Epstein-Barr Virus_Encoded LMP1 Upregulates MicroRNA-21 to Promote the Resistance of Nasopharyngeal Carcinoma Cells to Cisplatin-Induced Apoptosis by Suppressing PDCD4 and Fas-L
Figure 2
LMP1 confers resistance to cisplatin in NPC cells.
(A) CNE2 and HONE1 stable cell lines were treated with 0 or 2.0 μg/ml cisplatin. The MTT assay was used to determine the cell proliferation every 24 h for six days (* P < 0.05; ** P < 0.01 vs vector control cells). (B) Colony formation assays were performed for HONE1/LMP1 and HONE1/Vector cells, which were treated with the indicated doses of cisplatin. The upper panel shows the images of the colonies stained by crystal violet. The lower panel is the survival fraction from three independent experiments (* P < 0.05, *** P < 0.001 vs vector control cells). (C) Stably transfected cell lines were treated with various concentrations of cisplatin for 72 h to determine the IC50 of cisplatin. The IC50 values are presented as the means ± SD of three independent experiments. The IC50 is the concentration of chemotherapeutic drug (e.g., cisplatin) that causes a 50% inhibition of tumor cell growth. (D) The analysis of apoptosis by Hoechst-33342 staining for disintegrated nuclei in CNE2/LMP1 and CNE2/Vector cells treated with 3 μg/ml cisplatin for 72 h. Representative images (200x) at the indicated time points are shown. The white arrows indicate the cells with obviously broken nuclei. (E) The flow cytometry analysis of apoptosis by Annexin V/PI staining in CNE2/LMP1 and CNE2/Vector cells treated with 3 μg/ml cisplatin for indicated time points (left panel) or with cisplatin at various concentrations for 72h (right panel). The histograms shown are the means ± SD from three independent experiments (ns, not significant; * P < 0.05; ** P < 0.01 vs vector control cells). (F) C666-1 cells were transfected with siLMP1 or control siRNA (NC) for 24 h, exposed to 4 μg/ml cisplatin for 48 h or not and assayed to detect the apoptosis rate. The histograms shown are the means ± SD of three independent experiments (ns, not significant; ** P < 0.01 vs control siRNA transfected cells).