Podocyte depletion/ regeneration in evolution & reversal of diabetic nephropathy

描述(申请人提供)：糖尿病肾病(DN)是美国终末期肾病的最大单一原因。目前的糖尿病治疗方法在逆转已确定的并发症方面并不有效，例如糖尿病肾病。我们最近报道了一种新的糖尿病肾病小鼠模型，即具有ob/ob瘦素缺乏突变的BTBR小鼠株，该突变与人类糖尿病肾病非常相似，包括早期足细胞丢失(PODO)。我们提供的初步数据清楚地表明，在我们的模型中可以实现肾病的可逆性。在这项建议中，我们以这些观察为基础，定义和优化逆转糖尿病肾病的策略，重点放在两个可能在糖尿病肾病及其逆转的发病机制中起关键作用的基本机制：PODO的耗竭和再生。我们在我们的模型中展示了令人兴奋的数据，足字节数可以通过反转dN来恢复。在我们的第一个具体目标中，我们制定了一个策略来确定再生PODO的来源。利用谱系追踪研究，我们将测试邻近的顶叶上皮细胞PECs是否可以作为再生PODOS的局部祖细胞。我们将测试几种干预措施，包括人类糖尿病肾病的常用疗法，以测试它们对逆转糖尿病肾病缺乏疗效是否与它们无法促进PODO#的恢复有关。我们将把我们的观察扩展到人类肾脏活检的糖尿病肾病，以直接将我们在小鼠身上的观察结果转化为人类疾病。在我们的第二个特定目标中，我们将研究促进PODO丢失和促进再生的机制，重点是由被认为是糖尿病肾病足细胞损伤的主要原因的活性氧诱导的线粒体氧化应激所引起的损伤。我们利用足细胞特异性和系统清除mtROS的策略来测试这些方法是否可以消除糖尿病肾病的进展和/或结合PODO#的恢复来促进其逆转。我们通过创造以线粒体受限的方式诱导过表达ROS清道夫过氧化氢酶的转基因小鼠，以及通过注射具有减少线粒体氧化应激能力的新型多肽来追求这些策略。总之，拟议的研究将验证我们的中心假设：PODOS的丢失是糖尿病肾病特征性病变发展的早期和近期步骤，进一步的PODO丢失和更新是同时活跃的过程，PODO丢失的预防在很大程度上限制了糖尿病肾病的发展，最终糖尿病肾病的修复需要恢复PODO数。这些研究的影响将是1)建立新的范式，即在糖尿病肾病中实现足部再生，并将重点放在PECs在糖尿病肾病的进化和修复上。2)首次建立一种机制，通过对PODO的mtROS损伤是否是糖尿病肾病的因果关系和发展所必需的高度特异性的测试，来完成PODO的再生以及通过该机制可以逆转糖尿病肾病。3)如果成功，一种新型的mtROS小分子抑制剂可以恢复PODO的丢失，这可能为一类具有逆转人类糖尿病肾病潜力的新型治疗药物提供原理证明。 公共卫生相关性：这些研究使用了一种新的糖尿病肾病小鼠模型，该模型具有我们实验室的特征(患有瘦素缺乏的BTBR小鼠品系)，当瘦素被取代时，该模型是可逆的。这笔赠款探索了可逆性的潜在机制(替换通常在人类和实验性糖尿病肾病中丢失的肾脏细胞群)，重点是足细胞，这是肾脏中一种独特的细胞类型。然后，我们将测试新的治疗方法，基于纠正线粒体(存在于所有细胞中的细胞器)的代谢损伤，这些损伤专门针对足细胞线粒体，作为促进糖尿病肾病逆转的一种方式。

DESCRIPTION (provided by applicant): Diabetic Nephropathy (DN) is the largest single cause of end-stage renal disease in the United States. Current therapies for diabetes are not effective in reversing established complications such as DN. We have recently reported a new murine model of DN, the BTBR mouse strain with the ob/ob leptin deficiency mutation that closely resembles human DN including early loss of podocytes (podo). We present preliminary data clearly demonstrating that reversibility of nephropathy can be achieved in our model. In this proposal, we build upon these observations to define and optimize strategies for reversal of DN with a focus on two fundamental mechanisms that may be pivotal in the pathogenesis of DN and its reversal: depletion of podos and their regeneration. We show in our model exciting data that podocyte number can be restored with reversal of DN. In our first specific aim, we develop a strategy to identify the source of the regenerating podos. Using lineage tracing studies, we will test whether neighboring parietal epithelial cells PECs can serve as a local progenitor cell niche for regenerating podos. We will test several interventions including commonly used therapies for human DN to test whether their lack of efficacy for reversal of DN is linked with their inability to promote restoration of podo #. We will extend our observations to human kidney biopsies of DN to directly translate our observations in mice to the human disease. In our second specific aim, we will investigate mechanisms that potentiate podo loss and those that facilitate regeneration, focusing on injury induced by mitochondrial oxidative stress induced by reactive oxygen species (mtROS), considered a principal cause of podocyte injury in DN. We utilize strategies of podocyte specific and systemic scavenging of mtROS to test whether these approaches can abrogate progression of DN and/or promote its reversal in conjunction with restoration of podo #. We pursue these strategies both by creation of transgenic mice that inducibly overexpress the ROS scavenger catalase in a mitochondrially restricted fashion, as well as by administration of novel peptides characterized by their ability to reduce mitochondrial oxidative stress. In aggregate, the proposed studies will enable testing of our central hypothesis: loss of podos is an early and proximate step in the development of the characteristic lesions of DN, that further podo loss and renewal are concurrently active processes, that prevention of podo loss substantially limits the development of DN, and ultimately that repair of DN requires restoration of podo number. The impact of these studies will be 1) to establish new paradigms that podo regeneration in DN can be achieved, and that place a new emphasis on PECs in the evolution and repair of DN. 2) to establish for the first time a mechanism by which podo regeneration is accomplished and by which reversal of DN may be achieved, by employing highly specific tests of whether mtROS injuries to podos are causal and required for the development of DN. 3) If successful, restoration of podo loss by a novel small molecule inhibitor of mtROS could serve as a proof of principle for a new class of therapeutic agents with potential to reverse human DN. PUBLIC HEALTH RELEVANCE: These studies use a new mouse model of diabetic kidney disease characterized by our laboratory (the BTBR mouse strain with leptin deficiency) that is reversible when leptin is replaced. This grant explores mechanisms underlying reversibility (replacement of kidney cell populations that are typically lost in human and experimental diabetic kidney disease) with emphasis on the podocyte, a unique cell type in the kidney. We then will test new therapies, based on correcting metabolic injuries to mitochondria (an organelle present within all cells), that are specifically directed to podocyte mitochondria, as a way to promote reversal of diabetic kidney disease.