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Stable expression of programmed cell death 4 gene in glioma cell line and its influence on proliferation of tumor cells
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Abstract:
Abstract Objective: To establish a glioma cell line U251 stably expressing programmed cell death 4(PDCD4) gene, and to observe the influence of exogenous PDCD4gene on the proliferation and cell cycle of U251 cells.Methods: Recombinant eukaryotic expression vector pEGFPPDCD4 was transfected into human glioma cell line U251 by Lipofectamine 2000, and the U251 cells stably expressing PDCD4 were established by G418 selection. Reverse transcription polymerase chain reaction (RTPCR) and Western blotting were employed to detect the expression ofPDCD4 mRNA and protein. Furthermore, cell proliferation and colony forming ability were determined by cell counting and colony formation assay; the cell cycle was detected by FACS. Results: High expression of PDCD4 mRNA and protein was observed in U251 cells transfected with pEGFPPDCD4, whereas no PDCD4 mRNA and protein expression was detected in the nontransfected and vectortransfected cells. Further, cells transfected with pEGFPPDCD4 grew more slowly and had lower colony formation rate than cells of the other two control groups(P<0.01). Moreover, transfection of PDCD4 significantly reduced the number of cells at the G2/M phase and increased the cells at the S phase(P<0.05). Conclusion: Tumor suppressor gene PDCD4 can effectively inhibit cell proliferation and colony formation by altering their cell cycle.
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Project Supported:
Supported by the Scientific Research Foundation for Outstanding Young Scientists of Shandong Province (No.2006BS03064);Chinese Postdoctoral Science Foundation (No.20070411096).
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