Effects of RNAi-mediated Gene Silencing of LRIG3 Expression on Cell Cycle and Survival of Glioma Cells

【摘要】 The effects of RNAi-mediated gene silencing of LRIG3 expression on cell cycle and survival of human glioma cell line GL15 and the possible mechanisms were explored. The plasmids pGenesil2-LRIG3-shRNA1 and pGenesil2-LRIG3-shRNA2 were transfected into GL15 glioma cells respectively by using Metafectine, and the transfected cells that stably suppressed LRIG3 expression were selected by G418. The control cells were transfected with negative control shRNA. The changes in LRIG3 mRNA and protein levels were measured by RT-PCR and Western blot. The apoptosis rate and cell cycle were analyzed by flow cytometry. As compared with the negative shRNA-transfected GL15 cells, LRIG3 mRNA expression in GL15 cells transfected with pGenesil2-LRIG3-shRNA1 and pGenesil2-LRIG3-shRNA2 was silenced by 52.4%, 63.8%, and LRIG3 protein expression was re-duced by 50.9% and 67.4% respectively. The LRIG3-specific siRNA transfected cells had higher proliferation rate than control cells. Cell cycle analysis showed that silencing LRIG3 increased the percentage of G2/M phase cells and the proliferation index significantly (P<0.01). Silencing LRIG3 could inhibit the apoptosis of GL15 cells (P<0.05). These findings suggest that the siRNA targeting LRIG3 gene shows a dramatic inhibitory effect on RNA transcription and protein expression, then promoting the proliferation of GL15 cells, arresting GL15 cells in G2/M phase, and suppressing apoptosis of GL15 cells.更多还原

【Abstract】 The effects of RNAi-mediated gene silencing of LRIG3 expression on cell cycle and survival of human glioma cell line GL15 and the possible mechanisms were explored. The plasmids pGenesil2-LRIG3-shRNA1 and pGenesil2-LRIG3-shRNA2 were transfected into GL15 glioma cells respectively by using Metafectine, and the transfected cells that stably suppressed LRIG3 expression were selected by G418. The control cells were transfected with negative control shRNA. The changes in LRIG3 mRNA and protein levels were measured by RT-PCR and Western blot. The apoptosis rate and cell cycle were analyzed by flow cytometry. As compared with the negative shRNA-transfected GL15 cells, LRIG3 mRNA expression in GL15 cells transfected with pGenesil2-LRIG3-shRNA1 and pGenesil2-LRIG3-shRNA2 was silenced by 52.4%, 63.8%, and LRIG3 protein expression was re- duced by 50.9% and 67.4% respectively. The LRIG3-specific siRNA transfected cells had higher proliferation rate than control cells. Cell cycle analysis showed that silencing LRIG3 increased the percentage of G2/M phase cells and the proliferation index significantly (P<0.01). Silencing LRIG3 could inhibit the apoptosis of GL15 cells (P<0.05). These findings suggest that the siRNA targeting LRIG3 gene shows a dramatic inhibitory effect on RNA transcription and protein expression, then promoting the proliferation of GL15 cells, arresting GL15 cells in G2/M phase, and suppressing apoptosis of GL15 cells.更多还原