Understanding the Function of Tamm-Horsfall Protein in Acute Kidney Injury

描述（由申请人提供）： 急性肾损伤（阿基）是住院退伍军人死亡率和发病率的主要决定因素。了解阿基的病理生理学对于开发治疗至关重要。本申请提出研究Tamm-Horsfall蛋白（THP）在阿基中的作用。这种蛋白质的功能尚不清楚，仅在肾脏的粗升支（TAL）中表达。本研究的直接目标是描述THP在肾损伤和炎症中的新保护作用。PI的长期目标是了解THP在阿基期间的作用如何调节，以达到预防和治疗目的。中心假设是Tamm-Horsfall蛋白调节由外髓质缺血性损伤引发的炎症反应;这种作用需要THP与周围肾小管元件相互作用。该假设是基于PI产生的强有力的初步数据制定的，这些数据表明THP在阿基中具有保护作用，并且与S3节段的炎症和损伤较少相关，S3节段是肾脏外髓质中TAL的相邻小管。以下具体目标将用于调查这一假设。目的1将定义THP和近端小管在缺血性损伤中的相互作用，从而支持THP-S3直接相互作用。这一目标将确定THP从TAL释放的基底外侧路径及其与S3节段上受体相互作用的意义。目标2将确定THP调节的肾损伤中的特定炎症通路。目的是确定THP的保护作用部分是通过抑制特定的炎症趋化因子途径。这项研究将涉及使用THP敲除和野生型小鼠。用于阿基的动物模型是通过肾蒂钳夹实现的肾缺血-再灌注损伤。体外研究将涉及近端小管和粗升支的细胞系。其他技术包括：免疫荧光共聚焦显微镜、免疫金电子显微镜、流式细胞术、真实的时间PCR、沉默RNA、蛋白质印迹、激光捕获显微切割和原位杂交。这项研究是创新的，因为它将揭示THP的新的调节功能，这些功能不仅可能与肾损伤有关，而且可能与其他形式的急性或慢性肾脏疾病有关。加强THP在肾脏中的作用的未来策略将具有重要的治疗意义。 公共卫生相关性： 肾功能恶化或急性肾衰竭是住院退伍军人发病率和死亡率的重要因素。不幸的是，除了透析的支持性治疗外，目前还没有确定的治疗方法。我们拟议的研究将研究一种蛋白质Tamm-Horsfall蛋白（THP）的作用，这种蛋白质可以在损伤的情况下保护肾脏。因此，该项目与退伍军人健康有着重要的直接关系。更好地了解THP的作用，为未来的治疗策略提供了希望，通过增强这种蛋白质的保护作用来改善肾衰竭的结局。

DESCRIPTION (provided by applicant): Acute kidney injury (AKI) is a major determinant of mortality and morbidity in hospitalized veterans. Understanding the pathophysiology of AKI is essential for the development of therapy. This application proposes to study the role of Tamm-Horsfall protein (THP) in AKI. This protein has unclear functions and is expressed exclusively in thick ascending limbs (TAL) of the kidney. The immediate goal of this research is to characterize a novel protective role of THP in kidney injury and inflammation. The PI's long term goal is to understand how the role of THP can be modulated during AKI for preventative and therapeutic purposes. The central hypothesis is that Tamm-Horsfall protein regulates the inflammatory response triggered by ischemic injury in the outer medulla; an effect that requires an interaction of THP with the surrounding tubular elements. This hypothesis has been formulated on the basis of strong preliminary data produced by the PI showing that THP is protective in AKI and is associated with less inflammation and injury to S3 segments, which are neighboring tubules to TAL in the kidney outer medulla. The following specific aims will be used to investigate this hypothesis. Aim 1 will define the interaction between THP and proximal tubules in ischemic injury, thereby supporting a direct THP-S3 interaction. This aim will define the basolateral path for THP release from TAL and the significance of its interaction with receptors on S3 segments Aim 2 will identify specific inflammatory pathways in kidney injury that are modulated by THP. The goal is to establish that the protective effect of THP is in part through the inhibition of a specific inflammatory chemokine pathway. This research will involve the use of THP knockout and wild type mice. The animal model used for AKI is renal ischemia-reperfusion injury achieved through renal pedicel clamping. In vitro studies will involve cell lines for proximal tubules and thick ascending limbs. Other techniques used in this work include: Immunofluoresence confocal microscopy, immuno-gold electron microscopy, flow cytometry, real time-PCR, silencing RNA, western blot, laser capture microdissection and in-situ hybridization. This research is innovative, because it will uncover novel regulatory functions for THP that may be relevant not only in kidney injury but also in other forms of acute or chronic renal disease. Future strategies that enhance the role of THP in the kidney will be of important therapeutic significance. PUBLIC HEALTH RELEVANCE: Deterioration of kidney function, or acute kidney failure is an important contributing factor to the morbidity and mortality of hospitalized veterans. Unfortunately, there is no established treatment to date for this disease except supportive care with dialysis. Our proposed research will study the role of a protein, Tamm-Horsfall Protein (THP), that may protect the kidneys in the setting of injury. Therefore, this project has improtant direct relevance to veterans health. A better understanding of the role of THP holds promise for future therapeutic stategies that improve outcomes of kidney failure by enhancing the protective role of this protein.