Delayed disappearance of h-VEGF165 mRNA and protein under regulation of hypoxic response element
- Author:
- No author available
- Journal Title:
- ACTA PHYSIOLOGICA SINICA
- Issue:
- 3
- DOI:
- 10.3321/j.issn:0371-0874.2006.03.013
- Key Word:
- 血管内皮生长因子;低氧反应元件;心肌细胞
Abstract： 血管内皮生长因子(vascular endothelial growth factor,VEGF)转基因可促进心肌缺血区血管生成,改善心脏功能,然而长期高水平表达又会引起诸多副作用.为调控VEGF表达,在启动子区加入低氧反应元件(hypoxic response element,HRE)作为调控开关,研究氧环境对VEGF基因mRNA及蛋白产物表达的影响.重组腺相关病毒(recombinant adeno-associ-ated virus,rAAV)作为载体(rAAV-HRE-h-VEGF165),转染离体培养大鼠心肌细胞,在常氧/缺氧(氧浓度1%)/缺氧复氧条件下进行培养,用酶联免疫特异性测定(enzymelinkedimmunosorbent assay,ELISA)方法检测培养液h-VEGF165蛋白浓度,细胞免疫荧光染色观测细胞内h-VEGF165蛋白表达,逆转录聚合酶链反应(reverse transcription polymerase chain reaction,RT-PCR)方法检测细胞h-VEGF165 mRNA表达.结果显示:未转染组、常氧培养组均无h-VEGF165mRNA及其蛋白表达;缺氧培养组h-VEGF165mRNA及蛋白均表达;缺氧复氧4 h组的h-VEGF165mRNA消失,培养液中h-VEGF165蛋白减少,细胞内h-VEGF165蛋白存在;缺氧复氧8 h及12 h组的h-VEGF165mRNA消失,培养液和细胞内h-VEGF165蛋白均消失.研究表明,在HRE调控下,缺氧可促使h-VEGF165基因表达,复氧后,h-VEGF165mRNA表达停止,蛋白产物延迟消失.
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