3D organoid-derived human glomeruli for personalised podocyte disease modelling and drug screening

Abstract

The podocytes within the glomeruli of the kidney maintain the filtration barrier by forming interdigitating foot processes with intervening slit diaphragms, disruption in which results in proteinuria. Studies into human podocytopathies to date have employed primary or immortalised podocyte cell lines cultured in 2D. Here we compare 3D human glomeruli sieved from induced pluripotent stem cell-derived kidney organoids with conditionally immortalised human podocyte cell lines, revealing improved podocyte-specific gene expression, maintenance in vitro of polarised protein localisation and an improved glomerular basement membrane matrisome compared to 2D cultures. Organoid-derived glomeruli retain marker expression in culture for 96 h, proving amenable to toxicity screening. In addition, 3D organoid glomeruli from a congenital nephrotic syndrome patient with compound heterozygous NPHS1 mutations reveal reduced protein levels of both NEPHRIN and PODOCIN. Hence, human iPSC-derived organoid glomeruli represent an accessible approach to the in vitro modelling of human podocytopathies and screening for podocyte toxicity.

Kidney organoids: accurate models or fortunate accidents

“…Such studies suggest that the level of podocyte identity present in the glomeruli of organoids is superior to any previous model of this complex and structurally constrained cell type. Indeed, using induced pluripotent stem cell lines from congenital nephrotic syndrome patients carrying distinct point mutations in NPHS1, both Tanigawa et al (2018) and Hale et al (2018) show that the resulting organoid glomeruli display podocyte defects consistent with the patient phenotype. This is a powerful proof of concept for disease modeling using organoids.…”

“…The individual characterization of human renal cell types within organoids is being assisted via the development of fluorescent reporter tools generated using CRISPR/Cas9 gene editing (Sharmin et al 2016;Boreström et al 2018;Hale et al 2018;Howden et al 2018;van den Berg et al 2018;Vanslambrouck et al 2019). By targeting fluorescent reporter proteins to the loci of cell type-specific genes, it has been possible to visualize these cell types within the organoids in vitro (Sharmin et al 2016;Boreström et al 2018) and in vivo after transplantation (van den Berg et al 2018) as well as isolate and characterize individual cell types from within organoids using fluorescence activated cell sorting ( Fig.…”

“…This approach was applied for the temporal transcriptional analysis of the maturing glomerulus using a MAFBmTAGBFP reporter line that marks the forming podocytes ( Fig. 3M; Hale et al 2018) and has been shown recently to effectively enrich for a variety of cell types from within complex organoids, including Cold Spring Harbor Laboratory Press on July 6, 2020 -Published by genesdev.cshlp.org Downloaded from endothelium, nephron progenitor, proximal tubule, and distal tubule ( Fig. 3I-N; Vanslambrouck et al 2019).…”

“…Major challenges remain around structure, functional maturation, and scale, all of which will impact on the feasibility and accuracy of applications including disease modeling, toxicological screening, and regenerative approaches. Despite this, a number of studies have shown a capacity to recapitulate genetic forms of kidney disease in vitro, including nephronophthisis and congenital nephrotic syndrome, using patient-derived iPSC (Forbes et al 2018;Hale et al 2018;Tanigawa et al 2018). What is possible outside of the embryo is to modify culture format to optimize such models for one or other application, even when this does not necessarily involve creating a perfect model of the developing organ.…”

Ureteric Bud Cells Programmed from Embryonic Stem Cells Obtain Competence for Secondary Induction in the Kidney

“…The kidney organoids derived from PSCs have been shown to be able to mimic the in vivo kidney structures development and function in vitro (Takasato et al, 2015. The renal organoids in the four-dimensional (4D) (3D plus time) culture system self-organize highly complex tissue specific morphology, that are sufficient to model tissue development, disease and injury (Boreström et al, 2018, Kim et al, 2017, Taguchi and Nishinakamura, 2017, Lorna et al, 2018. The in vitro kidney organoid system provides new regenerative strategies as the combination of genome editing and stem cell technologies allow to generate personalized kidney organoids, which provide powerful tool for kidney disease treatment and drug toxicity screening (Czerniecki et al, 2018).…”