Abstract
Human-pluripotent-stem-cell-derived kidney cells (hPSC-KCs) have important potential for disease modelling and regeneration. Whether the hPSC-KCs can reconstitute tissue-specific phenotypes is currently unknown. Here we show that hPSC-KCs self-organize into kidney organoids that functionally recapitulate tissue-specific epithelial physiology, including disease phenotypes after genome editing. In three-dimensional cultures, epiblast-stage hPSCs form spheroids surrounding hollow, amniotic-like cavities. GSK3β inhibition differentiates spheroids into segmented, nephron-like kidney organoids containing cell populations with characteristics of proximal tubules, podocytes and endothelium. Tubules accumulate dextran and methotrexate transport cargoes, and express kidney injury molecule-1 after nephrotoxic chemical injury. CRISPR/Cas9 knockout of podocalyxin causes junctional organization defects in podocyte-like cells. Knockout of the polycystic kidney disease genes PKD1 or PKD2 induces cyst formation from kidney tubules. All of these functional phenotypes are distinct from effects in epiblast spheroids, indicating that they are tissue specific. Our findings establish a reproducible, versatile three-dimensional framework for human epithelial disease modelling and regenerative medicine applications.
Highlights
Human-pluripotent-stem-cell-derived kidney cells have important potential for disease modelling and regeneration
Human pluripotent stem cells are capable of extensive self-renewal and can differentiate into diverse somatic cell types and tissues. hPSCs are genetically diverse, including thousands of human embryonic stem cell and induced pluripotent stem cell lines with patient-specific or genetargeted mutations3–6. hPSCs have emerged as a powerful and reproducible source of diverse human tissues for disease modelling and regeneration. hPSCs resemble the implantation-stage human epiblast, a tissue that forms the axes for the developing embryo and cannot be studied in living human embryos owing to ethical considerations[2,7,8,9,10]
Because these experiments were performed with Mouse ESCs (mESCs), which more closely resemble the more primitive inner cell mass (ICM) than the epiblast, it remains unclear whether the observed rosettes truly represent epiblast and whether hPSCs could form similar structures[8,12,13,14,15,16]
Summary
Human-pluripotent-stem-cell-derived kidney cells (hPSC-KCs) have important potential for disease modelling and regeneration. Knockout of the polycystic kidney disease genes PKD1 or PKD2 induces cyst formation from kidney tubules All of these functional phenotypes are distinct from effects in epiblast spheroids, indicating that they are tissue specific. W Present address: Division of Nephrology, Department of Medicine, University of Washington School of Medicine, 850 Republican Street, PO Box 358056, Seattle, Washington 98109, USA Both undifferentiated stem cells and terminally differentiated somatic cells form epithelia. Mouse ESCs (mESCs) were recently shown to form polarized rosettes with small cavities when surrounded by Matrigel extracellular matrix, suggesting the possibility of modelling early amniotic cavity formation in the epiblast[11] Because these experiments were performed with mESCs, which more closely resemble the more primitive inner cell mass (ICM) than the epiblast, it remains unclear whether the observed rosettes truly represent epiblast and whether hPSCs could form similar structures[8,12,13,14,15,16]. Better understanding of human epiblast-stage biology may lead to improvements in the directed differentiation of hPSCs into specific cell types and organoids
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