PRECONDITIONING--PMN ADHESION AND MICROVASCULAR INJURY

我们实验室和其他实验室最近的研究表明，中性粒细胞 黏附于血管内皮细胞是产生微血管的必要条件 缺血后骨骼肌功能障碍和心肌细胞坏死。 对这一事实的认识导致了针对以下问题的重大研究努力 评价抑制白细胞黏附的潜力作为一种新的方法 再灌注损伤的治疗探讨。初步数据来自 我们的实验室表明，缺血预适应(IPC，一种现象) 使组织对有害物质的影响具有抵抗力 通过预先短暂、重复暴露延长缺血和再灌流时间 血管闭塞期间)通过以下方式预防I/R引起的肌肉坏死 抑制再灌流过程中白细胞的黏附和迁移。这个 本申请中概述的项目的总体目标是确定 IPC减轻白细胞黏附和黏附的机制 穿过毛细血管后静脉的迁移，微血管屏障的破坏， 骨骼肌毛细血管无复流和随后的心肌细胞坏死 暴露于长时间的缺血和再灌注(I/R)。我们的工作 假说是IPC将减轻微血管功能障碍和 缺血后骨骼肌肌细胞坏死的机制 在IPC期间和在 长时间缺血后再灌流。为了解决这个问题，我们建议 要确定：1)IPC是否会减弱白细胞黏附和 移行、毛细血管无血流、小静脉蛋白渗漏和心肌细胞 I/R引起的坏死；2)在脑缺血再灌注过程中产生的腺苷的作用。 预适应期内IPC的保护作用 在长时间缺血后的再灌流中表现明显；3)是否 IPC期间腺苷A/1受体激活的有益作用 周期通过一种涉及激活ATP敏感的机制发生 钾通道；4)IPC诱导的5‘- 大鼠脑持续缺血再灌流过程中核苷酸酶活性的变化 IPC的保护作用；5)腺苷的产生是否 在预适应的骨骼肌再灌流过程中增加 为IPC提供的保护行动做出了贡献 腺苷A/2受体。为了实现这些目标，我们将利用 活体内显微方法定量白细胞黏附和 小鼠的移行、小静脉蛋白渗漏和毛细血管无复流 提睾肌。IPC对I/I诱导的心肌细胞坏死的影响 还将接受检查。组织腺苷水平将通过高 高效液相色谱。拟议的研究不仅应该 大大提高我们对IPC的机制的理解 减少骨骼肌微血管功能障碍和心肌细胞坏死 受到随后长时间的缺血和再灌注期的影响，但 也应该为药物治疗提供一个理论基础 以I/R为特征的疾病

Recent studies from our laboratory and others indicate that neutrophil adherence to vascular endothelium is required to produce microvascular dysfunction and myocyte necrosis in postischemic skeletal muscle. Recognition of this fact has led to a major research effort directed at evaluating the potential for inhibition of leukocyte adhesion as a novel approach to the treatment of reperfusion injury. Preliminary data from our laboratory indicates that ischemic preconditioning (IPC, a phenomenon n which a tissue is rendered resistant to the deleterious effects of prolonged ischemia and reperfusion by prior exposure to brief, repeated periods of vascular occlusion) prevents muscle necrosis induced by I/R by inhibiting leukocyte adherence and emigration during reperfusion. The overall goal of the projects outlined in this application is to determine the mechanisms by which IPC attenuates leukocyte adhesion to and emigration across postcapillary venules, microvascular barrier disruption, capillary no-reflow, and myocyte necrosis in skeletal muscles subsequently exposed to prolonged ischemia and reperfusion (I/R). Our working hypothesis is that IPC will attenuate microvascular dysfunction and myocyte necrosis in postischemic skeletal muscles via a mechanism that involves adenosine receptor activation during the period of IPC and during reperfusion after prolonged ischemia. To address this issue, we propose to determine: 1) whether IPC will attenuate leukocyte adhesion and emigration, capillary no-flow, venular protein leakage, and myocyte necrosis induced by I/R; 2) the role of adenosine produced during the preconditioning period in the protective effects of IPC that become apparent during reperfusion after prolonged ischemia; 3) whether the beneficial effects of adenosine A/1-receptor activation during the IPC period occur by a mechanism that involves activation of ATP-sensitive potassium channels; 4) the role of IPC-induced increases in 5'- nucleotidase activity during reperfusion after sustained ischemia in the protective actions of IPC; and 5) whether adenosine production is increased during reperfusion of preconditioned skeletal muscles and contributed to the protective actions afforded by IPC by activating adenosine A/2-receptors. To accomplish these aims, we will utilize intravital microscopic approaches to quantitate leukocyte adhesion and emigration, venular protein leakage, and capillary no-reflow in the mouse cremaster muscle. The influence of IPC on I/-induced myocyte necrosis will also be examined. Tissue adenosine levels will be measured by high performance liquid chromatography. The proposed studies should not only substantially improve our understanding of the mechanisms whereby IPC reduces microvascular dysfunction and myocyte necrosis in skeletal muscles subjected to subsequent prolonged periods of ischemia and reperfusion but should also provide a rationale for the pharmacologic treatment of disorders characterized by I/R.