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目的研究模拟低氧条件对舌鳞癌SCC-15细胞增殖、周期、凋亡的影响,及SOX2和OCT4蛋白、mRNA的表达变化。方法体外培养舌鳞癌SCC-15细胞,加入模拟低氧物去铁胺(desferrioxamine,DFO)模拟低氧环境,为模拟低氧组,同时设置不加DFO的常氧组为对照,采用CCK-8法检测SCC-15细胞的增殖率;流式细胞技术检测SCC-15细胞的周期和细胞凋亡率;蛋白质印迹法检测常氧组和模拟低氧组SCC-15细胞中低氧诱导因子1α(HIF-1α)、SOX2及OCT4蛋白的表达;qPCR检测常氧组和模拟低氧组SCC-15细胞中HIF-1α、SOX2及OCT4mRNA的表达。结果与常氧组相比,模拟低氧组SCC-15细胞的增殖受到抑制(P<0.05),G1期细胞所占比例上升、S+G2期细胞所占比例下降(P<0.05),细胞凋亡率未见明显变化(P>0.05);蛋白质印迹法结果显示,与常氧组比较,模拟低氧组SCC-15细胞中HIF-1α、SOX2、OCT4蛋白的表达均升高,且HIF-1α、OCT4蛋白的升高幅度大于SOX2蛋白,差异有统计学意义(P<0.05,P<0.01);qPCR结果显示,两组SCC-15细胞中HIF-1αmRNA水平差异无统计学意义(P>0.05),而模拟低氧组SCC-15细胞中SOX2、OCT4mRNA的表达水平高于常氧组(P<0.05)。结论 DFO可以有效模拟低氧环境,促进SOX2和OCT4的蛋白和mRNA表达水平升高。
Objective To investigate the effects of simulated hypoxia on proliferation, cycle and apoptosis of SCC-15 cells and the expression of SOX2 and OCT4 protein and mRNA. Methods SCC-15 cells were cultured in vitro. Simulated hypoxia environment was simulated by simulated hypoxia desferrioxamine (DFO). Simulated hypoxia group and normoxia group without DFO were used as control. CCK- 8 method was used to detect the proliferation rate of SCC-15 cells. The cell cycle and apoptosis rate of SCC-15 cells were detected by flow cytometry. The expressions of HIF-1α The expressions of HIF-1α, SOX2 and OCT4 protein were detected by qPCR. The expressions of HIF-1α, SOX2 and OCT4 mRNA in normoxia group and mock hypoxia group were detected by qPCR. Results Compared with normoxia group, the proliferation of SCC-15 cells in hypoxia group was inhibited (P <0.05), the proportion of cells in G1 phase increased, the proportion of cells in S + G2 phase decreased (P <0.05) Compared with normoxia group, the expression of HIF-1α, SOX2 and OCT4 protein in SCC-15 cells in simulated hypoxia group were increased, and the expression of HIF (P <0.05, P <0.01). The qPCR results showed that there was no significant difference in the level of HIF-1αmRNA between the two groups of SCC-15 cells (P <0.05, P <0.01) > 0.05). However, SOX2 and OCT4 mRNA expressions in SCC-15 cells in hypoxia group were higher than those in normoxia group (P <0.05). Conclusion DFO can effectively simulate hypoxic environment and promote the expression of SOX2 and OCT4 protein and mRNA levels.