Human Id3 expression in lung adenocarcinoma cell line A549 and its inhibiory effect on cell growth

Objective:To investigate the inhibitory effect of inhibitor of differentiation 3(Id3)on growth of human lung adenocarcinoma cell line A549.Methods:Recombinant eukaryotic expression vector pEGFP/Id3 was constructed and transfected into A549 cells by liposome-mediated method.Expression of pEGFP/Id3 in A549 cells was analyzed by flow cytometry(FCM),fluorescence microscopy,semi-quantitative RT-PCR and immunocytochemistry.The growth inhibitory rate of A549 cells was examined by MTT assay;cell cycle change was evaluated by PI(propidium iodide)staining method.Cell apoptotic rate and nuclear morphology were detected by Annexin V/7-AAD and Hoechst33258 staining. Results:The recombinant eukaryotic expression vector pEGFP/Id3 was successfully constructed.The expression of EGFP reached the peak 48-72 h after transfection;the expresion of pEGFP-transfected group was higher than that of the pEGFP/ Id3 group.RT-PCR and immunocytochemistry staining showed that Id3 mRNA and protein were effectively expressed in pEGFP/Id3-transfected A549 cells.The growth of cells in pEGFP/Id3 transfeeted cells was significantly inhibited 48-72 h after transfection(P0.01).More cells were blocked in G_0/G_1 phase in pEGFP/Id3-transfected group compared with pEGFP group(P0.05).Annexin V/7-AAD showed that the apoptotic rate of pEGFP/Id3 group([10.67±2.60]%) were significantly higher than those of the control group([2.35±0.95]%)and pEGFP group([3.39±2.21]%)(P0.05).Hoechst33258 staining also showed that the cells in pEGFP/Id3 group had typical apoptotic morphology. Conclusion:Exogenous Id3 gene expression in A549 cells can induce cell growth inhibition and apoptosis of A549 cells.