MECHANISMS OF LIPID A TOXICITY IN RENAL PROXIMAL TUBULES

描述：（改编自研究者摘要）急性肾衰竭 是严重细菌感染常见且往往致命的后果， 人类 研究人员先前的研究表明， 一氧化氮合酶（NOS）负责细胞毒性 细菌内毒素（脂多糖）在肾近端的作用 小管。 脂质A，脂多糖的生物活性部分， 通过释放细胞内Ca 2+，其反过来激活NOS，导致NO 一代 虽然在几分钟内检测到NO生成， 2+更晚。 然而，细胞毒性可以通过以下方式预防：（a） 抑制Ca 2+释放，（B）NOS抑制剂，或（c）抗氧化剂。 的 拟议的研究将解决这一假设，即NOS的早期激活， 和随后的氧化应激是脂质A的主要决定因素 毒性 在具体目标1和2中，调查人员将审查 由脂质A和NO触发的生化事件，最终导致 细胞毒 在具体目标3中，他们将研究NOS的作用 内毒素诱导的急性肺损伤模型中的活化和氧化剂产生 肾衰竭 为实现上述所有目标， 建议; 1。他们将研究Ca 2+在激活 使用两种互补方法的近端小管NOS。 一是 使用部分纯化的制剂进行酶表征。 的 第二种方法利用用脂质A刺激的完整的分离的小管。 2.）的情况。研究人员将研究NO生成在 脂质A的细胞毒性。PI将评价NOS的有效性 抑制剂、NO清除剂和氧化剂清除剂以及Ca 2+释放 抑制剂对亚致死至致死损伤的发展。 3.）第三章的PI 将研究NO在体内内毒素诱导的肾衰竭中的作用。 他们将使用内毒素诱导的肾衰竭模型， NO和氧化剂在脂A和脂多糖诱导的急性肾损伤中的作用 在与体外试验相似的时间过程中发生的失败 问题研究 了解NOS活性的调节和 NO生成的后果对于理解 一氧化氮在近曲小管病理生理中的作用及新疗法的开发 来治疗人类的疾病

DESCRIPTION: (Adapted from the Investigator's Abstract) Acute renal failure is a frequent and often fatal consequence of severe bacterial infection in humans. The investigators's previous studies indicate that activation of the enzyme nitric oxide synthase (NOS) is responsible for the cytotoxic effects of bacterial endotoxin (lipopolysaccharide) in the kidney proximal tubule. Lipid A, the biologically active moiety of lipopolysaccharide, acts by releasing intracellular Ca2+ which in turn activates NOS resulting in NO generation. Although NO generation is detected within minutes, cell death occurs much 2+ later. Nevertheless, cytotoxicity can be prevented by (a) inhibiting Ca2+ release, (b) inhibitors of NOS, or (c) antioxidants. The proposed studies will address the hypothesis that early activation of NOS and subsequent oxidant stress are the major determinants of lipid A toxicity. In specific aims 1 and 2, the investigators will examine the biochemical events triggered by lipid A and NO that ultimately lead to cytotoxicity. In specific aim 3 they will examine the role of NOS activation and oxidant generation in a model of endotoxin-induced acute renal failure. To accomplish all of the above, the following specific aims are proposed; 1.) they will examine the role of Ca2+ in the activation of proximal tubule NOS using two complimentary approaches. The first is enzymatic characterization using a partially purified preparation. The second approach utilizes intact isolated tubules stimulated with lipid A. 2.) the investigators will examine the role of NO generation in the cytotoxicity of lipid A. The PI will evaluate the effectiveness of NOS inhibitors, NO scavengers and oxidant scavengers, and Ca 2+ release inhibitors on the development of sublethal to lethal injury. 3.) The PI will examine the role of NO in endotoxin-induced renal failure in vivo. Using a model of endotoxin-induced renal failure, they will examine the role of NO and oxidants in lipid A and lipopolysaccharide-induced acute renal failure that develop over a similar time course as in their in vitro studies. An understanding of the regulation of NOS activity and the consequence of NO generation is crucial to the understanding of the role of NO in proximal tubule pathophysiology and the development of new therapies to treat the human disease.