A Toolbox to Characterize Human Induced Pluripotent Stem Cell-Derived Kidney Cell Types and Organoids

Area of Science:

• Developmental Biology
• Stem Cell Biology
• Regenerative Medicine

Background:

• Studying human kidney development in vivo is challenging.
• Reporter lines are valuable for dissecting mouse kidney morphogenesis.
• Human induced pluripotent stem cells (iPSCs) offer a feasible model for studying human kidney development.

Purpose of the Study:

• To generate and validate fluorescence reporter iPSC lines for studying human kidney organoid development.
• To enable visualization and isolation of specific kidney cell populations within organoids.
• To facilitate applications like lineage tracing and protocol optimization.

Main Methods:

• CRISPR/Cas9 gene editing was used to create ten fluorescence reporter iPSC lines.
• Lines were designed to identify nephron progenitors, podocytes, proximal/distal nephron, and ureteric epithelium.
• Directed differentiation protocols were used to generate kidney organoids, followed by immunofluorescence, confocal microscopy, flow cytometry, and cell sorting.

Main Results:

• All generated iPSC reporter lines successfully formed well-patterned kidney organoids.
• Reporter expression accurately reflected endogenous gene and protein expression.
• These lines facilitated the isolation and characterization of specific kidney cell types and enabled time-lapse imaging and transplantation experiments.

Conclusions:

• A suite of validated fluorescence iPSC reporter lines for human kidney organoid morphogenesis has been generated.
• This reporter toolbox allows for visualization and isolation of key cell populations.
• These tools enhance the study of kidney organoids and their relevance to human kidney development.