ROLE OF CASPASES IN ISCHEMIC ACUTE RENAL FAILURE

描述(改编自申请者摘要)：急性肾缺血 衰竭(ARF)是一种常见的临床综合征，其发病率一直很高 死亡率。早期肾近端小管(PT)细胞极性丧失和 后来的肾小管细胞坏死被很好地描述为缺血性ARF。然而， 这些关键事件背后的机制尚未阐明。 基于初步数据和新发现的 半胱氨酸蛋白酶，半胱氨酸蛋白酶，作为其他脑缺血损伤的介质 器官，我们认为促炎性半胱氨酸蛋白酶-1，“启动者”半胱氨酸酶 8和9和“刽子手”caspase-3 8和9和“刽子手”caspase-3 在缺血性ARF期间被激活。这会导致细胞骨架的分解。 蛋白，它导致NAK-ATPase从 肾小管的基底外侧表面。在活体内，这些事件的意义 会减少PT对钠的重吸收，增加远端钠的输送 到致密黄斑，增加小管小球反馈，最后是 肾小球滤过率(GFR)降低。在第一个具体目标中，使用 Northern印迹分析、免疫印迹和特定的荧光底物， 存在于近端小管并在缺血期间被激活的caspase将被 已确认身份。在第二个具体目标中，caspase作为调解人的重要性 将在大鼠和小鼠的肾动脉夹模型中检查缺血性ARF的情况 使用特定的半胱氨酸酶抑制剂和基因敲除小鼠。肾功能和 形态学(肾脏组织学)的相关性将被记录在案。第三 特定目标将检测缺血时半胱氨酸酶的细胞靶点 利用体内缺血性ARF的补充研究，分离悬液中的PT 而且是在体外。具体地说，caspase与另一种 将对半胱氨酸蛋白酶进行研究。此外，脑缺血的影响与 如果没有半胱氨酸天冬氨酸氨基转移酶抑制剂，将使用免疫印迹和 免疫组织化学。这项资助中提出的总体假设提供了 一种综合的病理生理方案，根据该方案，肾PT损伤可导致 ARF的标志，即GFR的下降。这些研究与以下方面的相关性 临床ARF是实质性的，结果应该提供改变的线索 缺血性ARF的病程。

DESCRIPTION (Adapted from the Applicant's Abstract): Ischemic acute renal failure (ARF) is a common clinical syndrome which continues to have a high mortality rate. The early loss of renal proximal tubule (PT) cell polarity and the later tubular cell necrosis are well described is ischemic ARF. However, the mechanism underlying these pivotal events has not yet been elucidated. Based on preliminary data and established role of the newly discovered group of cysteine proteases, the caspases, as mediators of ischemic injury in other organs, we propose that the proinflammatory caspase-1, the "initiator" caspases 8 and 9 and the "executioner" caspase-3 8 and 9 and the "executioner" caspase-3 are activated during ischemic ARF. This causes breakdown of the cytoskeletal protein, spectrin, which leads to dissociation of NaK-ATPase from the basolateral surface of the tubule. In vivo, the significance of these events would be decreased PT reabsorption of sodium, increased distal sodium delivery to the macula densa, increased tubuloglomerular feedback and finally a decreased glomerular filtration rate (GFR). In the first specific aim, using northern blot analysis, immunoblotting and specific fluorogenic substrates, the caspases present in the proximal tubule and activated during ischemia will be identified. In the second specific aim, the importance of caspases as mediators of ischemic ARF will be examined in a renal artery clamp model in rats and mice using specific caspase inhibitors and knockout mice. Renal function and morphological (renal histology) correlates will be documented. The third specific aim will examine the cellular targets of caspases during ischemia using complementary studies in ischemic ARF in vivo, isolated PT in suspension and in vitro. Specifically, the interaction between caspases and the other cysteine protease, calpain, will be studied. Also the effect of ischemic with and without caspases inhibitors will be structured using immunoblotting and immunohistochemistry. The overall hypothesis presented in this grant provides an integrated pathophysiological schema whereby renal PT damage can lead to the hallmark of ARF, namely the fall in GFR. The relevance of these studies to clinical ARF is substantial and the results should provide leads to altering the course of ischemic ARF.