目的观察体外缺氧条件下肝癌细胞株HepG2中缺氧诱导因子-α(HIF-α)和甲硫氨酸腺苷转移酶2A(MAT2A)的表达,探讨HIF-α在低氧条件下对MAT2A基因表达的调控作用。方法构建人MAT2A启动子真核表达载体质粒,CoCl2化学模拟肿瘤缺氧环境,检测缺氧条件下MAT2A启动子活性,HIF-α和MAT2A在HepG细胞中的共定位、mRNA和蛋白水平的表达。以及小干扰RNA(siRNA)沉默HIF-α后对MAT2A基因表达的影响。结果缺氧24h能使HepG2细胞活性增加到高峰(41.26±2.34),同时MAT2A基因的表达也较常氧时显著升高(P〈0.01),siRNA转染HepG2细胞后能显著下调HIF-α蛋白表达(抑制率90%),并导致MAT2A基因的表达也受到明显抑制。结论缺氧促使HepG2细胞中HIF-α在蛋白水平表达升高,缺氧时HIF-α能上调MAT2A基因的表达水平。
Objective To observe the expression of hypoixa inducible factor-1α (HIF-α) and me- thionine adenosyltransferase-2A (MAT2A) gene in HepG2 cells under hypoxia in vitro, and explore the regulation of MAT2A gene expression by HIF-1α. Methods Human MAT2A promoter eukaryotic expres- sion vector was constructed. CoC12 was used as a chemical hypoxia-inducible reagent to mimic tumor hy- poxic microenvironment. MAT2A promoter activity, and mRNA and protein expression of HIF-αwere de- tected in the HepG2 cells transfected with RNA interference (RNAi) originated by small interfering RNA (siRNA). The change of MAT2A gene expression was observed after HIF-1α gene silencing. Results Under hypoxia for 24 h, HepG2 cell activity was increased to (41.26 ± 2. 34) , and the mRNA and protein expression levels of MAT2A were up-regulated (P 〈 0. 01 ). After siRNA targeting, the HIF-1 α was down- regulated efficiently in HepG2 cells ( inhibition ratio: 90% ), and MAT2A gene was down-regulated as well. Conclusion Hypoxia can increase protein level of HIF-1α in HepG2 ceils, and HIF-1α up-regu- lates the gene expression of MAT2A.