NEUTROPHILS IN ISCHEMIA REPERFUSION INJURY IN SHOCK

休克缺血再灌注损伤中的中性粒细胞

• 批准号: 2444729
• 负责人: ROBERT K WINN
• 金额: $ 17.47万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-04-01 至 1999-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2444729
• 关键词: CD antigens; Primates; body temperature; cardiopulmonary resuscitation; enzyme linked immunosorbent assay; gene expression; hemorrhagic shock; human subject; hypothermia; immunocytochemistry; in situ hybridization; ischemia; laboratory rabbit; leukocyte activation /transformation; leukocyte adhesion molecules; messenger RNA; monoclonal antibody; neutrophil; northern blottings; phagocytosis; phorbols; reperfusion; selectins; tissue /cell culture; vascular endothelium; CD antigens; Primates; body temperature; cardiopulmonary resuscitation; enzyme linked immunosorbent assay; gene expression; hemorrhagic shock; human subject; hypothermia; immunocytochemistry; in situ hybridization; ischemia; laboratory rabbit; leukocyte activation /transformation; leukocyte adhesion molecules; messenger RNA; monoclonal antibody; neutrophil; northern blottings; phagocytosis; phorbols; reperfusion; selectins; tissue /cell culture; vascular endothelium

A significant portion of the injury that occurs following ischemia and reperfusion has been established to occur during the reperfusion period. This reperfusion injury is leukocyte mediated and blocking leukocyte adherence with monoclonal antibodies provides significant protection from injury. Likewise, mild hypotheremia (1-4 degrees C) during ischemia and reperfusion results in a significant reduction in injury. Ischemia-reperfusion is seen in a variety of clinical disorders including myocardial infarction, stroke, organ transplantation, hemorrhagic shock and traumatic injuries. Clearly, this broad spectrum of clinical disorders that may have a component of reperfusion injury places it as a major cause of mortality and morbidity in the United States. Prolonged ischemia in these disorders results in significant injury due to tissue hypoxia and rapid return of perfusion will reduce this type of injury. However, reperfusion of previously ischemic tissue can initiate an inflammatory response that results in significant additional tissue injury. Blocking leukocytes adhesion has been shown experimentally to significantly reduce portion of this reperfusion injury. Likewise, mild hypothermia has been shown to reduce the injury that occurs in ischemia and reperfusion. The hypotheses of this project are: 1) The expression and/or activation of leukocyte adhesion molecules on endothelial cells or on neutrophils (pMNs) is dependent on small changes in temperature (1 to 4 degrees C). Adhesion molecule function is reduced during mild hypothermia, thus providing protection from ischemia and reperfusion injury. and 2) Significant tissue protection results from mild hypothermia during the immediate post reperfusion period (4-8 hours). Therefore, maintaining patients slightly hypothermic during this period may provide protection from injury and be of significant therapeutic benefit. The specific aims are: Aim 1 A: To determine the stimulated expression of adhesion molecules CD62P, CD62E, CD106, and CD54 as well as expression of mRNA by cultured HUVECs using ELISA and Northern blot analysis at 33, 35, 37, and 39 degrees C. Aim 1 B: By understanding the mechanism of hypothermia-mediated inhibition of gene expression in endothelial cells, it will be possible to reverse or activate the effects of hypothermia in vivo - Aim 2: To measure indicators of PMN function and activation following stimulation of leukocytes with PMA, heat killed bacteria, and C5a. Temperature will be set at 33 degrees C, 35 degrees C, 37 degrees C and 39 degrees C. Aim 3: To determine the effect of mild hypothermia on leukocyte rolling and adherence under flow conditions at temperatures of 33, 35, 37, and 39 degrees C. Aim 4: To determine the quantitative changes in expression of CD62P, CD62E and CD54 and their mRNA in rabbit ear ischemia- reperfusion at ambient temperatures of 22 degrees C and 24 degrees C. Aim 5: To determine the timing of hypothermia that is effective in preventing tissue injury following ischemia and reperfusion. Aim 6: To determine the protective effects of reduced temperature following hemorrhagic shock and resuscitation. The experiments of this project will use cell and molecular biology techniques and will be completed by using in vivo and in vitro experiments.

在缺血和 在再灌注期间已经建立了再灌注。 这种再灌注损伤是白细胞介导的，堵塞白细胞 遵守单克隆抗体可提供明显的保护 受伤。 同样，温和的性交（1-4度C） 缺血和再灌注会显着减少损伤。 在多种临床疾病中可以看到缺血 - 重灌注 包括心肌梗塞，中风，器官移植， 出血性休克和创伤性伤害。显然，这个广阔的范围 可能具有再灌注损伤成分的临床疾病 将其视为统一死亡率和发病率的主要原因 国家。这些疾病的长期缺血导致了重要的 由于组织缺氧引起的伤害和灌注迅速恢复将减少 这种类型的伤害。但是，先前缺血组织的再灌注 可以发起炎症反应，从而导致重要 额外的组织损伤。已显示阻塞白细胞粘附 实验可显着减少此再灌注的一部分 受伤。同样，温和的体温过低可以减少损伤 这发生在缺血和再灌注中。该项目的假设 是：1）白细胞粘附分子的表达和/或激活 在内皮细胞或中性粒细胞上（PMN）取决于小 温度变化（1至4度C）。粘附分子功能 在温和体温过低期间减少 缺血和再灌注损伤。 2）明显的组织保护 在直接再灌注期间轻度体温过低的结果 时期（4-8小时）。因此，维持患者略有低温 在此期间，可能会提供免受伤害的保护 显着的治疗益处。具体目的是：目标1 A： 确定粘附分子CD62P的刺激表达，CD62E， CD106和CD54以及使用培养的HUVEC表达mRNA的表达 Elisa和Northern印迹分析，分别为33、35、37和39度。 1 B：通过了解低温介导的抑制的机制 内皮细胞中基因表达的表达，可以反向 或激活体内体温过低的影响 - 目标2：测量 刺激后PMN功能和激活的指标 白细胞带有PMA，热量杀死细菌和C5A。温度将是 设置为33度C，35度C，37度C和39度。 3：确定温和体温过低对白细胞滚动的影响 在33、35、37和39的温度下流动条件下的依从性 学位C.目标4：确定表达的定量变化 CD62P，CD62E和CD54及其在兔耳朵缺血中的mRNA- 在22度和24度的环境温度下再灌注。 目标5：确定有效的体温过低的时间 缺血和再灌注后防止组织损伤。目标6：到 确定降低温度的保护作用 出血性休克和复苏。该项目的实验 将使用细胞和分子生物学技术，并将通过 使用体内和体外实验。