scholarly journals Fate-mapping within human kidney organoids reveals conserved mammalian nephron progenitor lineage relationships

2018 ◽  
Author(s):  
Sara E Howden ◽  
Jessica M Vanslambrouck ◽  
Sean B Wilson ◽  
Ker Sin Tan ◽  
Melissa H Little
Keyword(s):  
Stem Cell ◽  
Pluripotent Stem Cell ◽  
Kidney Development ◽  
Collecting Duct ◽  
Kidney Tissue ◽  
Human Kidney ◽  
Lineage Tracing ◽  
Nephron Progenitor ◽  
Kidney Organoids

AbstractWhile mammalian kidney morphogenesis has been well documented, human kidney development is poorly understood. Here we combine reprogramming, CRISPR/Cas9 gene-editing and organoid technologies to study human nephron lineage relationships in vitro. Early kidney organoids contained a SIX2+ population with a transcriptional profile akin to human nephron progenitors. Lineage-tracing using gene-edited induced pluripotent stem cell (iPSC) lines revealed that SIX2-expressing cells contribute to nephron formation but not to the putative collecting duct epithelium. However, Cre-mediated temporal induction of the SIX2+ lineage revealed a declining capacity for these cells to contribute to nephron formation over time. This suggests human kidney organoids, unlike the developing kidney in vivo, lack a nephron progenitor niche capable of both self-renewal and ongoing nephrogenesis. Nonetheless, human iPSC-derived kidney tissue maintains previously identified lineage relationships supporting the utility of pluripotent stem cell-derived kidney organoids for interrogating the molecular and cellular basis of early human development.

scholarly journals Human pluripotent stem cell-derived kidney organoids for personalized congenital and idiopathic nephrotic syndrome modeling

2021 ◽  
Author(s):  
Jitske Jansen ◽  
Bartholomeus T van den Berge ◽  
Martijn van den Broek ◽  
Rutger J Maas ◽  
Brigith Willemsen ◽  
...  
Keyword(s):  
Nephrotic Syndrome ◽  
Stem Cell ◽  
Pluripotent Stem Cell ◽  
Idiopathic Nephrotic Syndrome ◽  
Super Resolution ◽  
Induced Pluripotent Stem Cell ◽  
Glomerular Injury ◽  
Kidney Organoids

Nephrotic syndrome (NS) is characterized by severe proteinuria as a consequence of kidney glomerular injury due to podocyte damage. In vitro models mimicking in vivo podocyte characteristics are a prerequisite to resolve NS pathogenesis. Here, we report human induced pluripotent stem cell derived kidney organoids containing a podocyte population that heads towards adult podocytes and were superior compared to 2D counterparts, based on scRNA sequencing, super-resolution imaging and electron microscopy. In this study, these next-generation podocytes in kidney organoids enabled personalized idiopathic nephrotic syndrome modeling as shown by activated slit diaphragm signaling and podocyte injury following protamine sulfate treatment and exposure to NS plasma containing pathogenic permeability factors. Organoids cultured from cells of a patient with heterozygous NPHS2 mutations showed poor NPHS2 expression and aberrant NPHS1 localization, which was reversible after genetic correction. Repaired organoids displayed increased VEGFA pathway activity and transcription factor activity known to be essential for podocyte physiology, as shown by RNA sequencing. This study shows that organoids are the preferred model of choice to study idiopathic and congenital podocytopathies.

scholarly journals 3D kidney organoids for bench-to-bedside translation

2020 ◽  
Author(s):  
Navin Gupta✉ ◽  
Emre Dilmen ◽  
Ryuji Morizane
Keyword(s):  
Collecting Duct ◽  
Developmental Toxicity ◽  
Human Kidney ◽  
Great Promise ◽  
Duct Systems ◽  
Recent Developments ◽  
Induced Pluripotent ◽  
Kidney Organoids

Abstract The kidneys are essential organs that filter the blood, removing urinary waste while maintaining fluid and electrolyte homeostasis. Current conventional research models such as static cell cultures and animal models are insufficient to grasp the complex human in vivo situation or lack translational value. To accelerate kidney research, novel research tools are required. Recent developments have allowed the directed differentiation of induced pluripotent stem cells to generate kidney organoids. Kidney organoids resemble the human kidney in vitro and can be applied in regenerative medicine and as developmental, toxicity, and disease models. Although current studies have shown great promise, challenges remain including the immaturity, limited reproducibility, and lack of perfusable vascular and collecting duct systems. This review gives an overview of our current understanding of nephrogenesis that enabled the generation of kidney organoids. Next, the potential applications of kidney organoids are discussed followed by future perspectives. This review proposes that advancement in kidney organoid research will be facilitated through our increasing knowledge on nephrogenesis and combining promising techniques such as organ-on-a-chip models.

Pluripotent stem cell-derived kidney organoids: An in vivo-like in vitro technology

2016 ◽  
Vol 790 ◽  
pp. 12-20 ◽  
Author(s):  
Frans Schutgens ◽  
Marianne C. Verhaar ◽  
Maarten B. Rookmaaker
Keyword(s):  
Stem Cell ◽  
Pluripotent Stem Cell ◽  
Kidney Organoids

scholarly journals Characterization of the Hoechst 33342 side population from normal and malignant human renal epithelial cells

2008 ◽  
Vol 295 (3) ◽  
pp. F680-F687 ◽  
Author(s):  
Sanjai K. Addla ◽  
Mick D. Brown ◽  
Claire A. Hart ◽  
Vijay A. C. Ramani ◽  
Noel W. Clarke
Keyword(s):  
Stem Cell ◽  
Epithelial Cells ◽  
Side Population ◽  
Kidney Tissue ◽  
Human Kidney ◽  
Proliferative Potential ◽  
Stem Cell Markers ◽  
Hoechst 33342 ◽  
Tissue Samples

The fundamental changes which predispose for renal cell carcinoma (RCC) are poorly characterized. It is hypothesized that “cancer stem cells” may be influential in carcinogenesis, and the epithelial side population (SP) is enriched for stemlike cells in other epithelial cancers. In this study, we have isolated and characterized the SP and non-SP (NSP) populations from normal (NK) and malignant (RCC) human kidney tissue. NK specimens were taken from patients undergoing non-renal cancer surgery and paired malignant and macroscopically normal tissue samples were taken from patients undergoing surgery for RCC. The Hoechst 33342 dye efflux technique was used to isolate epithelial SP and NSP from normal and malignant human renal tissue. Cellular subpopulations were phenotyped for lineage, cell cycle, and putative stem cell markers, and functionally characterized using in vitro colony-forming and proliferation assays. The SP constituted 3.8 ± 0.4 and 5.9 ± 0.9% of epithelial cells in NK and RCC, respectively, of which 14.1 ± 3.5 and 13.2 ± 3.6% were shown to be in G0. SP cells demonstrated greater proliferative potential in colony-forming efficiency, long-term culture, and spheroids assays and were shown to be maintained upon tissue culture passage. We have shown that the renal SP is enriched for quiescent cells, with a high proliferative capacity and stemlike properties. The population is, however, heterogeneous, confirming that the terms “SP cell” and “stem cell” cannot be used interchangeably.

scholarly journals High throughput single cell RNA-seq of developing mouse kidney and human kidney organoids reveals a roadmap for recreating the kidney

2017 ◽  
Author(s):  
Alexander N. Combes ◽  
Belinda Phipson ◽  
Luke Zappia ◽  
Kynan T. Lawlor ◽  
Pei Xuan Er ◽  
...  
Keyword(s):  
Single Cell ◽  
Kidney Tissue ◽  
Human Kidney ◽  
Population Based ◽  
Mouse Kidney ◽  
Rna Seq ◽  
Kidney Morphogenesis ◽  
Human Ipsc ◽  
Kidney Organoids

AbstractRecent advances in our capacity to differentiate human pluripotent stem cells to human kidney tissue are moving the field closer to novel approaches for renal replacement. Such protocols have relied upon our current understanding of the molecular basis of mammalian kidney morphogenesis. To date this has depended upon population based-profiling of non-homogenous cellular compartments. In order to improve our resolution of individual cell transcriptional profiles during kidney morphogenesis, we have performed 10x Chromium single cell RNA-seq on over 6000 cells from the E18.5 developing mouse kidney, as well as more than 7000 cells from human iPSC-derived kidney organoids. We identified 16 clusters of cells representing all major cell lineages in the E18.5 mouse kidney. The differentially expressed genes from individual murine clusters were then used to guide the classification of 16 cell clusters within human kidney organoids, revealing the presence of distinguishable stromal, endothelial, nephron, podocyte and nephron progenitor populations. Despite the congruence between developing mouse and human organoid, our analysis suggested limited nephron maturation and the presence of ‘off target’ populations in human kidney organoids, including unidentified stromal populations and evidence of neural clusters. This may reflect unique human kidney populations, mixed cultures or aberrant differentiation in vitro. Analysis of clusters within the mouse data revealed novel insights into progenitor maintenance and cellular maturation in the major renal lineages and will serve as a roadmap to refine directed differentiation approaches in human iPSC-derived kidney organoids.

scholarly journals Graft immaturity and safety concerns in transplanted human kidney organoids

2019 ◽  
Vol 51 (11) ◽  
pp. 1-13 ◽  
Author(s):  
Sun Ah Nam ◽  
Eunjeong Seo ◽  
Jin Won Kim ◽  
Hyung Wook Kim ◽  
Hong Lim Kim ◽  
...  
Keyword(s):  
Therapeutic Option ◽  
Kidney Tissue ◽  
Human Kidney ◽  
Ips Cells ◽  
Attractive Potential ◽  
Transplanted Kidney ◽  
Filtration Barrier ◽  
Kidney Organoids

AbstractFor chronic kidney disease, regeneration of lost nephrons with human kidney organoids derived from induced pluripotent stem (iPS) cells is proposed to be an attractive potential therapeutic option. It remains unclear, however, whether organoids transplanted into kidneys in vivo would be safe or functional. Here, we purified kidney organoids and transplanted them beneath the kidney capsules of immunodeficient mice to test their safety and maturity. Kidney organoid grafts survived for months after transplantation and became vascularized from host mouse endothelial cells. Nephron-like structures in grafts appeared more mature than kidney organoids in vitro, but remained immature compared with the neighboring mouse kidney tissue. Ultrastructural analysis revealed filtration barrier-like structures, capillary lumens, and tubules with brush border in the transplanted kidney organoids, which were more mature than those of the kidney organoids in vitro but not as organized as adult mammalian kidneys. Immaturity was a common feature of three separate differentiation protocols by immunofluorescence analysis and single cell RNA sequencing. Stroma of transplanted kidney organoid grafts were filled with vimentin-positive mesenchymal cells, and chondrogenesis, cystogenesis, and stromal expansion were observed in the long term. Transcription profiles showed that long-term maintenance after kidney organoid transplantation induced transcriptomic reprogramming with prominent suppression of cell-cycle-related genes and upregulation of extracellular matrix organization. Our data suggest that kidney organoids derived from iPS cells may be transplantable but strategies to improve nephron differentiation and purity are required before they can be applied in humans as a therapeutic option.

scholarly journals In Vivo Assessment of Size-Selective Glomerular Sieving in Transplanted Human Induced Pluripotent Stem Cell–Derived Kidney Organoids

2020 ◽  
Vol 31 (5) ◽  
pp. 921-929 ◽  
Author(s):  
Cathelijne W. van den Berg ◽  
Angela Koudijs ◽  
Laila Ritsma ◽  
Ton J. Rabelink
Keyword(s):  
Stem Cell ◽  
Molecular Mass ◽  
Glomerular Filtration ◽  
Pluripotent Stem Cell ◽  
Disease Modeling ◽  
Left Kidney ◽  
Kidney Tissue ◽  
Human Pluripotent Stem Cell ◽  
Filtration Barrier ◽  
Kidney Organoids

BackgroundThe utility of kidney organoids in regenerative medicine will rely on the functionality of the glomerular and tubular structures in these tissues. Recent studies have demonstrated the vascularization and subsequent maturation of human pluripotent stem cell–derived kidney organoids after renal subcapsular transplantation. This raises the question of whether the glomeruli also become functional upon transplantation.MethodsWe transplanted kidney organoids under the renal capsule of the left kidney in immunodeficient mice followed by the implantation of a titanium imaging window on top of the kidney organoid. To assess glomerular function in the transplanted human pluripotent stem cell–derived kidney tissue 1, 2, and 3 weeks after transplantation, we applied high-resolution intravital multiphoton imaging through the imaging window during intravenous infusion of fluorescently labeled low and high molecular mass dextran molecules or albumin.ResultsAfter vascularization, glomerular structures in the organoid displayed dextran and albumin size selectivity across their glomerular filtration barrier. We also observed evidence of proximal tubular dextran reuptake.ConclusionsOur results demonstrate that human pluripotent stem cell–derived glomeruli can develop an appropriate barrier function and discriminate between molecules of varying size. These characteristics together with tubular presence of low molecular mass dextran provide clear evidence of functional filtration. This approach to visualizing glomerular filtration function will be instrumental for translation of organoid technology for clinical applications as well as for disease modeling.

scholarly journals A simple bioreactor-based method to generate kidney organoids from pluripotent stem cells

2017 ◽  
Author(s):  
Aneta Przepiorski ◽  
Veronika Sander ◽  
Tracy Tran ◽  
Jennifer A. Hollywood ◽  
Brie Sorrenson ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Kidney Development ◽  
Kidney Tissue ◽  
Experimental System ◽  
Human Kidney ◽  
Cost Effective ◽  
Cost Effective Method ◽  
Induced Pluripotent ◽  
Kidney Organoids

SummaryKidney organoids generated from human pluripotent stem cells have the potential to revolutionize how kidney development and injury are studied. Current protocols are technically complex and suffer from poor reproducibility and high reagent costs restricting scalability. To overcome these issues, we have established a simple, inexpensive and robust method to grow kidney organoids in bulk from human induced pluripotent stem cells. Our organoids develop tubular structures by day (d) 8 and show optimal tissue morphology at d14. A comparison with fetal human kidney suggests that d14 organoid renal structures most closely resemble ‘capillary loop’ stage nephrons. We show that deletion of HNF1B, a transcription factor linked to congenital kidney defects, interferes with tubulogenesis, validating our experimental system for studying renal developmental biology. Taken together, our protocol provides a fast, efficient and cost-effective method for generating large quantities of human fetal kidney tissue, enabling the study of normal and aberrant human renal development.

scholarly journals Kidney Organoids and Tubuloids

Cells ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 1326 ◽  
Author(s):  
Fjodor A. Yousef Yengej ◽  
Jitske Jansen ◽  
Maarten B. Rookmaaker ◽  
Marianne C. Verhaar ◽  
Hans Clevers
Keyword(s):  
High Throughput Screening ◽  
Drug Targets ◽  
Kidney Development ◽  
Disease Modeling ◽  
Kidney Diseases ◽  
Collecting Duct ◽  
Distal Tubule ◽  
Metanephric Mesenchyme ◽  
Kidney Organoids

In the past five years, pluripotent stem cell (PSC)-derived kidney organoids and adult stem or progenitor cell (ASC)-based kidney tubuloids have emerged as advanced in vitro models of kidney development, physiology, and disease. PSC-derived organoids mimic nephrogenesis. After differentiation towards the kidney precursor tissues ureteric bud and metanephric mesenchyme, their reciprocal interaction causes self-organization and patterning in vitro to generate nephron structures that resemble the fetal kidney. ASC tubuloids on the other hand recapitulate renewal and repair in the adult kidney tubule and give rise to long-term expandable and genetically stable cultures that consist of adult proximal tubule, loop of Henle, distal tubule, and collecting duct epithelium. Both organoid types hold great potential for: (1) studies of kidney physiology, (2) disease modeling, (3) high-throughput screening for drug efficacy and toxicity, and (4) regenerative medicine. Currently, organoids and tubuloids are successfully used to model hereditary, infectious, toxic, metabolic, and malignant kidney diseases and to screen for effective therapies. Furthermore, a tumor tubuloid biobank was established, which allows studies of pathogenic mutations and novel drug targets in a large group of patients. In this review, we discuss the nature of kidney organoids and tubuloids and their current and future applications in science and medicine.

scholarly journals PKD1-Dependent Renal Cystogenesis in Human Induced Pluripotent Stem Cell-Derived Ureteric Bud/Collecting Duct Organoids

2020 ◽  
Vol 31 (10) ◽  
pp. 2355-2371 ◽  
Author(s):  
Shohei Kuraoka ◽  
Shunsuke Tanigawa ◽  
Atsuhiro Taguchi ◽  
Akitsu Hotta ◽  
Hitoshi Nakazato ◽  
...  
Keyword(s):  
Stem Cell ◽  
Kidney Disease ◽  
Pluripotent Stem Cell ◽  
Collecting Duct ◽  
Induced Pluripotent Stem Cell ◽  
Cyst Formation ◽  
Ureteric Bud ◽  
Induced Pluripotent ◽  
Nephron Progenitor ◽  
Camp Stimulation

BackgroundAutosomal dominant polycystic kidney disease (ADPKD) is the most common hereditary kidney disease leading to renal failure, wherein multiple cysts form in renal tubules and collecting ducts derived from distinct precursors: the nephron progenitor and ureteric bud (UB), respectively. Recent progress in induced pluripotent stem cell (iPSC) biology has enabled cyst formation in nephron progenitor–derived human kidney organoids in which PKD1 or PKD2, the major causative genes for ADPKD, are deleted. However, cysts have not been generated in UB organoids, despite the prevalence of collecting duct cysts in patients with ADPKD.MethodsCRISPR-Cas9 technology deleted PKD1 in human iPSCs and the cells induced to differentiate along pathways leading to formation of either nephron progenitor or UB organoids. Cyst formation was investigated in both types of kidney organoid derived from PKD1-deleted iPSCs and in UB organoids generated from iPSCs from a patient with ADPKD who had a missense mutation.ResultsCysts formed in UB organoids with homozygous PKD1 mutations upon cAMP stimulation and, to a lesser extent, in heterozygous mutant organoids. Furthermore, UB organoids generated from iPSCs from a patient with ADPKD who had a heterozygous missense mutation developed cysts upon cAMP stimulation.ConclusionsCysts form in PKD1 mutant UB organoids as well as in iPSCs derived from a patient with ADPKD. The organoids provide a robust model of the genesis of ADPKD.

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