基因沉默缺氧诱导因子-1α对BxPC-3细胞株糖酵解基因表达的影响  

Effect of hypoxia inducible factor-1α gene silencing on glycolysis correlated gene expression inBxPC-3 cell line

作　　者：蒋奕[1] 吴国豪[1] 何国栋[1] 庄秋林[1] 张波[1] 韩寓嵩[1] 

机构地区：[1]复旦大学附属中山医院普外科,上海200032

出　　处：《中华实验外科杂志》2012年第11期2211-2214,共4页Chinese Journal of Experimental Surgery

摘　　要：目的观察沉默缺氧诱导因子（HIF）-1α基因表达对于缺氧微环境下人胰腺癌BxPC-3细胞株中糖酵解相关基因表达的影响。方法通过合成小干扰RNA（（siRNA））转染沉默BxPC-3细胞株中HIF-1α基因表达。将细胞分为空白对照组（BxPC-3）、空白质粒转染对照组（GFP）和HIF-1α基因沉默组（sh-HIF-1α），分别在正常环境（21．0％O2）和缺氧环境（0．5％-1．0％O2）中培养48h后，检测各组细胞中糖酵解相关基因[如丙酮酸激酶1（PDK-1）、乳酸脱氢酶A（LDH-A）和柠檬酸合成酶（CS）]的表达以及细胞糖酵解产物乳酸含量的改变。结果与正常环境比较，缺氧培养后sh-HIF-1α组细胞内HIF-1α mRNA增加了12．4％，蛋白增加了1．6倍，显著低于BxPC-3组和GFP组（P〈0．05），PDK-1和LDH-A的表达也明显低于两个对照组（P〈0．05）。相应的sh-HIF-1α组的乳酸含量为（4．8±0．3）nmol／10。个细胞，明显低于BxPC-3组（9．1±0．5）nmol／10^6个细胞和GFP组（9．2±0．5）nmol／10。个细胞（P〈0．05）。相反，缺氧时BxPC-3组和GFP组细胞中与线粒体氧化磷酸化相关的CS基因表达明显下降，低于其在sh-HIF-1α组细胞中的表达（P〈0．05）。结论缺氧可以诱导BxPC-3细胞株中HIF-hx基因高表达并促进糖酵解相关的PDK-1和LDH-A等基因表达。而通过沉默HIF-1α基因，可以抑制上述基因的表达，并促进CS基因表达，从而抑制肿瘤细胞的糖酵解代谢。Objective To investigate the impact of hypoxia inducible factor （HIF） -1αgene silen- cing on glycolysis correlated gene expression in human pancreatic cancer cell line （ BxPC-3 ） under hypoxic conditions. Methods The siRNA was synthesized to silence the expression of HIF-1α in BxPC-3 cell line. The transfected cells were divided into three groups ： blank control group （ BxPC-3 ）, empty plasmid vector transfection group （GFP） and HIF-1α gene silencing group （sh-HIF-1α）. All cells were cultured under either normoxic （21.0% O2 ） or hypoxic （0. 5%-1.0% 02 ） conditions for 48 h. Then pyruvate de- hydrogenase kinase 1 （PDK-1） , lactate dehydrogenase A （LDH-A）, and citrate synthase （CS） expression and the lactate acid level were analyzed respectively. Results Under hypoxie conditions, the expression of HIF-1α mRNA was increased by 12.4% and HIF-1α protein increased by 1.6 folds correspondingly, which were.significantly lower than in BxPC-3 and GFP groups （P 〈 0. 05 ）. PDK-1 and LDH-A expression levels were also lower in sh-HIF-1α group than the other two control groups. Accordingly, the final product of glycolysis, lactate acid level in sh-HIF-1α group [ （4. 8 ±0. 3） nmol/10^6 cells] was less than in BxPC-3 group [（9. 1 ±0.5） nmol/10^6 cells] and GFP group [（9.2 ±0.5） nmol/10^6 cells] （P〈0. 05）. In con- trast, the expression levels of CS mRNA and protein in BxPC-3 and GFP groups were repressed and lower than in sh-HIF-1α group （P 〈 0. 05）. Conclusion Hypoxia would induce HIF-1α expression in BxPC-3 cell line, which upregulated the glycolysis correlated PDK-1 and LDH-A expression. C, ene silencing of HIF-1α could inhibit PDK-1 and LDH-A, and promote the CS expression. As a result, the glycolysis pathway was greatly attenuated under hypoxic conditions.

关 键 词：缺氧诱导因子-1Α 糖酵解 丙酮酸激酶1 乳酸脱氢酶A 柠檬酸合成酶 

分 类 号：R730.2[医药卫生—肿瘤]