Objectives: To examine the effects of RUNX3 on cell proliferation and apoptosis and the expression level of Smad4 mRNA in the bladder cancer cell line of T24 by transfection with recombinant plasmid of pIRES-EGFP-RUNX3.
Methods: The recombinant plasmid of pIRES-EGFP-RUNX3 was constructed successfully. Cultured T24 cells were divided into three groups, including control group, empty vector group,and recombinant plasmid group. The cells in empty vector group and recombinant plasmid group were respectively transfected by pIRES-EGFP and pIRES-EGFP-RUNX3 The cells were harvested at 24 h after the transfection, the variation of cell morphology was examined by fluorescence microscopy. The cell apoptosis was detected by flow cytometry. The expression level of RUNX3 and Smad4 mRNA was measured by RT-PCR.
Results: Cell death was observed in two transfection groups. At 24 h after transfection,the apoptosis rate was (3.23±0.45)% in control group, (8.98±1.62)% in empty vector group and (43.61±2.69)% in recombinant plasmid group. The expression level of RUNX3 mRNA was 2.79±0.36,detected only in recombinant plasmid group, which was significantly up-regulated compared with the other two groups (P<0.05).
Conclusions: The expression level of Smad4 mRNA was up-regulated by transfection with pIRES-EGFP-RUNX3,which also inhibited cell proliferation and promoted cell apoptosis.The tumor suppressor gene of RUNX3 could regulate the bladder cancer cell proliferation and apoptosis by TGF-β/Smad signaling pathway.
Keywords: Bladder cancer; RUNX3; Smad4; Transforming growth factor-β.