摘要
目的探讨低氧反应元件(hypoxic response element,HRE)对缺血、复灌心肌转导人血管内皮生长因子165(human vascular endous growth factor 165,hVEGF165)基因表达的调控作用。方法在猪冠状动脉回旋支起始处以钛夹阻断回旋支血流,建立可复灌的心肌缺血模型。携带9拷贝HRE(9HRE)-hVEGF165的腺相关病毒转染模型心肌,转染后4天,复灌组二次开胸打开钛夹恢复缺血区血供。取各组心肌组织,RT-PCR法测定hVEGF165mRNA的表达情况;免疫组化及免疫印迹法测定hVEGF165蛋白的表达情况;Ⅷ因子免疫组化染色检测局部毛细血管情况。结果缺血心肌转导hVEGF165基因后,hVEGF165蛋白表达量明显增高,而复灌组表达明显减少(P<0.01)。结论9HRE作为调控缺血心肌组织内转导的hVEGF165基因的氧敏感开关灵敏、高效,使hVEGF165基因在缺血时高度表达,而复灌后表达水平下降。
Objective To investigate how the nine copies hypoxic response elements （9HRE） regulate the expression of human vascular endothelial growth factor 165 （hVEGF165） gene transferred by adeno- associated vector into ischemia and reperfusion myocardium in vivo. Methods 24 male swine ischemic myocardium models were made through left lateral fourth intercostal thoracotomy by placing a titanium clamp on the proximal left circumflex artery. Recombinant adeno - associated viral （rAAV - 9HRE - hVEGF165 ） vectors or PBS was injected into the ischemic myocardium. The titanium clamp in reperfusion group was released under thoracotomy 4 days after gene transfer. 8 days after gene transfer, Ⅷ factor staining was performed, hVEGF165 protein was tested by Western blot and i mmunochemistry, hVEGF165 mRNA was detected after reverse transcription polymerase chain reaction （RT- PCR）. Results The expression of hVEGF165 increased in ischemic myocardium significantly and decreased rapidly after reperfusion. Conclusion The expression level of transferred hVEGF165 gene rises in ischemia and declines under reperfusion. 9HRE could regulate the gene expression in response to the oxygen environment.
出处
《徐州医学院学报》
CAS
2006年第5期381-385,共5页
Acta Academiae Medicinae Xuzhou
基金
江苏省教育厅重点资助项目(03JKA310140)
关键词
低氧反应元件
基因调控
人血管内皮生长因子165
腺相关病毒
心肌
hypoxic response elements
gene regulation
human vascular endothelial growth factor 165
recombinant adeno- associated virus
myocardium
