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Generation of patterned kidney organoids that recapitulate the adult kidney collecting duct system from expandable ureteric bud progenitors.

ABSTRACT: Current kidney organoids model development and diseases of the nephron but not the contiguous epithelial network of the kidney's collecting duct (CD) system. Here, we report the generation of an expandable, 3D branching ureteric bud (UB) organoid culture model that can be derived from primary UB progenitors from mouse and human fetal kidneys, or generated de novo from human pluripotent stem cells. In chemically-defined culture conditions, UB organoids generate CD organoids, with differentiated principal and intercalated cells adopting spatial assemblies reflective of the adult kidney's collecting system. Aggregating 3D-cultured nephron progenitor cells with UB organoids in vitro results in a reiterative process of branching morphogenesis and nephron induction, similar to kidney development. Applying an efficient gene editing strategy to remove RET activity, we demonstrate genetically modified UB organoids can model congenital anomalies of kidney and urinary tract. Taken together, these platforms will facilitate an enhanced understanding of development, regeneration and diseases of the mammalian collecting duct system.

SUBMITTER: Zeng Z

PROVIDER: S-EPMC8206157 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Human ureteric bud organoids recapitulate branching morphogenesis and differentiate into functional collecting duct cell types.

Project description:Directed differentiation of human pluripotent stem cells (hPSCs) into functional ureteric and collecting duct (CD) epithelia is essential to kidney regenerative medicine. Here we describe highly efficient, serum-free differentiation of hPSCs into ureteric bud (UB) organoids and functional CD cells. The hPSCs are first induced into pronephric progenitor cells at 90% efficiency and then aggregated into spheres with a molecular signature similar to the nephric duct. In a three-dimensional matrix, the spheres form UB organoids that exhibit branching morphogenesis similar to the fetal UB and correct distal tip localization of RET expression. Organoid-derived cells incorporate into the UB tips of the progenitor niche in chimeric fetal kidney explant culture. At later stages, the UB organoids differentiate into CD organoids, which contain >95% CD cell types as estimated by single-cell RNA sequencing. The CD epithelia demonstrate renal electrophysiologic functions, with ENaC-mediated vectorial sodium transport by principal cells and V-type ATPase proton pump activity by FOXI1-induced intercalated cells.

| S-EPMC9957856 | biostudies-literature

Generation of kidney ureteric bud and collecting duct organoid

Project description:A platform for generating expandable, branching and gene-editable ureteric bud organoid from primary mouse and human ureteric bud progenitor cells and human pluripotent stem cells, and its maturation into collecting duct organoid.

2021-05-11 | GSE149109 | GEO

<i>PKD1</i>-Dependent Renal Cystogenesis in Human Induced Pluripotent Stem Cell-Derived Ureteric Bud/Collecting Duct Organoids.

Project description: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.

| S-EPMC7609014 | biostudies-literature

Generation of kidney ureteric bud and collecting duct organoid

Project description:Generation of kidney ureteric bud and collecting duct organoid

| PRJNA627400 | ENA

HNF1B controls epithelial organization and cell polarity during ureteric bud branching and collecting duct morphogenesis

Project description:Kidney development depends critically on proper ureteric bud branching giving rise to the entire collecting duct system. The transcription factor HNF1B is required for the early steps of ureteric bud branching. Yet, the molecular and cellular events regulated by HNF1B are poorly understood. We report that specific removal of Hnf1b from the ureteric bud leads to defective cell-cell contacts and apico-basal polarity during the early branching events. High resolution ex vivo imaging combined with a membranous fluorescent reporter strategy show decreased mutant cell-rearrangements during mitosis-associated cell dispersal and severe epithelial disorganisation. Molecular analysis reveals downregulation of Gdnf-Ret pathway components and suggests that HNF1B acts both upstream and downstream of Ret-signaling by directly regulating Gfrα1 and Etv5. Subsequently, Hnf1b-deletion leads to massively mispatterned ureteric tree network, defective collecting duct differentiation and disrupted tissue architecture leading to cystogenesis. Consistently, mRNA-seq analysis performed from E15.5 control and mutant kidneys shows that the most impacted genes encode intrinsic cell-membrane components with transporter activity. Our study uncovers a fundamental and recurrring role of HNF1B in epithelial organization during early ureteric bud branching and further patterning and differentiation of the collecting duct system. Article submitted to Development. Authors: Audrey Desgrange, Claire Heliot,Ilya Skovorodkin, Saad U. Akram, Janne Heikkilä, Veli-Pekka Ronkainen, Ilkka Miinalainen I., Seppo J. Vainio and Silvia Cereghini

2017-12-15 | GSE106085 | GEO

Mouse Embryonic Stem Cell-Derived Ureteric Bud Progenitors Induce Nephrogenesis.

Project description:Generation of kidney organoids from pluripotent stem cells (PSCs) is regarded as a potentially powerful way to study kidney development, disease, and regeneration. Direct differentiation of PSCs towards renal lineages is well studied; however, most of the studies relate to generation of nephron progenitor population from PSCs. Until now, differentiation of PSCs into ureteric bud (UB) progenitor cells has had limited success. Here, we describe a simple, efficient, and reproducible protocol to direct differentiation of mouse embryonic stem cells (mESCs) into UB progenitor cells. The mESC-derived UB cells were able to induce nephrogenesis when co-cultured with primary metanephric mesenchyme (pMM). In generated kidney organoids, the embryonic pMM developed nephron structures, and the mESC-derived UB cells formed numerous collecting ducts connected with the nephron tubules. Altogether, our study established an uncomplicated and reproducible platform to generate ureteric bud progenitors from mouse embryonic stem cells.

| S-EPMC7072223 | biostudies-literature

Ret-dependent cell rearrangements in the Wolffian duct epithelium initiate ureteric bud morphogenesis.

Project description:While the genetic control of renal branching morphogenesis has been extensively described, the cellular basis of this process remains obscure. GDNF/RET signaling is required for ureter and kidney development, and cells lacking Ret are excluded from the tips of the branching ureteric bud in chimeric kidneys. Here, we find that this exclusion results from earlier Ret-dependent cell rearrangements in the caudal Wolffian duct, which generate a specialized epithelial domain that later emerges as the tip of the primary ureteric bud. By juxtaposing cells with elevated or reduced RET activity, we find that Wolffian duct cells compete, based on RET signaling levels, to contribute to this domain. At the same time, the caudal Wolffian duct transiently converts from a simple to a pseudostratified epithelium, a process that does not require Ret. Thus, both Ret-dependent cell movements and Ret-independent changes in the Wolffian duct epithelium contribute to ureteric bud formation.

| S-EPMC2762206 | biostudies-literature

Production of kidney organoids arranged around single ureteric bud trees, and containing endogenous blood vessels, solely from embryonic stem cells.

Project description:There is intense worldwide effort in generating kidney organoids from pluripotent stem cells, for research, for disease modelling and, perhaps, for making transplantable organs. Organoids generated from pluripotent stem cells (PSC) possess accurate micro-anatomy, but they lack higher-organization. This is a problem, especially for transplantation, as such organoids will not be able to perform their physiological functions. In this study, we develop a method for generating murine kidney organoids with improved higher-order structure, through stages using chimaeras of ex-fetu and PSC-derived cells to a system that works entirely from embryonic stem cells. These organoids have nephrons organised around a single ureteric bud tree and also make vessels, with the endothelial network approaching podocytes.

| S-EPMC9307805 | biostudies-literature

Generation of mitochondria-rich kidney organoids from expandable intermediate mesoderm progenitors reprogrammed from human urine cells under defined medium.

Project description:BackgroundThe kidneys require vast amounts of mitochondria to provide ample energy to reabsorb nutrients and regulate electrolyte, fluid, and blood pressure homeostasis. The lack of the human model hinders the investigation of mitochondria homeostasis related to kidney physiology and disease.ResultsHere, we report the generation of mitochondria-rich kidney organoids via partial reprogramming of human urine cells (hUCs) under the defined medium. First, we reprogrammed mitochondria-rich hUCs into expandable intermediate mesoderm progenitor like cells (U-iIMPLCs), which in turn generated nephron progenitors and formed kidney organoids in both 2D and 3D cultures. Cell fate transitions were confirmed at each stage by marker expressions at the RNA and protein levels, along with chromatin accessibility dynamics. Single cell RNA-seq revealed hUCs-induced kidney organoids (U-iKOs) consist of podocytes, tubules, and mesenchyme cells with 2D dominated with mesenchyme and 3D with tubule and enriched specific mitochondria function associated genes. Specific cell types, such as podocytes and proximal tubules, loop of Henle, and distal tubules, were readily identified. Consistent with these cell types, 3D organoids exhibited the functional and structural features of the kidney, as indicated by dextran uptake and transmission electron microscopy. These organoids can be further matured in the chick chorioallantoic membrane. Finally, cisplatin, gentamicin, and forskolin treatment led to anatomical abnormalities typical of kidney injury and altered mitochondria homeostasis respectively.ConclusionsOur study demonstrates that U-iKOs recapitulate the structural and functional characteristics of the kidneys, providing a promising model to study mitochondria-related kidney physiology and disease in a personalized manner.

| S-EPMC9569036 | biostudies-literature

Rac1 promotes kidney collecting duct integrity by limiting actomyosin activity.

Project description:A polarized collecting duct (CD), formed from the branching ureteric bud (UB), is a prerequisite for an intact kidney. The small Rho GTPase Rac1 is critical for actin cytoskeletal regulation. We investigated the role of Rac1 in the kidney collecting system by selectively deleting it in mice at the initiation of UB development. The mice exhibited only a mild developmental phenotype; however, with aging, the CD developed a disruption of epithelial integrity and function. Despite intact integrin signaling, Rac1-null CD cells had profound adhesion and polarity abnormalities that were independent of the major downstream Rac1 effector, Pak1. These cells did however have a defect in the WAVE2-Arp2/3 actin nucleation and polymerization apparatus, resulting in actomyosin hyperactivity. The epithelial defects were reversible with direct myosin II inhibition. Furthermore, Rac1 controlled lateral membrane height and overall epithelial morphology by maintaining lateral F-actin and restricting actomyosin. Thus, Rac1 promotes CD epithelial integrity and morphology by restricting actomyosin via Arp2/3-dependent cytoskeletal branching.

| S-EPMC8563289 | biostudies-literature

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