shRNA-3. The apoptosis rate increase was induced by paclitaxel after GLI1 down-regulation. The expressions of Bel-2 and CyclinD1 were decreased and the expression of Caspase3 was increased. GLI1 shRNA visibly inhibited the proliferation of ovarian cancer cells and induced the cell apoptosis; GLI1 signaling could induce cell proliferation by activating CyclinD1.