DEVELOPMENTAL ROLE OF THE GENE SIX2 IN KIDNEY NEPHRON ENDOWMENT

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 与那些出生时具有典型肾单位天赋的人相比，肾脏发育不佳导致出生时肾单位数量不足可能会产生终生后果，并与以后患心血管-肾脏疾病的显著风险相关。目前的建议的长期目标是确定驱动成功的肾单位分化的细胞机制，以实现最佳的肾单位捐赠。具体地说，我们将探索同源盒基因Sine Oculis 2(SIX2)的作用，并确定SIX2是否参与了肾脏器官发生过程中肾单位数量的建立。利用RNAi方法，实验旨在确定肾发生早期SIX2转录的失活是否会导致输尿管芽生长显著减少，从而减少器官型肾脏外植体培养中的肾单位数量。此外，使用发育过程中缺乏SIX2表达的遗传性肾发育不良小鼠模型(Brachyrrhine(BR)突变小鼠)，我们将采取系统的方法将SIX2重新引入BR小鼠的肾脏外植体，并确定外源SIX2是否刺激输尿管芽的分支，从而增强肾单位的生成。此外，我们将把含有SIX2基因的250kb Bac基因整合到BR小鼠基因组中，试图增加肾单位的数量，从而挽救与SIX2缺乏相关的肾脏表型缺陷。在抢救之后，我们将评估成年BR小鼠的肾脏生理特征是否恢复。这项提议将利用BR小鼠品系的优势，该品系表现出SIX2基因表达的单倍性不足，并且是世界上唯一可用的菌落。同样，我们已经充分描述了成年BR小鼠的生理学特征，显示出高血压和慢性肾功能衰竭，这有助于我们的表型救援实验。综合考虑，这些实验将提供构成SIX2在肾脏形态发生中的遗传基础的基本见解，并具体确定SIX2是否直接决定发育中的肾脏中的肾单位捐赠。这项研究的结果也将强调SIX2在成人病变肾脏可能的肾单位重建方法中的重要性。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Suboptimal kidney development resulting in an in-born deficit in nephron number can have lifelong consequences and is associated with a significant risk of cardiovascular-renal disease in later life, compared to those born with a typical nephron endowment. The long-term goal of the current proposal is to identify cellular mechanisms that drive successful nephron differentiation to achieve optimal nephron endowment. Specifically, we will explore the roles of the homeobox gene sine oculis 2 (six2), and determine whether six2 is involved in the establishment of nephron number during kidney organogenesis. Using an RNAi methodology, experiments are designed to determine whether inactivation of six2 transcription early in nephrogenesis results in a significant decrease in ureteric bud growth and, therefore, nephron number in an organotypic kidney explant culture. Further, using a mouse model of heritable renal hypoplasia that lacks sufficient expression of six2 during development (the Brachyrrhine (Br) mutant mouse), we will take a systematic approach to re-introduce six2 to kidney explants that are prepared from Br mice and determine if exogenous six2 stimulates branching of the ureteric buds leading to an enhanced nephron production. Additionally, we will incorporate a 250 kb Bac containing six2 gene into the Br mouse genome to attempt to increase nephron number and, thereby rescuing the defective renal phenotype associated with six2 deficiency. Following the rescue we will assess if renal physiological features are restored in the adult Br mouse. This proposal will take advantage of the Br mouse strain that displays haploinsufficiency of six2 gene expression and is the only working colony in the world. Similarly, we have fully characterized the physiology of the adult Br mouse that demonstrates hypertension and chronic renal failure facilitating our phenotypic rescue experiments. When considered together, these experiments will provide the fundamental insights that constitute the genetic basis of six2 in renal morphogenesis and specifically establish whether six2 directly determines nephron endowment in the developing kidney. Results from this study will also underscore the importance of six2 in possible nephron restoration approaches in the adult diseased kidney.