Effects Of E2F-1 Gene Silencing On Biological Characteristics Of Human Glioma U251 Cells

Objective: To explore the influence of the E2F-1 transcription factor on the proliferation and invasion of glioma U251 cells.Brain glioma is the most common intracranial malignant tumor across different age groups,accounting for more than 50% of intracranial primary tumors.Surgical resection assisted with chemoradiotherapy is the main therapeutic regimen for glioma patients.Glioma has high infiltrative growth,and recurrence frequently occurs postoperatively.Recent studies have applied multilocus microsatellite typing in glioma,and the development of therapeutic targeted drugs for glioma,as a more effective and accurate treatment,has become a new direction of glioma treatment.E2F-1 is an important transcription factor that regulates cyclins.E2F-1 binds heterogeneous protein DP1/2 and functions as a critical transcription factor that regulates the cell cycle and apoptosis.E2F-1 is abnormally expressed in various malignant tumors and is an important pathogenic factor during tumorigenesis and progression.However,few studies have examined E2F-1 and brain glioma.In this study,we generated a recombinant lentiviral vector for sh RNA-mediated downregulation E2F-1 in U251 glioma cells.The silencing effect of the lentiviral vector on E2F-1 in U251 cells was verified by measuring m RNA and protein expression of E2F-1.MTT and Transwell assays were used to examine changes of proliferative and invasive capabilities of U251 cells after silencing E2F-1 gene.These results will help provide a basis for understanding the influence and mechanism of E2F-1 on the biological behaviors of U251 cells and may uncover new pathways for brain glioma treatment.Materials and methods: The human glioma U251 cell strain was used as a glioma cell line in this study.Establishment of cell line with E2F-1 gene interfered by lentivirus.Human glioma U251 cell strain was used as a study subject.E2F-1 gene silencing mediated by lentivirus made E2F-1 protein lowly express to establish cell model with silenced E2F-1 and the negative control,and screen cell line with stable expression.According to the difference in infected lentivirus,U251 cells were divided into E2F-1-sh RNA group,NC-sh RNA group,and normally cultured U251 cell group(Blank group).E2F-1 silencing was evaluated in the stable cell lines by real-time PCR and western blot analyses.U251 cell proliferation and invasion were examined using MTT and Transwell assays.Statistical analysis was performed using SPSS 18.0 and Graph Pad Prism 7.Data were expressed as mean ± SD and analyzed by one-way analysis of variance.P < 0.05 was considered to indicate statistical significance.Results: Fluorescence microscope observation of the U251 stable cell lines revealed more than 90% green fluorescent cells,indicating high infection efficiency.Real-time PCR and western blot confirmed significantly decreased E2F-1 m RNA and protein levels,respectively,in the E2F-1-sh RNA U251 stable cell line compared with the negative control(NC-sh RNA)cells and parental U251 cells(P < 0.05).The NC-sh RNA cells and parental cells showed no significant differences in E2F1 m RNA or protein expression(P > 0.05).MTT assays revealed that E2F-1-sh RNA U251 cells showed significantly reduced cell growth compared with the NC-sh RNA cells and U251 cells(P < 0.05).No significant difference was found between the NC-sh RNA and U251 cells(P > 0.05).Transwell experiments revealed significantly reduced numbers of invading tumor cells in the E2F-1-sh RNA U251 cells compared with NC-sh RNA and U251 cells(P < 0.05).No significant difference was found between the NC-sh RNA and U251 cells(P > 0.05).Conclusion: Silencing of the E2F-1 gene can significantly inhibit the proliferative and invasive capabilities of glioma U251 cells.Future studies should examine the mechanism of the E2F-1 transcription factor in brain glioma and evaluate its potential as a target for glioma treatment.