The Cytoskeletal System in Preservation Injury

描述(由申请人提供)：肾移植是终末期肾功能衰竭患者唯一可行的治疗方法。然而，严重的器官短缺让美国数以千计的人在等待名单上遭受痛苦和死亡。如果这些移植物能够得到妥善保存，扩大捐赠者池以包括高风险捐赠者可能会解决这一短缺问题。然而，器官采集后所遭受的器官保存损伤还不是很清楚，在过去的20年里，这一领域几乎没有取得重大进展。因此，这项建议的目的是确定涉及细胞骨架系统的肾脏保存损伤的新的分子机制，并利用这一知识指导未来的治疗方法。初步数据显示，细胞骨架蛋白Ezrin在肾脏冷藏过程中失效，增强Ezrin的表达显著保护肾小管上皮细胞免受保存损伤。相反，Ezrin的击倒作用正好相反，这意味着因果关系。这项建议将通过使用分子和遗传方法改变Ezrin的功能，在细胞和肾脏冷藏损伤模型中测试Ezrin诱导的细胞保护的可能机制。这些研究将确定Ezrin在其适当的配置下，是否在冷缺血和再灌流期间保护细胞膜结构和保存细胞线粒体功能。肾上皮细胞模型将用于细胞表达不同数量的野生型和突变型Ezrin蛋白。还将使用表达突变Ezrin基因的转基因Ezrin敲入小鼠的分离肾小管和整个肾脏建立更高阶的保存损伤模型。在这些研究结束时，我们将收集有用的分子信息，了解Ezrin如何参与冷藏肾脏保存损伤，以及这种参与是否与临床相关。这些见解，加上之前确定的机制，可能会提出改善肾脏保存的可能治疗目标，这将允许从高风险捐赠者那里获取不那么健壮的肾脏，以帮助缓解供者短缺的问题。 公共卫生相关性：肾移植是终末期肾功能衰竭患者的首选治疗方法，但可用的可用供体肾脏不足。这种扩大捐赠者库的需要增加了了解如何更好地保存器官以供移植的需要，这反过来又需要对器官保存损伤的机制有更全面的了解。这一建议与人类健康相关，因为它将弥合我们在理解肾脏从捐赠者身上移除后如何损伤的基本分子机制方面的一些关键知识空白。这将揭示集中治疗的新治疗靶点，并有助于缓解捐赠者短缺。

DESCRIPTION (provided by applicant): Kidney transplantation is the only viable treatment for patients with end stage renal failure. However, the severe organ shortage leaves thousands in the US to suffer and die on waiting lists. Expanding the donor pool to include high risk donors may solve this shortage if those grafts could be preserved properly. However, organ preservation injury that is suffered after organ harvest is not well understood and little major advancement have been made in this area in the last 20 years. Therefore, the objective of this proposal is to identify novel molecular mechanisms of renal preservation injury involving the cytoskeleton system and to use this knowledge to direct future therapeutic approaches. Preliminary data show that the cytoskeleton protein ezrin fails during renal cold storage and that augmenting expression of ezrin significantly protects renal tubule epithelial cells from preservation injury. Conversely, ezrin knock-down does the opposite, which suggests causation. This proposal will test likely mechanisms of ezrin-induced cytoprotection in cell and kidney cold storage injury models by using molecular and genetic approaches to alter ezrin functionality. These studies will determine if ezrin, in its proper configuration, protects cell membrane structure and preserves cell mitochondrial function during cold ischemia and reperfusion. Renal epithelial cell models will be used where the cells express different amounts of wild type and mutant ezrin protein. Higher order models of preservation injury using isolated renal tubules and whole kidneys derived from transgenic ezrin knock-in mice expressing mutant ezrin genes will also be used. At the end of these studies, we will have gathered useful molecular information of how ezrin is involved in cold storage renal preservation injury and if this involvement is clinically relevant. These insights, together with previously identified mechanisms, may suggest likely therapeutic targets to improve renal preservation, which will allow harvesting of less robust kidneys from high risk donors to help alleviate the donor shortage. PUBLIC HEALTH RELEVANCE: Kidney transplantation is the treatment of choice for patients in end stage renal failure but there are not enough usable donor kidneys available. This need to expand the donor pool has increased the need to understand how to better preserve organs for transplantation, which in turn requires a more complete understanding of the mechanisms of organ preservation injury. This proposal is relevant to human health because it will bridge some of the key knowledge gaps in our understanding of the basic molecular mechanisms of how kidneys are injured after they are removed from the donor. This will reveal novel therapeutic targets to focus treatments and help alleviate the donor shortage.