A Novel Mouse Model of Podocyte Injury

描述(由申请人提供)：一种新的足细胞损伤小鼠模型：遗传学研究强调了肾小球上皮细胞(足细胞)在肾小球疾病过程发展中的重要性。此外，最近的研究表明足细胞耗竭在获得性肾小球疾病如糖尿病肾病的发病机制中起着重要作用。事实上，越来越多的证据表明，足细胞耗竭可能是导致进行性肾损伤的最终共同途径，这是肾小球疾病过程的特征。通过使用基因操纵的动物，研究人员研究肾小球疾病的能力得到了显著提高。在这些通过基因操作的动物中，最突出的是老鼠，它们已经成为在脊椎动物中进行基因操作的首选动物。不幸的是，小鼠对许多在啮齿动物身上发展的肾小球疾病模型具有抵抗力。这一限制大大削弱了研究人员使用基因操纵的小鼠研究肾小球疾病过程的能力。为了回应名为“NIDDK相关疾病的动物模型”的项目公告(PA-07-012)，我们建议开发一种足细胞损伤的小鼠模型，该模型将允许可复制的和分级的足细胞损伤。建议的模型应有助于肾小球疾病的诊断、预防和/或治疗干预措施的临床前测试，因此，建议的实验与本计划的公告直接相关。在实验中，我们将使用足细胞特异的podocin启动子来靶向表达酵母酶胞嘧啶脱氨酶(CD)到肾小球上皮细胞。这种酶催化药物5-氟胞嘧啶(5-FC)转化为5-氟尿嘧啶(5-FU)，5-FU是一种代谢产物，既能抑制DNA和RNA的合成，又能促进足细胞等没有活性分裂的细胞的死亡。在拟议的研究中，我们将1.建立在肾小球足细胞特异性表达CD的转基因(TG)小鼠，并确定5-FC产生足细胞损伤的剂量依赖能力以及足细胞损伤的可逆性；2.通过将CD TG动物与秋田鼠杂交，在1型糖尿病遗传模型中测试足细胞耗竭在加强肾小球损伤中的作用。拟议的小鼠模型的成功开发将使研究人员能够测试足细胞损伤引起的疾病的预防和/或治疗干预措施，并使用基因操纵的小鼠来确定治疗人类肾小球疾病的新靶点。公共卫生相关性：小鼠是在脊椎动物中进行基因操作的首选动物。不幸的是，小鼠对在啮齿动物身上形成的肾脏疾病动物模型具有抵抗力。小鼠模型的这种局限性极大地限制了研究人员利用基因操纵的小鼠研究肾脏疾病的能力。这项赠款申请的目标是开发一种肾脏疾病的小鼠模型，该模型将促进使用基因操纵的小鼠对肾脏疾病的诊断、预防和/或治疗干预进行临床前测试。

DESCRIPTION (provided by applicant): A novel mouse model of podocyte injury: Genetic studies have highlighted the importance of glomerular epithelial cells (podocytes) in the development of glomerular disease processes. Moreover, recent studies suggest an important role for podocyte depletion in the pathogenesis of acquired glomerular diseases such as diabetic nephropathy. Indeed, accumulating evidence suggests that podocyte depletion may be a final common pathway causing progressive renal injury characteristic of glomerular disease processes. The ability of investigators to study glomerular diseases has been significantly enhanced by the use of genetically manipulated animals. Preeminent among these genetically manipulated animals are mice which have become the animals of choice for performing genetic manipulations in vertebrates. Unfortunately, mice are resistant to many of the glomerular disease models developed in rodents. This limitation has significantly impaired the ability of investigators to study glomerular disease processes using genetically manipulated mice. In response to the Program Announcement (PA-07-012) entitled, "Animal models of NIDDK-relevant diseases", we propose to develop a mouse model of podocyte injury that will permit reproducible and graded degrees of podocyte damage. The proposed model should facilitate the preclinical testing of diagnostic, preventive and/or therapeutic interventions in glomerular diseases and, therefore, the proposed experiments are directly relevant to this Program Announcement. For the experiments, we will use the podocyte specific podocin promoter to target expression of the yeast enzyme cytosine deaminase (CD) to glomerular epithelial cells. This enzyme catalyzes the conversion of the drug 5- flucytosine (5-FC) to 5-fluorouracil (5-FU), a metabolite that inhibits both DNA and RNA synthesis and, in turn, promotes death in cells that are not activity dividing such as podocytes. In the proposed studies, we will 1. Create transgenic (TG) mice expressing CD specifically in glomerular podocyte and determine the dose dependent ability of 5-FC produce podocyte injury as well as reversibility of the podocyte damage, and 2. Test the role of podocyte depletion in augmenting glomerular injury in a genetic model of type 1 diabetes mellitus by crossing CD TG animals with Akita mice. The successful development of the proposed mouse model will permit investigators to test preventive and/or therapeutic interventions in diseases caused by podocyte injury as well as to use genetically manipulated mice to identify novel targets for the treatment of glomerular diseases in humans. PUBLIC HEALTH RELEVANCE: Mice are the animals of choice for performing genetic manipulations in vertebrates. Unfortunately, mice are resistant to animal models of kidney disease developed in rodents. This limitation of mouse models has significantly limited the ability of investigators to study kidney diseases using genetically manipulated mice. The goal of this grant application is to develop a mouse model of kidney disease that will facilitate the preclinical testing of diagnostic, preventive and/or therapeutic interventions in kidney diseases using genetically manipulated mice.