CARDIOCYTE RESPONSES TO HYPOXIA

描述（改编自申请人的摘要）：长期目标 本申请旨在定义心脏细胞的分子遗传反应， 氧化还原应激，分析介导这些反应的DNA调控元件， 为在缺血心肌中表达外源基因开辟了新的途径。 心肌缺血对心肌细胞产生两种主要形式的氧化还原应激 肌细胞：1）缺氧成分，由心肌细胞的严重程度决定。 缺血，和2）由再氧合引起的高氧应激。 两种形式的 压力会引发一系列复杂反应。 至少两种分子 可以区分对缺氧的遗传反应：快速的，立即的早期反应， 响应时间（几分钟到几小时）和延迟响应时间超过几个小时 天 原癌基因的诱导是早期反应的一部分， 糖酵解酶基因、骨骼α-肌动蛋白和内皮素的诱导 （ET-1）是晚期缺氧反应基因的实例。 无论是氧气 传感机制，也不是第二信使，中继氧化还原信号， 知道的 对氧化应激的反应必须迅速，因为 活性氧中间体是高度活性的，并立即氧化 细胞蛋白质和脂质，而不扩散远离其位点， 一代 研究提出了调查氧化还原调节 心肌细胞和血管内皮细胞中的特异性基因。 的 严重氧化还原应激对锌指转录因子的影响将是 表征了 糖酵解酶和ET-1基因的分子分析是 提出了确定缺氧负责元件（HRE）和蛋白质 把他们绑在一起 最后，研究提出了调查在体内 与HRE连接的报告基因的表达。 DNA将被输送到 心脏通过直接注射，心脏将经受缺血，并且 将报告基因表达与不含HRE的构建体的表达进行比较。

DESCRIPTION (adapted from the applicant's abstract): The long term goals of this application are to define molecular genetic responses of heart cells to redox stress, analyze DNA regulatory elements that mediate these responses, and define a new approach to express foreign genes in ischemic myocardium. Myocardial ischemia imposes two primary forms of redox stress on cardiac myocytes: 1) a hypoxic component determined by the severity of the ischemia, and 2) hyperoxic stress caused by reoxygenation. Both forms of stress trigger a complex series of reactions. At least two molecular genetic responses to hypoxia can be distinguished: a rapid, immediate early response (minutes to hours) and a late response that occurs over several days. Induction of proto-oncogenes is part of the early response, and induction of glycolytic enzyme genes, skeletal a-actin, and endothelin (ET-1) are examples of late hypoxia response genes. Neither the oxygen sensing mechanism nor the second messengers that relay the redox signals are known. The response to oxidative stress is obligatorily rapid since reactive oxygen intermediates are highly reactive and immediately oxidize cellular proteins and lipids without diffusing far from their sites of generation. Studies are proposed to investigate redox regulation of specific genes in cardiac myocytes and vascular endothelial cells. The impact of severe redox stress on zinc finger transcription factors will be characterized. Molecular analyses of glycolytic enzymes and ET-1 genes are proposed to identify hypoxia responsible elements (HREs) and the proteins that bind them. Finally, studies are proposed to investigate the in vivo expression of reporter genes linked to HREs. DNA will be delivered into the heart by direct injection, the heart will be subjected to ischemia, and the reporter expression will be compared with that from constructs without HREs.