siRNA沉默HIF-1α、HIF-2α、HIF-1β在不同缺氧状态下对BGC-823胃癌细胞VEGF表达的影响

发布时间：2018-05-12 11:11

本文选题：胃癌 + 乏氧诱导因子-1α　；参考：《新疆医科大学》2017年硕士论文

【摘要】：目的:观察不同缺氧条件下胃腺癌细胞系BGC-823中HIF-1α、HIF-2α、HIF-1β和VEGF的表达,探讨三种HIF因子在低氧条件下调控胃癌血管生成的作用差异。方法:分别用150、200、250、275μmol/L Co Cl2模拟肿瘤内部缺氧,缺氧时间分别为:2h、4h、8h、12h、24h、48h、72h。采用实时定量聚合酶链反应(Real-time PCR)和Western blot法检测HIF-1α、HIF-2α、HIF-1β和VEGF的mRNA及蛋白表达情况。进一步运用RNA干扰(RNA interference,RNAi)技术采用小干扰RNA(small interfering RNA,siRNA)沉默BGC-823细胞,比较三者沉默VEGF的效果。结果:1.与常氧时比较,缺氧后HIF-1α、HIF-2α、VEGF的m RNA及蛋白表达均随CoCl2浓度增加、时间延长而增加,但三者变化趋势不一致,HIF-1α在缺氧早期迅速升高,之后快速下降;HIF-2α和VEGF则在缺氧晚期表达最高,之后保持稳定。HIF-1βmRNA和蛋白在常氧或缺氧下表达均无明显变化(P0.05)。2.细胞分别特异性转染HIF-1α、HIF-2α和HIF-1βsiRNA后均能有效沉默各自基因的表达,同时VEGF的表达水平也随之降低,且随HIF-2α的下调幅度最显著(P0.05)。结论:1.BGC-823胃癌细胞中HIF-1α、HIF-2α和HIF-1β对乏氧刺激表现出不同的应答方式,HIF-1α和HIF-2α可能是胃癌适应乏氧环境的主要缺氧诱导因子;2.RNA干扰能抑制三种HIF基因的表达,并不同程度减少下游VEGF的表达,相较于HIF-1α和HIF-1β沉默途径,切断HIF-2α可能会更有效阻止胃癌血管生成。
[Abstract]:Objective: to observe the expression of HIF-1 伪 -HIF-2 伪 -hIF-1 尾 and VEGF in gastric adenocarcinoma cell line BGC-823 under different hypoxia conditions, and to explore the difference of three HIF factors in regulating angiogenesis of gastric cancer under hypoxia. Methods: 150200250275 渭 mol/L Co Cl2 was used to simulate the anoxia in the tumor. The anoxia time of the tumor was 12: 2, 4 h, 8 h, 12 h, 24 h, 48 h, 72 h, respectively. Real-time polymerase chain reaction (Real-time PCR) and Western blot were used to detect the expression of mRNA and protein in HIF-1 伪 -HIF-2 伪 HIF-1 尾 and VEGF. RNA interference siRNAs were further used to silence BGC-823 cells. The effect of VEGF silencing was compared among the three groups. The result is 1: 1. The expression of m RNA and protein of HIF-1 伪 -HIF-2 伪 -VEGF increased with the increase of CoCl2 concentration and time after hypoxia compared with that of normoxic hypoxia. However, the changes of HIF-1 伪 increased rapidly at the early stage of hypoxia, and then decreased rapidly. The expression of HIF-2 伪 and VEGF was the highest in the late stage of hypoxia. After that, the expression of. HIF-1 尾 mRNA and protein remained stable under normoxic or hypoxic conditions. There was no significant change in the expression of HIF-1 尾 mRNA and protein under normoxic or hypoxic conditions. The specific transfection of HIF-1 伪 -HIF-2 伪 and HIF-1 尾 siRNA could effectively silence the expression of their respective genes, and the level of VEGF expression also decreased, and the down-regulation of HIF-2 伪 was the most significant (P 0.05). Conclusion HIF-1 伪 HIF-2 伪 and HIF-1 尾 have different responses to hypoxia stimulation in BGC-823 gastric cancer cells. HIF-1 伪 and HIF-2 伪 may be the main hypoxia inducible factors for gastric cancer to adapt to hypoxia. 2. RNA interference can inhibit the expression of three HIF genes. Compared with HIF-1 伪 and HIF-1 尾 silencing pathway, HIF-2 伪 may be more effective in preventing angiogenesis in gastric cancer than in HIF-1 伪 and HIF-1 尾 silencing pathway.
【学位授予单位】：新疆医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.2

【参考文献】