Generation of New Mouse Models of Low Nephron Numbers to Understand Pathogenesis of AKI and CKD in Humans Born Preterm

摘要 由于晚期妊娠，早产(~24-37周妊娠)的人口正在迅速增长 医疗护理。不幸的是，在36周前出生的人类肾脏发育不完全， 低肾单位数，这被认为会增加慢性肾脏疾病(CKD)的风险 以及晚年的高血压。早产儿急性肾损伤(AKI)的发生率也很高。到期 由于动物模型有限，肾活检标本少，AKI和CKD的发病机制以及 在这一人群中，对肾脏损伤和修复的细胞反应知之甚少。我们开发了新的 通过抑制Ret酪氨酸激酶建立先天性低肾单位小鼠模型(对照组的50-70%) 在肾脏发育过程中。初步研究表明，低肾单位数的肾脏有 加速缺血再灌注损伤(IRI)后AKI向CKD的转变。因为低的肾单位数是 已知会导致肾小球高滤过和更高的代谢需求，以吸收高滤过负荷的 在单个肾单位水平，我们用自噬检查肾脏的应激反应。我们发现 低肾单位数的肾脏在IRI后4周有更多的缺氧和更高的自噬反应。 因此，我们计划进行研究，以检验以下假设：1)不发达地区肾脏储备不足 肾脏导致慢性肾脏病的高风险，2)高代谢应激可能超过自噬补偿 并导致CKD的发展。目标1将生成并验证新的鼠标模型，其范围为Low 肾单位数量(对照组的25%-70%)与人类发育不全的肾脏相似。我们要买小说 使用携带突变的Ret等位基因的小鼠进行化学和遗传方法的研究 对小分子抑制剂的敏感性。我们计划用这种化学物质来抑制工程Ret的活性 在妊娠中后期，特异性抑制或删除输尿管芽中的Ret基因。肾脏发育和 将描述肾脏的结构和功能。目标2我们将使用这些新的鼠标模型来解决 肾单位数低的肾脏是否更容易发生AKI？ 损伤和不完全康复，以及2)在没有AKI或AKI既往史的情况下，CKD的风险增加 AKI暴露后CKD进展更快。我们还将研究临床相关的新生儿模型。 AKI使用低肾单位数的新生小鼠来测试AKI对发育中的肾脏的影响。目标 3将自噬描述为欠发达人群对低氧和代谢紊乱的应激反应 通过检测自噬在细胞质量控制中的作用以及在AKI到CKD转变过程中对肾脏的影响。 研究的目的是测试自噬能力和通量的下降是否响应于生理和 病理性应激对CKD的发生有贡献，减少代谢应激是否会延缓CKD的发生 发展。总的目标是获得急需的知识，这些知识可以在未来用于预防 并在不断增长的早产人口中治疗肾脏疾病。

Abstract The population of humans born preterm (~24-37 w gestation) is growing rapidly as a result of advanced medical care. Unfortunately, humans born before 36 w gestation have incomplete kidney development and low nephron numbers, which has been hypothesized to cause increased risk of chronic kidney disease (CKD) and hypertension later in life. Premature infants also have a high incidence of acute kidney injury (AKI). Due to limited animal models and few kidney biopsy samples, the pathogenesis of AKI and CKD as well as the cellular response to kidney injury and repair in this population is poorly understood. We have developed new mouse models of congenital low nephron numbers (50-70% of controls) by inhibiting Ret tyrosine kinase during kidney development. Preliminary studies showed that kidneys with low nephron numbers had an accelerated AKI to CKD transition following ischemia-reperfusion injury (IRI). Since low nephron numbers is known to cause glomerular hyperfiltration and higher metabolic demands for absorption of high filter load of Na+ at the single nephron level, we examined kidneys for stress response with autophagy. We found that kidneys with low nephron numbers were more hypoxic and had higher autophagic response 4 w post-IRI. Therefore, we have planned studies to test the following hypotheses: 1) Low renal reserve in underdeveloped kidneys contributes to high risk of CKD, and 2) High metabolic stress may exceed autophagic compensation and lead to CKD development. Aim 1 will generate and validate new mouse models with a range of low nephron numbers (25-70% of control) that resemble human underdeveloped kidneys. We will take novel chemical and genetic approaches using mice harboring a floxed and mutant Ret allele that renders it susceptibility to a small molecule inhibitor. We plan to inhibit the engineered Ret activity with the chemical inhibitor or delete Ret gene specifically in the ureteric bud during mid-late gestation. Kidney development and renal structure and function will be characterized. Aim 2 We will use these new mouse models to address the questions of whether kidneys with low nephron numbers: 1) are more susceptible to AKI with more severe injury and incomplete recovery, and 2) have increased risk of CKD in the absence of prior history of AKI or more rapid CKD development after AKI exposure. We will also study clinically relevant models of neonatal AKI using newborn mice with low nephron number to test the impact of AKI on the developing kidneys. Aim 3 will characterize autophagy as a stress response to hypoxia and metabolic perturbations in underdeveloped kidneys by examining the role of autophagy in cell quality control as well as during the AKI to CKD transition. Studies are designed to test whether a decline in autophagic capacity and flux in response to physiologic and pathologic stress contributes to CKD development, and whether reducing metabolic stress delays CKD development. The overall goal is to obtain much needed knowledge that could be used in the future to prevent and treat kidney disease in the growing population of humans born preterm.