OXIDANT AND PROTEASE INTERACTION IN ACUTE LUNG INJURY

中性粒细胞和巨噬细胞来源的炎症介质 参与急性和慢性肺病的发病机制 人们对损伤的理解还不完全。大多数研究都认为 分别为蛋白酶或氧自由基(氧化剂) 参与其中。这些研究表明氧化剂是至关重要的 参与多种类型的急性肺损伤和蛋白水解酶， 尤其是弹性蛋白酶在慢性肺损伤中的作用。然而，新兴的 证据，特别是来自体外研究的证据表明 这两类强效炎性细胞之间的相互作用 介体可能在组织损伤的演变过程中起关键作用。 该提案将以系统的方式解决氧化剂和 蛋白水解酶的相互作用及其与致病机制的关系 急性肺损伤。在初步的体外研究中，我们将识别 大鼠中性粒细胞产生的蛋白酶和氧化剂的谱 和巨噬细胞，因为我们用大鼠进行体内研究 肺损伤的症状。我们有兴趣确定大鼠白细胞 具有与人类细胞相同类型的蛋白酶和氧化剂 在初步研究中，情况似乎就是这样。大鼠 中性粒细胞产生次氯酸(HOCL)，具有潜伏性 金属酶就像人类的中性粒细胞一样，产生 大量的弹性淀粉酶。在其他体外研究中，我们将 评估氧化剂-蛋白酶在破坏过程中的相互作用 细胞外基质蛋白。根据初步研究， 似乎氧化剂可以直接影响这些成分 增强后续的蛋白质分解，我们将详细研究如何 氧化剂和蛋白水解酶相互作用，促进这些物质的分解。 细胞外基质蛋白。 在相关的体内研究中，我们将解决这个问题 蛋白水解酶在两种急性肺损伤模型中的作用 似乎需要氧化剂才能启动组织的损伤 受伤。由于抗氧化剂的抑制作用似乎是时间 依赖于我们将做一个完整的时间进程研究来确定 确切地说，在损伤开始后多久，抗氧化剂 能有效抑制损伤。蛋白酶的证据 参与这两种急性肺损伤模型的人将 评估如下。首先，在肺灌洗中，我们将寻找 损伤的动物体内有游离酶活性的证据 初步研究表明他们确实存在。其次，我们 将寻找细胞外基质蛋白存在的证据 灌洗液，特别是它们是否有证据 蛋白水解酶。在初步研究中，我们发现这似乎 事实就是如此。在干预性研究中，我们将确定 抗氧化剂改变细胞外基质产物的水平，并 创伤大鼠灌洗液中的蛋白酶。最后，我们会 通过使用蛋白水解酶直接评估蛋白水解酶的作用 抑制剂。这项研究应提供有关 蛋白水解酶-氧化剂相互作用在糖尿病发病机制中的重要作用 急性肺损伤。

The inflammatory mediators derived from neutrophils and macrophages involved in the pathogenesis of acute as well as chronic lung injury are incompletely understood. Most studies have looked separately at either protease or oxygen radical (oxidant) involvement. These studies suggest that oxidants are critically involved in many types of acute lung injury and proteases, particularly elastase, in chronic lung injury. However, emerging evidence, particularly from in-vitro studies suggests that interactions between these two potent classes of inflammatory mediators may be critical in the evolution of the tissue injury. This proposal will address in a systematic fashion oxidant and protease interactions and how this relates to the pathogenesis of acute lung injury. In initial studies in-vitro we will identify the spectrum of proteases and oxidants produced by rat neutrophils and macrophages since we are using the rat for the in-vivo studies of lung injury. We are interested in determining if rat leukocytes have the same types of proteases and oxidants as the human cells and in preliminary studies this appears to be the case. Rat neutrophils produce hypochlorous acid (HOCL), have latent metalloenzymes just like the human neutrophil and produce significant amounts of elastase. In other in-vitro studies we will assess oxidant-protease interactions in the destruction of extracellular matrix proteins. Based on preliminary studies it appears as if oxidants can directly affect these components to enhance subsequent proteolysis and we will study in detail how oxidants and proteases interact to enhance the breakdown of these extracellular matrix proteins. In correlative in-vivo studies we will address the question of the role that proteases play in two experimental models of acute lung injury which appear to require oxidants to initiate the tissue injury. Since inhibition by antioxidants appears to be time dependent we will do a complete time course study to determine precisely how long after initiation of the injury that antioxidants can effectively suppress the injury. Evidence of protease involvement in these two models of acute lung injury will be assessed as follows. First, in lung lavage we will look for evidence of free protease activity in the injured animals with preliminary studies revealing that they are present. Secondly, we will be looking for evidence of extracellular matrix proteins in the lavage fluid and specifically whether or not they show evidence of proteolysis. In preliminary studies we find that this appears to be the case. In interventional studies we will determine if antioxidants alter the levels of extracellular matrix products and proteases in the lavage fluid of injured rats. Finally, we will assess directly the role that proteases play by the use of protease inhibitors. This study should provide important information on the importance of protease-oxidant interactions in the pathogenesis of acute lung injury.