Aim: To observe the growth inhibitory effect of wild-type Kras2 gene on a colonic adenocarcinoma cell line Caco-2.
Methods: Recombinant plasmid pCI-neo-Kras2 with wild type Kras2 open reading frame was constructed. The Caco-2 cells were transfected with either pCI-neo or pCI-neo-Kras2 using Lipofectamine 2000. The expression of wild type Kras2 was examined by Northern blot analysis. And the expression of wild type Kras2 protein was examined by Western blot analysis. The effects of wild-type Kras2 on cell proliferation were analyzed by monotetrazolium (MTT) assay, meanwhile analyses of cell cycle and spontaneous apoptosis rate were carried out by flow cytometry (FCM).
Results: The plasmid of pCI-neo-Kras2 was successfully established. The growth rate of cells transfected with pCI-neo-Kras2 was significantly lower than the control cells transfected with the empty pCI-neo vector (P<0.05). Cell cycle analysis revealed arrest of the pCI-neo-Kras2 transfected cells in G0/G1 phases, decreased DNA synthesis and decreased fractions of cells in S phase. The proliferative index of cells transfected with pCI-neo-Kras2 was decreased compared with the control cells (49.78% vs 64.21%), while the apoptotic rate of Caco-2 cells with stable Kras2 expression increased (0.30% vs 0.02%).
Conclusion: The wild-type Kras2 gene effectively inhibits the growth of the colonic adenocarcinoma cell line Caco-2.