缺氧诱导因子-1在缺氧诱导一氧化氮合成酶基因表达中的作用

目的 研究缺氧诱导因子-1(HIF-1)在一氧化氮合成酶基因缺氧诱导反应中的作用。方法 体外合成具有HIF-1特异结合位点的DNA片段(红细胞生成素3'-增强子片段),借助脂质体,转入培养的鼠主动脉内皮细胞和肺微血管细胞,用半定量RT-PCR方法测定诱导型一氧化氮合成酶(iNOS)mRNA。结果 (1)大鼠主动脉内皮细胞、肺微血管内皮细胞在常氧下培养,有iNOS基因表达;(2)缺氧能诱导这两种细胞iNOS基因表达增加;(3)野生型EPO3'-增强子片段能阻断缺氧对内皮细胞iNOS基因表达的诱导作用,而突变片段则无此作用。结论 在iNOS基因序列中,可能存在EPO3'-端增强子片段,其参与内皮细胞的缺氧反应。

Effects of hypoxia-inducible factor 1 on the hypoxia induced expression of the nitric oxide synthase gene in the endothelial cells

Objective To investigate the effects of HIF-1 on the hypoxia induced expression of nitric oxide synthase gene in the endothelial cells. Method Wild and mutant EPO3'-enhancer DNA fragments were synthesized in vitro. Cultured endothelial cells were transfected with wild or mutant fragments. Total RNA was extracted from endothelial cells exposed to normoxia or hypoxia for 4 hours. The amount of iNOS mRNA was measured by semiquantitative RT-PCR. Results (1)There was expression of iNOS gene in the aortic endothelial cells and pulmonary vascular endothelial cells in normoxia. (2)hypoxia increased iNOS gene expression in these two kinds of endothelial cells. (3)The wild EPO3'-enhancer fragments inhibited the hypoxia-induced expression of iNOS gene, whereas the mutant fragments did not. Conclusion The increase of iNOS gene expression in hypoxia might result from the combination of activated hypoxia inducible factor 1 with EPO3'-enhancer-like sequence in iNOS gene.