Background & Objective: Serine/threonine kinase (AKT) mediates a downstream signal transduction pathway of epidermal growth factor receptor (EGFR), and plays an important role in survival and apoptosis of tumor cells. This study was designed to investigate regulation of antisense AKT2 (AS-AKT2) and dominant-negative AKT2 (DN-AKT2) constructs on proliferation and apoptosis of glioma cell line TJ905.
Methods: AS-AKT2 and DN-AKT2 were transfected into TJ905 cells, 3 stably transfected clones were randomly selected from each group, and amplified for further study. The expression of AKT2 was detected by in situ hybridization and Western blot. Proliferation rate of TJ905 cells was evaluated by MTT assay and Ki67 labeling index (Ki67 LI), cell apoptosis was detected by TUNEL method.
Results: AKT2 expression was significantly inhibited in TJ905 cells transfected with AS-AKT2, while no significant change observed in TJ905 cells transfected with DN-AKT2. Compared with parental TJ905 cells and TJ905 cells transfected with empty vector, the 6-day survival rates of TJ905 cells transfected with AS-AKT2, and DN-AKT2 dramatically dropped to 49.3%-51.5%, and 50.5%-55.2% (P< 0.001). The Ki67 LI of TJ905 cells transfected with AS-AKT2, and DN-AKT2 dropped to 57.5%-59.3% and 59.2%-61.0%, lower than those of parental TJ905 cells (76.5%), and TJ905 cells transfected with empty vector (74.8%) (P < 0.05). Nearly no apoptotic cells found in parental TJ905 cells or TJ905 cells transfected with empty vector, however, apoptosis indexes (AIs) of TJ905 cells transfected with AS-AKT2, and DN-AKT2 prominently increased to 8.8%-8.5%, and 7.5%-7.8% (P< 0.05). The inhibitory effect of AS-AKT2 on cell growth was statistically stronger than that of DN-AKT2 (P< 0.001), while no difference was detected in Ki67 LI and AI between AS-AKT2 and DN-AKT2 transfected TJ905 cells.
Conclusion: AKT2 pathway may modulate proliferation and apoptosis of glioma cells, indicating that it may be a potential therapeutic target of gene therapy of malignant glioma.
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