Use of kidney organoids to investigate developmental programs and the effects of fetal exposures

Abstract In utero exposures to insults such as pregestational diabetes can have significant repercussions for proper growth and development. Exposure to the diabetic milieu can lead to diabetic embryopathy, a disruption in normal cellular programs during critical stages of development which leads to congenital anomalies affecting

multiple organs. Kidney anomalies resulting from genetic and in utero perturbations are the leading cause of chronic kidney disease in children, markedly affecting quality of life. However, there is a significant gap in our understanding of how such insults affect these early developmental programs, precluding the generation of

interventional strategies. The limitations of mouse models and differences between human and mouse kidney programs necessitate the development of alternate systems in which to study in utero exposures. Kidney organoids represent an attractive model for their recapitulation of developmental programs in an easily

manipulatable environment. Yet, studies with kidney organoids thus far have almost exclusively focused on genetic diseases and injury affecting cells of the mature nephron. To this end, we will exploit iPSC-derived kidney organoids for their ability to recapitulate human developmental processes and test how in utero

exposures to the pregestational diabetic milieu affect kidney development programs. We will focus on programs of nephron progenitors, the precursor of all the cells in the nephron, and podocytes, a nephron progenitor descendant critical to establishing and maintaining proper blood filtration. We will mimic the

pregestational diabetic environment in culture and interrogate the effects on nephron progenitor programs by single cell (sc)RNA-seq and the assay for transposase-accessible chromatin with sequencing (ATAC-seq) as the organoids progress through this stage of differentiation (Aim 1). We will also assess the effects on maturing

podocytes utilizing similar methodology (Aim 2). We expect to identify alterations to the normal differentiation programs of these cells and uncover critical pathways affected by the diabetic milieu. These findings will serve as the foundations for future studies and aid the development of novel interventional strategies then help

mitigate the effects of in utero insults. Our studies will enable new directions for kidney organoid research, laying the groundwork for future studies into in utero insults and congenital anomalies in this ex vivo, manipulatable system.