HEME OXYGENASE-1 GENE EXPRESSION IN ENDOTOXIC SHOCK

内毒素休克是严重克- 以血管平滑肌细胞为特征的阴性感染 松弛和严重低血压。 一氧化氮（NO），通过 诱导型一氧化氮合酶（iNOS）途径，是一个主要的贡献者，在这一点上， 过程 然而，我们实验室最近的调查表明， 血红素加氧酶（HO），一种产生一氧化碳（CO）的酶， 在血红素代谢过程中，也可能参与 内毒素休克血流动力学损害 诱导型HO（HO-1）是 在接受内毒素的大鼠的血管组织中显著增加， HO活性的非选择性抑制剂消除了内毒素诱导的 低血压此外，HO-1 mRNA的诱导在 基因转录 除了转录因子与 它们的DNA结合结构域，适当的染色质环境是 对基因转录很重要。 通过改变染色质结构， 结构蛋白将转录因子组装成增强子 复杂. 一种这样的结构蛋白，高迁移率蛋白 （HMG）-I（Y），已显示可促进 转录因子转化为复合物， 协同地增加基因转录。 我们的初步数据显示， 实验表明，与HO-1一样，HMG-I（Y）mRNA和蛋白在 血管平滑肌细胞的IL-1 β和内毒素的作用。 vivo. 此外，HMG-I（Y）结合HO-1的5'侧翼序列， 基因 在本申请中，我们提出测试HO-1在以下方面的作用： 内毒素休克，并研究调节内毒素休克的分子机制。 血管平滑肌细胞中HO-1的转录诱导。 提出了三个具体目标。 目的1：确定HO-1的作用 内毒素诱导的低血压和死亡， HO-1基因 我们假设HO-1基因的缺失将导致 预防体内内毒素诱导的低血压和死亡。 目标2： 确定特定的DNA序列（顺式作用元件）， 对血管平滑肌细胞中HO-1基因诱导很重要， 探讨染色质在HO-1基因调控中的作用 转录。 我们假设顺式作用蛋白 具有反式作用因子的元素，在综合环境中， HO-1基因转录。 目标3：识别和描述 这些DNA结合蛋白（反式作用因子）， 与HMG-I（Y）的相互作用对HO-1基因的诱导起重要的调节作用 通过血管平滑肌细胞中IL-1 β的转录。 这些 研究应该为HO-1的调节提供新的见解， 内毒素休克的发病机制。

Endotoxic shock is a life-threatening consequence of severe gram- negative infection characterized by vascular smooth muscle cell relaxation and severe hypotension. Nitric oxide (NO), through the inducible NO synthase (iNOS) pathway, is a major contributor in this process. However, recent investigations from our laboratory suggest that heme oxygenase (HO), an enzyme that generates carbon monoxide (CO) in the course of heme metabolism, may also be involved in the hemodynamic compromise of endotoxic shock. Inducible HO (HO-1) is dramatically increased in vascular tissue from rats receiving endotoxin, and non-selective inhibitors of HO activity abrogate endotoxin-induced hypotension. In addition, the induction of HO-1 mRNA is at the level of gene transcription. Beyond an interaction of transcription factors with their DNA binding domains, an appropriate chromatin environment is important for gene transcription. By changing chromatin structure, architectural proteins assemble transcription factors into an enhancer complex. One such architectural protein, high mobility groups protein (HMG)-I(Y), has been shown to promote cooperative binding of transcription factors into a complex that efficiently and synergistically increases gene transcription. Preliminary data from our laboratory show that like HO-1, HMG-I(Y) mRNA and protein is induced in vascular smooth muscle cells by IL-1 beta in vitro and by endotoxin in vivo. Furthermore, HMG-I(Y) binds to 5' flanking sequence of the HO-1 gene. In this application we propose to test the role of HO-1 in endotoxic shock, and to study the molecular mechanisms regulating the transcriptional induction of HO-1 in vascular smooth muscle cells. Three specific aims are proposed. Aim 1: To determine the role of HO-1 in endotoxin-induced hypotension and death by generating mice deficient in the HO-1 gene. We hypothesize that deletion of the HO-1 gene will prevent endotoxin-induced hypotension and death in vivo. Aim 2: To identify the specific DNA sequences (cis-acting elements) that are important for HO-1 gene induction in vascular smooth muscle cells, and to determine the role of chromatin in regulating HO-1 gene transcription. We hypothesize that the interaction of cis-acting elements with trans-acting factors, in an integrated setting, mediates HO-1 gene transcription. Finally, Aim 3: To identify and characterize those DNA-binding proteins (trans-acting factors), that in conjunction with HMG-I(Y), are important for regulating the induction of HO-1 gene transcription by IL-1 beta in vascular smooth muscle cells. These studies should provide new insight into the regulation of HO-1, and the pathogenesis of endotoxic shock.