LIVER/LUNG INTERACTIONS DURING GRAM NEGATIVE ENDOTOXEMIA

描述(改编自申请人的摘要)：在革兰氏阴性期间 细菌性败血症，一种肝-肺炎症轴，易患危重 炎症与抗炎因子表达失衡所致的器官损伤 炎性基因产物。这种细菌感染后的器官功能障碍被认为是 因还原-氧化引起的继发性缺血-缺氧应激 (氧化还原)敏感的转录因子和随后的扩增 关键细胞因子和非细胞因子表达介导的炎症反应 基因。这项拟议的研究的目的是检验这一假设 肝和肺的细菌后氧限制不同 调节一组确定的氧化还原敏感转录的激活 因子，从而改变细胞因子的表达方向相反和 器官特有的方式。实验的设计是为了确定 肝脏与肺供氧在调节中的继发性减少 核因子-kB(核因子-kB)激活蛋白的菌后反式激活 (AP)-1、NFIL-6和cAMP反应元件结合蛋白(CREB)。这个 这些变化的生物学意义将通过研究 伴随的炎性细胞因子(肿瘤坏死因子-α、白介素1-α、 IL-1β)、抗炎细胞因子(IL-6、IL-10)、前列腺素(PG)H 合酶-2和诱导型一氧化氮合酶在这些器官中的表达 在异位器官灌流期间与氧化还原状态有关的系统。实验 也被设计来定义细菌感染后蛋白：DNA相互作用 低氧应激和复氧通过评估这些的激活 库普弗细胞和肺泡巨噬细胞中的转录因子。结果数据 将确定是否存在涉及增强的COX-2和CREB的自动调节循环 PGE2依赖的活性抑制细菌后细胞因子的表达 机制以特定于器官的方式进行。核子激活的作用 蛋白缺氧诱导因子-1在内毒素诱导细胞因子中的协同调节作用 随后的低氧过程中的基因表达将被评估。这些终端 也将在有和没有低氧应激的清醒大鼠中进行分析 细菌性感染后先前存在的肝功能障碍。并行的 研究表明，顺式作用的DNA序列赋予缺氧抑制力 内毒素诱导RAW 264.7细胞的细胞因子启动子活性 利用肿瘤坏死因子-α和白介素1-β报告基因构建物将被识别。 这些垂直方向的研究结果应该会提供新的见解 细胞因子和诱导型一氧化氮合酶表达的转录调控 革兰氏阴性细菌性败血症的机制研究 改善肺损伤和多器官功能衰竭。

DESCRIPTION (Adapted from the applicant's abstract): During gram-negative bacteremic sepsis, a liver-lung axis of inflammation predisposes to critical organ injury due to an imbalanced expression of inflammatory vs. anti- inflammatory gene products. Such postbacteremic organ dysfunction is thought to be augmented by secondary ischemic-hypoxic stress owing to reduction-oxidation (redox)-sensitive transcription factors and subsequent amplification of inflammatory responses mediated by expression of key cytokine and noncytokine genes. The objective of the proposed research is to test the hypothesis that postbacteremic O2 limitation within the liver and the lungs differentially modulates the activation of a defined group of redox-sensitive transcription factors, thereby altering cytokine expression in a directionally- opposite and organ-specific manner. Experiments are designed to determine the effects of secondary reductions in the hepatic vs. pulmonary O2 supply in modulating postbacteremic transactivation of nuclear factor-kB (NF-kB), activator protein (AP)-1, NFIL-6, and the cyclic AMP response element binding protein (CREB). The biologic significance of these changes will be assessed by examining the concomitant expression of inflammatory cytokines (TNF-alpha, IL-1alpha, IL-1beta), anti-inflammatory cytokines (IL-6, IL-10), prostaglandin (PG) H synthase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in these organ systems in relation to redox status during ex situ organ perfusion. Experiments are also designed to define postbacteremic protein:DNA interactions during hypoxic stress and reoxygenation by assessing the activation of these transcription factors in Kupffer cells and alveolar macrophages. Resulting data will establish if an autoregulatory loop involving enhanced COX-2 and CREB activity suppresses postbacteremic cytokine expression by a PGE2-dependent mechanism in an organ-specific manner. The role of activation of the nuclear protein hypoxia inducible factor-1 in co-modulating endotoxin-induced-cytokine gene expression during subsequent hypoxia will be assessed. These endpoints will also be analyzed in conscious rats during hypoxic stress, with and without preexisting liver dysfunction following bacteremic infection. In parallel studies, the cis-acting DNA sequences that confer hypoxic suppressibility of endotoxin-induced cytokine promoter activity in RAW 264.7 cells transfected with TNF-alpha and IL-1beta reporter gene constructs will be identified. Results from these vertically-oriented studies should provide novel insights into the transcriptional regulation of cytokine and iNOS expression during gram-negative bacteremic sepsis while identifying mechanistic approaches to ameliorate lung injury and multiple organ failure.