Abstract

Human iPSC-derived kidney organoids have the potential to revolutionize discovery, but assessing their consistency and reproducibility across iPSC lines, and reducing the generation of off-target cells remain an open challenge. Here, we profile four human iPSC lines for a total of 450,118 single cells to show how organoid composition and development are comparable to human fetal and adult kidneys. Although cell classes are largely reproducible across time points, protocols, and replicates, we detect variability in cell proportions between different iPSC lines, largely due to off-target cells. To address this, we analyze organoids transplanted under the mouse kidney capsule and find diminished off-target cells. Our work shows how single cell RNA-seq (scRNA-seq) can score organoids for reproducibility, faithfulness and quality, that kidney organoids derived from different iPSC lines are comparable surrogates for human kidney, and that transplantation enhances their formation by diminishing off-target cells.

Highlights

  • Human induced pluripotent stem cells (iPSC)-derived kidney organoids have the potential to revolutionize discovery, but assessing their consistency and reproducibility across iPSC lines, and reducing the generation of off-target cells remain an open challenge

  • We profiled single cells at the day 0 (D0) iPSC state, at two critical milestones, day 7 (D7; immediately prior to cells being plated for 3D self-organization and nephrogenesis), and day 15 (D15; when growth factor treatment ends), and at day 29 (D29) when the organoids are mature

  • In this study, we present a comprehensive census of human kidney organoids in comparison to human adult and fetal kidneys at single-cell resolution spanning multiple iPSC cell lines, time points, protocols, and replicates including transplantation into mice, in a total of 450,118 cells

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Summary

Introduction

Human iPSC-derived kidney organoids have the potential to revolutionize discovery, but assessing their consistency and reproducibility across iPSC lines, and reducing the generation of off-target cells remain an open challenge. A comprehensive single-cell survey of kidney organoids from several iPSC lines is critically needed, as individual patient iPSCs become essential tools for precision medicine and drug development projects[7,19,20]. We address these critical questions by surveying organoid composition from more than 450,000 single cells across several iPSC lines, multiple replicates, differentiation protocols, developmental time, and after organoid transplantation into mouse, benchmarked against human fetal and adult kidney

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