ORCID Profile
0000-0003-0380-2263
Current Organisations
Novo Nordisk Foundation Centre for Stem Cell Medicine (reNEW)
,
University of Melbourne
,
Murdoch Children's Research Institute
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Research Topics
In Research Link Australia (RLA), "Research Topics" refer to ANZSRC FOR and SEO codes. These topics are either sourced from ANZSRC FOR and SEO codes listed in researchers' related grants or generated by a large language model (LLM) based on their publications.
Top 5 Research Topics
ANZSRC Field of Research (FoR)
Cell Development, Proliferation and Death | Biochemistry and Cell Biology | Genetics | Cellular Interactions (incl. Adhesion, Matrix, Cell Wall) | Developmental Genetics (incl. Sex Determination) | Nanobiotechnology | Simulation And Modelling | Plant Biology | Biochemistry and Cell Biology not elsewhere classified | Medical Biochemistry and Metabolomics not elsewhere classified | Infectious Agents | Information Storage, Retrieval And Management | Plant Cell and Molecular Biology | Biological Sciences Not Elsewhere Classified | Medical Biochemistry and Metabolomics | Other Biological Sciences | Gene Expression | Genetic Development (Incl. Sex Determination) | Central Nervous System | Regenerative Medicine (incl. Stem Cells and Tissue Engineering) |
ANZSRC Socio-Economic Objective (SEO)
Expanding Knowledge in the Biological Sciences | Nervous System and Disorders | Biological sciences | Plant Production and Plant Primary Products not elsewhere classified | Production of Biofuels (Biomass) | Urogenital System and Disorders | Information services not elsewhere classified | Cardiovascular System and Diseases | Expanding Knowledge in Engineering | Urogenital system and disorders | Blood Disorders | Reproductive system and disorders | Infectious Diseases
Related Links
Publications
Fate-mapping within human kidney organoids reveals conserved mammalian nephron progenitor lineage relationships
Publisher: Cold Spring Harbor Laboratory
Date: 10-2018
DOI: 10.1101/432161
Abstract: While 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.
Expression and Functional Analysis of Dkk1 during Early Gonadal Development
Publisher: S. Karger AG
Date: 2011
DOI: 10.1159/000327709
Abstract: WNT signalling plays a central role in mammalian sex determination by promoting ovarian development and repressing aspects of testis development in the early gonad. Dickkopf homolog 1 (DKK1) is a WNT signalling antagonist that plays critical roles in multiple developmental systems by modulating WNT activity. Here, we examined the role of DKK1 in mouse sex determination and early gonadal development. i Dkk1 /i mRNA was upregulated sex-specifically during testis differentiation, suggesting that DKK1 could repress WNT signalling in the developing testis. However, we observed overtly normal testis development in i Dkk1 /i -null XY gonads, and found no significant upregulation of i Axin2 /i or i Sp5 /i that would indicate increased canonical WNT signalling. Nor did we find significant differences in expression of key markers of testis and ovarian development. We propose that DKK1 may play a protective role that is not unmasked by loss-of-function in the absence of other stressors.
Collecting Duct-Derived Cells Display Mesenchymal Stem Cell Properties and Retain Selective In Vitro and In Vivo Epithelial Capacity
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 2015
Expression of Metanephric Nephron-Patterning Genes in Differentiating Mesonephric Tubules
Publisher: Wiley
Date: 12-04-2011
DOI: 10.1002/DVDY.22640
Cell–Cell Interactions Driving Kidney Morphogenesis
Publisher: Elsevier
Date: 2015
DOI: 10.1016/BS.CTDB.2014.12.002
Abstract: The mammalian kidney forms via cell-cell interactions between an epithelial outgrowth of the nephric duct and the surrounding nephrogenic mesenchyme. Initial morphogenetic events include ureteric bud branching to form the collecting duct (CD) tree and mesenchymal-to-epithelial transitions to form the nephrons, requiring reciprocal induction between adjacent mesenchyme and epithelial cells. Within the tips of the branching ureteric epithelium, cells respond to mesenchyme-derived trophic factors by proliferation, migration, and mitosis-associated cell dispersal. Self-inhibition signals from one tip to another play a role in branch patterning. The position, survival, and fate of the nephrogenic mesenchyme are regulated by ECM and secreted signals from adjacent tip and stroma. Signals from the ureteric tip promote mesenchyme self-renewal and trigger nephron formation. Subsequent fusion to the CDs, nephron segmentation and maturation, and formation of a patent glomerular basement membrane also require specialized cell-cell interactions. Differential cadherin, laminin, nectin, and integrin expression, as well as intracellular kinesin and actin-mediated regulation of cell shape and adhesion, underlies these cell-cell interactions. Indeed, the capacity for the kidney to form via self-organization has now been established both via the recapitulation of expected morphogenetic interactions after complete dissociation and reassociation of cellular components during development as well as the in vitro formation of 3D kidney organoids from human pluripotent stem cells. As we understand more about how the many cell-cell interactions required for kidney formation operate, this enables the prospect of bioengineering replacement structures based on these self-organizing properties.
Kidney side population reveals multilineage potential and renal functional capacity but also cellular heterogeneity
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 07-2006
A clinical overview of WT1 gene mutations
Publisher: Hindawi Limited
Date: 1997
DOI: 10.1002/(SICI)1098-1004(1997)9:3<209::AID-HUMU2>3.0.CO;2-2
Abstract: Bacterial studies based on growth curves are common in microbiology and related fields. Compared to the standard photometer and cuvette based protocols, bacterial growth curve measurements with microplate readers provide better temporal resolution, higher efficiency, and are less laborious, while analysis and interpretation of the microplate-based measurements are less straightforward. Recently, we developed a new analysis method for evaluating bacterial growth with microplate readers based on time derivatives. Here, we describe a detailed protocol for this development and provide the homemade program for the new analysis method.
CRIM1, a novel gene encoding a cysteine-rich repeat protein, is developmentally regulated and implicated in vertebrate CNS development and organogenesis
Publisher: Elsevier BV
Date: 02-2000
DOI: 10.1016/S0925-4773(99)00248-8
Abstract: Development of the vertebrate central nervous system is thought to be controlled by intricate cell-cell interactions and spatio-temporally regulated gene expressions. The details of these processes are still not fully understood. We have isolated a novel vertebrate gene, CRIM1/Crim1, in human and mouse. Human CRIM1 maps to chromosome 2p21 close to the Spastic Paraplegia 4 locus. Crim1 is expressed in the notochord, somites, floor plate, early motor neurons and interneuron subpopulations within the developing spinal cord. CRIM1 appears to be evolutionarily conserved and encodes a putative transmembrane protein containing an IGF-binding protein motif and multiple cysteine-rich repeats similar to those in the BMP-associating chordin and sog proteins. Our results suggest a role for CRIM1/Crim1 in CNS development possibly via growth factor binding.
Refining transcriptional programs in kidney development by integration of deep RNA-sequencing and array-based spatial profiling
Publisher: Springer Science and Business Media LLC
Date: 05-09-2011
The Kidney Research National Dialogue
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 10-2014
DOI: 10.2215/CJN.07310714
Modelling Cellular Interactions and Dynamics During Kidney Morphogenesis
Publisher: Springer Science and Business Media LLC
Date: 27-11-2021
DOI: 10.1007/S11538-021-00968-3
Abstract: Kidney disease and renal disorders account for a significant proportion of health complications in mid-late adulthood worldwide. Many renal deficiencies are due to improper formation of the kidneys before birth, which are caused by disorders in the developmental process that arise from genetic and/or environmental factors. Mathematical modelling can help build on experimental knowledge to increase our understanding of the complexities of kidney organogenesis. In this paper, we present a discrete cell-based model of kidney development. Specifically, we model the tip of the developing ureteric tree to investigate the behaviours of cap mesenchyme cells which are required to sustain ureteric tip growth. We find that spatial regulation of the differentiation of cap mesenchyme cells through cellular signalling is sufficient to ensure robust ureteric tip development. Additionally, we find that increased adhesion interactions between cap mesenchyme cells and the ureteric tip surface can lead to a more stable tip-cap unit. Our analysis of the various processes on this scale highlights essential components for healthy kidney growth and provides insight into mechanisms to be studied further in order to replicate the process in vitro.
Distinct but overlapping expression patterns of two vertebrate slit homologs implies functional roles in CNS development and organogenesis
Publisher: Elsevier BV
Date: 12-1998
DOI: 10.1016/S0925-4773(98)00174-9
Abstract: The Drosophila slit gene (sli) encodes a secreted leucine-rich repeat-containing protein (slit) expressed by the midline glial cells and required for normal neural development. A putative human sli homolog, SLIT1, has previously been identified by EST database scanning. We have isolated a second human sli homolog, SLIT2, and its murine homolog Slit2. Both SLIT1 and SLIT2 proteins show approximately 40% amino acid identity to slit and 60% identity to each other. In mice, both genes are expressed during CNS development in the floor plate, roof plate and developing motor neurons. As floor plate represents the vertebrate equivalent to the midline glial cells, we predict a conservation of function for these vertebrate homologs. Each gene shows additional but distinct sites of expression outside the CNS suggesting a variety of functions for these proteins.
PlexinA4 is necessary as a downstream target of Islet2 to mediate Slit signaling for promotion of sensory axon branching
Publisher: The Company of Biologists
Date: 08-2004
DOI: 10.1242/DEV.01228
Abstract: Slit is a secreted protein known to repulse the growth cones of commissural neurons. By contrast, Slit also promotes elongation and branching of axons of sensory neurons. The reason why different neurons respond to Slit in different ways is largely unknown. Islet2 is a LIM/homeodomain-type transcription factor that specifically regulates elongation and branching of the peripheral axons of the primary sensory neurons in zebrafish embryos. We found that PlexinA4, a transmembrane protein known to be a co-receptor for class III semaphorins,acts downstream of Islet2 to promote branching of the peripheral axons of the primary sensory neurons. Intriguingly, repression of PlexinA4 function by injection of the antisense morpholino oligonucleotide specific to PlexinA4 or by overexpression of the dominant-negative variant of PlexinA4 counteracted the effects of overexpression of Slit2 to induce branching of the peripheral axons of the primary sensory neurons in zebrafish embryos, suggesting involvement of PlexinA4 in the Slit signaling cascades for promotion of axonal branching of the sensory neurons. Colocalized expression of Robo, a receptor for Slit2, and PlexinA4 is observed not only in the primary sensory neurons of zebrafish embryos but also in the dendrites of the pyramidal neurons of the cortex of the mammals, and may be important for promoting the branching of either axons or dendrites in response to Slit, as opposed to the growth cone collapse.
Mentorship in Science: Response to AlShebli et al., Nature Communications 2020.
Publisher: Elsevier BV
Date: 2021
Autonomous Calcium Signaling in Human and Zebrafish Podocytes Controls Kidney Filtration Barrier Morphogenesis
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 07-2021
Abstract: Podocytes are critical to maintaining the kidney glomerular filtration barrier. Mutations in genes associated with development of nephrotic syndrome lead to elevated cytoplasmic calcium in podocytes and cause disruption of filtration barrier function. Whether calcium signaling plays a role in the initial formation of the filtration barrier is not known. Using live calcium imaging in two models, larval zebrafish and human kidney organoids, the authors demonstrate that podocyte calcium signaling is active during podocyte differentiation, is podocyte-cell autonomous, occurs independently of neighboring cell types, and is required for foot process and slit diaphragm formation. Their findings also show that developmental calcium signaling occurs by a different mechanism than disease-associated calcium perturbations, and represents a critical regulatory signal for podocyte morphogenesis and filtration barrier formation. Podocytes are critical to maintaining the glomerular filtration barrier, and mutations in nephrotic syndrome genes are known to affect podocyte calcium signaling. However, the role of calcium signaling during podocyte development remains unknown. We undertook live imaging of calcium signaling in developing podocytes, using zebrafish larvae and human kidney organoids. To evaluate calcium signaling during development and in response to channel blockers and genetic defects, the calcium biosensor GCaMP6s was expressed in zebrafish podocytes. We used electron microscopy to evaluate filtration barrier formation in zebrafish, and Fluo-4 to detect calcium signals in differentiating podocytes in human kidney organoids. Immature zebrafish podocytes (2.5 days postfertilization) generated calcium transients that correlated with interactions with forming glomerular capillaries. Calcium transients persisted until 4 days postfertilization, and were absent after glomerular barrier formation was complete. We detected similar calcium transients in maturing human organoid glomeruli, suggesting a conserved mechanism. In both models, inhibitors of SERCA or IP3 receptor calcium-release channels blocked calcium transients in podocytes, whereas lanthanum was ineffective, indicating the calcium source is from intracellular podocyte endoplasmic-reticulum stores. Calcium transients were not affected by blocking heartbeat or by blocking development of endothelium or endoderm, and they persisted in isolated glomeruli, suggesting podocyte-autonomous calcium release. Inhibition of expression of phospholipase C- γ 1, but not nephrin or phospholipase C- ε 1, led to significantly decreased calcium activity. Finally, blocking calcium release affected glomerular shape and podocyte foot process formation, supporting the critical role of calcium signaling in glomerular morphogenesis. These findings establish podocyte cell–autonomous calcium signaling as a prominent and evolutionarily conserved feature of podocyte differentiation and demonstrate its requirement for podocyte foot process formation.
Endothelial Dysfunction in the Brain
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 05-2021
DOI: 10.1161/STROKEAHA.120.032711
Abstract: The Coronavirus disease 2019 (COVID)-19 pandemic has already affected millions worldwide, with a current mortality rate of 2.2%. While it is well-established that severe acute respiratory syndrome-coronavirus-2 causes upper and lower respiratory tract infections, a number of neurological sequelae have now been reported in a large proportion of cases. Additionally, the disease causes arterial and venous thromboses including pulmonary embolism, myocardial infarction, and a significant number of cerebrovascular complications. The increasing incidence of large vessel ischemic strokes as well as intracranial hemorrhages, frequently in younger in iduals, and associated with increased morbidity and mortality, has raised questions as to why the brain is a major target of the disease. COVID-19 is characterized by hypercoagulability with alterations in hemostatic markers including high D-dimer levels, which are a prognosticator of poor outcome. Together with findings of fibrin-rich microthrombi, widespread extracellular fibrin deposition in affected various organs and hypercytokinemia, this suggests that COVID-19 is more than a pulmonary viral infection. Evidently, COVID-19 is a thrombo-inflammatory disease. Endothelial cells that constitute the lining of blood vessels are the primary targets of a thrombo-inflammatory response, and severe acute respiratory syndrome coronavirus 2 also directly infects endothelial cells through the ACE2 (angiotensin-converting enzyme 2) receptor. Being highly heterogeneous in their structure and function, differences in the endothelial cells may govern the susceptibility of organs to COVID-19. Here, we have explored how the unique characteristics of the cerebral endothelium may be the underlying reason for the increased rates of cerebrovascular pathology associated with COVID-19.
Cap mesenchyme cell swarming during kidney development is influenced by attraction, repulsion, and adhesion to the ureteric tip
Publisher: Elsevier BV
Date: 10-2016
DOI: 10.1016/J.YDBIO.2016.06.028
Abstract: Morphogenesis of the mammalian kidney requires reciprocal interactions between two cellular domains at the periphery of the developing organ: the tips of the epithelial ureteric tree and adjacent regions of cap mesenchyme. While the presence of the cap mesenchyme is essential for ureteric branching, how it is specifically maintained at the tips is unclear. Using ex vivo timelapse imaging we show that cells of the cap mesenchyme are highly motile. In idual cap mesenchyme cells move within and between cap domains. They also attach and detach from the ureteric tip across time. Timelapse tracks collected for >800 cells showed evidence that this movement was largely stochastic, with cell autonomous migration influenced by opposing attractive, repulsive and cell adhesion cues. The resulting swarming behaviour maintains a distinct cap mesenchyme domain while facilitating dynamic remodelling in response to underlying changes in the tip.
Macrophages in Renal Development, Injury, and Repair
Publisher: Elsevier BV
Date: 05-2010
DOI: 10.1016/J.SEMNEPHROL.2010.03.011
Abstract: Macrophages have long been regarded as classic mediators of innate immunity because of their production of proinflammatory cytokines and their ability to induce apoptotic cell death. As a result of such activities and the detrimental long-term effect of kidney inflammation, macrophages principally have been regarded as mediators of glomerular damage, tubular cell death, and the downstream fibrotic events leading to chronic kidney disease. Although this has been the accepted consequence of macrophage infiltration in kidney disease, macrophages also play a critical role in normal organ development, cell turnover, and recovery from injury in many organs, including the kidney. There is also a growing awareness that there is considerable heterogeneity of phenotype and function within the macrophage population and that a greater understanding of these different states of activation may result in the development of therapies specifically designed to capitalize on this variation in phenotype and cellular responses. In this review, we discuss the current understanding of induction and consequences of classic versus alternative macrophage activation and highlight what additional therapeutic options this may provide for the management of both acute and chronic kidney disease as well as renal cancer.
Regrow or repair: Potential regenerative therapies for the kidney
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 09-2006
Kidney organoids: accurate models or fortunate accidents
Publisher: Cold Spring Harbor Laboratory
Date: 10-2019
Abstract: There are now many reports of human kidney organoids generated via the directed differentiation of human pluripotent stem cells (PSCs) based on an existing understanding of mammalian kidney organogenesis. Such kidney organoids potentially represent tractable tools for the study of normal human development and disease with improvements in scale, structure, and functional maturation potentially providing future options for renal regeneration. The utility of such organotypic models, however, will ultimately be determined by their developmental accuracy. While initially inferred from mouse models, recent transcriptional analyses of human fetal kidney have provided greater insight into nephrogenesis. In this review, we discuss how well human kidney organoids model the human fetal kidney and how the remaining differences challenge their utility.
Vascular bioengineering of scaffolds derived from human discarded transplant kidneys using human pluripotent stem cell–derived endothelium
Publisher: Elsevier BV
Date: 05-2019
DOI: 10.1111/AJT.15200
+P5(D1S3309E), a novel target binding site for the Wilms' tumour suppressor 1 (WT1) gene, maps to human chromosome 1q21->q22
Publisher: S. Karger AG
Date: 1996
DOI: 10.1159/000134210
Abstract: The Wilms’ tumour suppressor 1 gene (WT1) encodes a zinc finger transcription factor critical for normal urogenital development. We have previously isolated a DNA fragment, +P5 (D1S3309E), to which all WT1 protein isoforms bind. Using PCR of a human × rodent somatic cell hybrid mapping panel, together with two-color fluorescence in situ hybridisation of +P5-containing cosmids and previously localised human chromosome 1q cosmids, we have mapped the +P5 fragment to chromosome 1q21→q22.
Three non‐overlapping regions of chromosome arm 11p allele loss identified in infantile tumors of adrenal and liver
Publisher: Wiley
Date: 10-1993
Abstract: Tumor and constitutional chromosome arm 11p genotypes were compared in 6 hepatoblastoma (HB) patients and 2 adrenal adenoma (AA) patients, with one HB patient and both AA patients displaying clinical features associated with the Beckwith-Wiedemann syndrome (BWS). Using up to 14 chromosome 11 polymorphic markers, loss of constitutional heterozygosity (LOH) was demonstrated in both AA patients and in 4 of 6 HB patients. This identified three distinct and non-overlapping regions of 11p within which LOH occurred, which were defined as lying distal to the gamma-globin locus (11p15.5), proximal to the gamma-globin locus but distal to 11p13 (LOH being detected at 11p15.1), and restricted to the 11p13 region. Specific LOH within each 11p15 region was observed in HB, and this represents the first demonstration by a single study of LOH clearly affecting separate regions of chromosome band 11p15 in a particular tumor type. One AA showed LOH restricted to 11p13 loci, implicating the involvement of the WT1 gene. The second AA patient presented with genitourinary abnormalities and we therefore examined sequences coding for 3 zinc finger domains of WT1 in both AAs. No point mutations were identified in sequence from either patient. Nonetheless our results indicate that 3 separate 11p loci may be significant in the development of tumors which arise in association with BWS.
Erratum: Kidney organoids from human iPS cells contain multiple lineages and model human nephrogenesis
Publisher: Springer Science and Business Media LLC
Date: 27-04-2016
DOI: 10.1038/NATURE17982
Does Renal Repair Recapitulate Kidney Development?
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 26-10-2016
Abstract: Over a decade ago, it was proposed that the regulation of tubular repair in the kidney might involve the recapitulation of developmental pathways. Although the kidney cannot generate new nephrons after birth, suggesting a low level of regenerative competence, the tubular epithelial cells of the nephrons can proliferate to repair the damage after AKI. However, the debate continues over whether this repair involves a persistent progenitor population or any mature epithelial cell remaining after injury. Recent reports have highlighted the expression of Sox9 , a transcription factor critical for normal kidney development, during postnatal epithelial repair in the kidney. Indeed, the proliferative response of the epithelium involves expression of several pathways previously described as being involved in kidney development. In some instances, these pathways are also apparently involved in the maladaptive responses observed after repeated injury. Whether development and repair in the kidney are the same processes or we are misinterpreting the similar expression of genes under different circumstances remains unknown. Here, we review the evidence for this link, concluding that such parallels in expression may more correctly represent the use of the same pathways in a distinct context, likely triggered by similar stressors.
DevKidCC allows for robust classification and direct comparisons of kidney organoid datasets
Publisher: Cold Spring Harbor Laboratory
Date: 20-01-2021
DOI: 10.1101/2021.01.20.427346
Abstract: Kidney organoids provide a valuable resource to understand kidney development and disease. Clustering algorithms and marker genes fail to accurately and robustly classify cellular identity between human pluripotent stem cell (hPSC)-derived organoid datasets. Here we present a new method able to accurately classify kidney cell subtypes, a hierarchical machine learning model trained using comprehensive reference data from single cell RNA-sequencing of human fetal kidney (HFK). We demonstrate the tool’s ( DevKidCC ) performance by application to all published kidney organoid datasets and a novel dataset. DevKidCC is available on Github and can be used on any kidney single cell RNA-sequence data.
Single cell analysis of the developing mouse kidney provides deeper insight into marker gene expression and ligand-receptor crosstalk
Publisher: The Company of Biologists
Date: 12-06-2019
DOI: 10.1242/DEV.178673
Abstract: Recent advances in the generation of kidney organoids and the culture of primary nephron progenitors from mouse and human have been based on knowledge of the molecular basis of kidney development in mice. Although gene expression during kidney development has been intensely investigated, single cell profiling provides new opportunities to further subsect component cell types and the signalling networks at play. Here, we describe the generation and analysis of 6732 single cell transcriptomes from the fetal mouse kidney [embryonic day (E)18.5] and 7853 sorted nephron progenitor cells (E14.5). These datasets provide improved resolution of cell types and specific markers, including sub ision of the renal stroma and heterogeneity within the nephron progenitor population. Ligand-receptor interaction and pathway analysis reveals novel crosstalk between cellular compartments and associates new pathways with differentiation of nephron and ureteric epithelium cell types. We identify transcriptional congruence between the distal nephron and ureteric epithelium, showing that most markers previously used to identify ureteric epithelium are not specific. Together, this work improves our understanding of metanephric kidney development and provides a template to guide the regeneration of renal tissue.
Nephron Progenitor Cells
Publisher: Elsevier
Date: 2014
Direct reprogramming to human nephron progenitor-like cells using inducible piggyBac transposon expression of SNAI2-EYA1-SIX1.
Publisher: Elsevier BV
Date: 05-2019
The M1 subunit of ribonucleotide reductase refines mapping of genetic rearrangements at chromosome 11p15
Publisher: Elsevier BV
Date: 04-1992
DOI: 10.1016/0165-4608(92)90216-U
Abstract: We report the first use of the ribonucleotide reductase M1 subunit (RRM1) locus as a marker to assist in defining genetic rearrangements at 11p15. Our s le consisted of 21 Wilms' tumors from 18 patients, and one adrenal adenoma from a patient with Beckwith-Wiedemann syndrome, preexisting chromosome 11 maps being refined by the use of the RRM1 locus in all cases. Significantly, one Wilms' tumor showed loss of heterozygosity at the RRM1 locus only, whereas the adrenal adenoma showed a maintenance of heterozygosity at the RRM1 locus, loss having been previously demonstrated at the c-Ha-ras locus. The relevance of this finding to the location of one or more disease-associated loci at 11p15 is discussed.
Motile nephron progenitors can exit and re-enter the niche over time
Publisher: Elsevier BV
Date: 07-2017
Integrating single-cell genomics pipelines to discover mechanisms of stem cell differentiation
Publisher: Elsevier BV
Date: 12-2021
DOI: 10.1016/J.MOLMED.2021.09.006
Abstract: Pluripotent stem cells underpin a growing sector that leverages their differentiation potential for research, industry, and clinical applications. This review evaluates the landscape of methods in single-cell transcriptomics that are enabling accelerated discovery in stem cell science. We focus on strategies for scaling stem cell differentiation through multiplexed single-cell analyses, for evaluating molecular regulation of cell differentiation using new analysis algorithms, and methods for integration and projection analysis to classify and benchmark stem cell derivatives against in vivo cell types. By discussing the available methods, comparing their strengths, and illustrating strategies for developing integrated analysis pipelines, we provide user considerations to inform their implementation and interpretation.
Allelic loss on chromosome 11p is a less frequent event in bilateral than in unilateral Wilms' tumours
Publisher: Elsevier BV
Date: 1992
DOI: 10.1016/0959-8049(92)90027-Y
Abstract: Analyses to detect loss of heterozygosity (LOH) were performed at 11 polymorphic loci on chromosome 11 and, using a polymorphic CA repeat sequence in the WT1 gene, on a series of 39 tumours from 28 unilateral and 10 tumours from 6 bilateral Wilms' tumour (WT) patients. LOH was seen in 13 out of 35 patients including 12 out of 29 unilateral tumours, but only one of 10 bilateral tumours. This suggests that bilateral WT represents a subgroup of WT in which tumour initiating events less frequently involve LOH on chromosome 11 and that either epigenetic events, point mutations or another non-chromosome 11p locus are important in bilateral tumours. The observation of LOH in one WT but not another WT in a bilateral WT patient provides evidence that these tumours arising in the same patient are not monoclonal proliferations and most likely arise via different molecular pathways.
Molecular genetic evidence for common pathogenesis of childhood and adult Wilms' tumor
Publisher: Elsevier BV
Date: 03-1989
DOI: 10.1016/0165-4608(89)90172-6
Abstract: A case of Wilms' tumor in an adult is reported, showing, by restriction fragment length polymorphism analysis of somatic and tumor DNA, the loss of alleles from the short arm of chromosome 11. Loss of alleles in this region has previously been reported in childhood Wilms' tumor. The findings of this study indicate that adult Wilms' tumor and childhood Wilms' tumor may share a common pathogenic pathway. These results may also be useful in differentiating between Wilms' tumor and renal cell carcinoma or sarcoma in adults when the histologic findings are unclear.
Making a Kidney Organoid Using the Directed Differentiation of Human Pluripotent Stem Cells
Publisher: Springer New York
Date: 2017
DOI: 10.1007/978-1-4939-6949-4_14
Abstract: An organoid can be defined as a three-dimensional organ-like structure formed from organ-specific progenitor cells. Organ progenitor cells were empirically found to self-organize three-dimensional tissues when they were aggregated and cultivated in vitro. While this nature power of progenitor cells has an amazing potential to recreate artificial organs in vitro, there had been difficulty to apply this technology to human organs due to the inaccessibility to human progenitor cells until human-induced pluripotent stem cell (hiPSC) was invented by Takahashi and Yamanaka in 2007. As embryonic stem cells do, hiPSCs also have pluripotency to give rise to any organs/tissues cell types, including the kidney, via directed differentiation. Here, we provide a detailed protocol for generating kidney organoids using human pluripotent stem cells. The protocol differentiates human pluripotent stem cells into the posterior primitive streak. This is followed by the simultaneous induction of posterior and anterior intermediate mesoderm that are subsequently aggregated and undergo self-organization into the kidney organoid. Such kidney organoids are comprised of all anticipated kidney cell types including nephrons segmented into the glomerulus, proximal tubule, loop of Henle, and distal tubule as well as the collecting duct, endothelial network, and renal interstitium.
PAX2 activates WNT4 expression during mammalian kidney development
Publisher: Elsevier BV
Date: 05-2006
Direct Transcriptional Reprogramming of Adult Cells to Embryonic Nephron Progenitors
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 09-2013
Crim1 has cell-autonomous and paracrine roles during embryonic heart development
Publisher: Springer Science and Business Media LLC
Date: 29-01-2016
DOI: 10.1038/SREP19832
Abstract: The epicardium has a critical role during embryonic development, contributing epicardium-derived lineages to the heart, as well as providing regulatory and trophic signals necessary for myocardial development. Crim1 is a unique trans-membrane protein expressed by epicardial and epicardially-derived cells but its role in cardiogenesis is unknown. Using knockout mouse models, we observe that loss of Crim1 leads to congenital heart defects including epicardial defects and hypoplastic ventricular compact myocardium. Epicardium-restricted deletion of Crim1 results in increased epithelial-to-mesenchymal transition and invasion of the myocardium in vivo, and an increased migration of primary epicardial cells. Furthermore, Crim1 appears to be necessary for the proliferation of epicardium-derived cells (EPDCs) and for their subsequent differentiation into cardiac fibroblasts. It is also required for normal levels of cardiomyocyte proliferation and apoptosis, consistent with a role in regulating epicardium-derived trophic factors that act on the myocardium. Mechanistically, Crim1 may also modulate key developmentally expressed growth factors such as TGFβs, as changes in the downstream effectors phospho-SMAD2 and phospho-ERK1/2 are observed in the absence of Crim1 . Collectively, our data demonstrates that Crim1 is essential for cell-autonomous and paracrine aspects of heart development.
Comparative gene expression analysis of genital tubercle development reveals a putative appendicular Wnt7 network for the epidermal differentiation
Publisher: Elsevier BV
Date: 08-2010
Generating Kidney Organoids from Human Pluripotent Stem Cells Using Defined Conditions
Publisher: Springer US
Date: 2020
Establishment of metanephros transplantation in mice highlights contributions by both nephrectomy and pregnancy to developmental progression
Publisher: S. Karger AG
Date: 09-2005
DOI: 10.1159/000087939
Abstract: i Background: /i It has been demonstrated that embryonic kidneys (metanephroi) xenotransplanted into the omentum of adult recipients continue to develop and display immune protection due to their more naïve immune presentation. To date, this has been achieved using rat, pig and human metanephroi, with unilateral nephrectomy (UNX) of recipient rats a requisite of renal development. The aim of this study was to adapt this approach for use in mice and examine the parameters affecting successful onward development in this species. i Methods: /i Metanephroi at embryonic age (E) 13.5 were transplanted either onto the body wall, abdominal fat pads or omentum of recipient isogenic C57/Bl6 mice using either sutures or polyglycolic acid mesh. Having established greatest success with polyglycolic acid mesh on the body wall, E12.5 and 15.5 days metanephroi from C57/Bl6 mice were then transplanted onto the body wall of control (non-pregnant non-UNX), UNX or 12.5 days post-coitum pregnant isogenic recipients. After 7 days, implanted tissue was harvested and examined using histology and immunohistochemistry for markers of renal maturation. The mean number of S-shaped bodies and glomeruli per section were recorded and statistically analysed for significant differences between all recipient groups and untransplanted metanephroi. The degree of development was scored qualitatively. i Results: /i Transplanted E12.5 metanephroi developed S-shaped bodies and glomeruli in all recipient groups, although there were statistically higher numbers of S-shaped bodies in UNX (n = 2) and pregnant recipients (n = 9) than in control recipients (n = 4). Continued development, as indicated by mature vascularized glomeruli, was only observed in those E15.5 metanephroi transplanted into pregnant recipients (n = 11) with a 15.5-fold increase in S-shaped bodies and 4-fold increase in glomeruli compared with control transplants (n = 12). i Conclusions: /i We have successfully established metanephros transplantation in mice and demonstrated enhancement of onward development of E12.5 metanephroi in response to both pregnancy and UNX. Using E15.5 metanephroi, continued development only occurred in pregnant recipients, implying pregnancy provides an environment conducive to continued organogenesis. This murine assay, when coupled with transgenically-tagged strains of mice, will allow the investigation of the relative contribution of donor and recipient cells to this process.
Author Correction: Lin28 and let-7 regulate the timing of cessation of murine nephrogenesis
Publisher: Springer Science and Business Media LLC
Date: 09-03-2020
DOI: 10.1038/S41467-020-14944-3
Abstract: An amendment to this paper has been published and can be accessed via a link at the top of the paper.
A Toolbox to Characterize Human Induced Pluripotent Stem Cell–Derived Kidney Cell Types and Organoids
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 06-09-2019
Abstract: Kidney organoids generated from human induced pluripotent stem cells (iPSCs) show great potential for modeling kidney diseases and studying disease pathogenesis. However, the relative accuracy with which kidney organoids model normal morphogenesis, as well as the maturity and identity of the renal cell types they comprise, remain to be fully investigated. The authors describe the generation and validation of ten fluorescent CRISPR/Cas9 gene-edited iPSC reporter lines specifically designed for the visualization, isolation, and characterization of cell types and states within kidney organoids, and demonstrate the use of these lines for cellular isolation, time-lapse imaging, protocol optimization, and lineage-tracing applications. These tools offer promise for better understanding this model system and its congruence with human kidney morphogenesis. The generation of reporter lines for cell identity, lineage, and physiologic state has provided a powerful tool in advancing the dissection of mouse kidney morphogenesis at a molecular level. Although use of this approach is not an option for studying human development in vivo , its application in human induced pluripotent stem cells (iPSCs) is now feasible. We used CRISPR/Cas9 gene editing to generate ten fluorescence reporter iPSC lines designed to identify nephron progenitors, podocytes, proximal and distal nephron, and ureteric epithelium. Directed differentiation to kidney organoids was performed according to published protocols. Using immunofluorescence and live confocal microscopy, flow cytometry, and cell sorting techniques, we investigated organoid patterning and reporter expression characteristics. Each iPSC reporter line formed well patterned kidney organoids. All reporter lines showed congruence of endogenous gene and protein expression, enabling isolation and characterization of kidney cell types of interest. We also demonstrated successful application of reporter lines for time-lapse imaging and mouse transplantation experiments. We generated, validated, and applied a suite of fluorescence iPSC reporter lines for the study of morphogenesis within human kidney organoids. This fluorescent iPSC reporter toolbox enables the visualization and isolation of key populations in forming kidney organoids, facilitating a range of applications, including cellular isolation, time-lapse imaging, protocol optimization, and lineage-tracing approaches. These tools offer promise for enhancing our understanding of this model system and its correspondence with human kidney morphogenesis.
The GUDMAP database – an online resource for genitourinary research
Publisher: The Company of Biologists
Date: 07-2011
DOI: 10.1242/DEV.063594
Abstract: The GenitoUrinary Development Molecular Anatomy Project (GUDMAP) is an international consortium working to generate gene expression data and transgenic mice. GUDMAP includes data from large-scale in situ hybridisation screens (wholemount and section) and microarray gene expression data of microdissected, laser-captured and FACS-sorted components of the developing mouse genitourinary (GU) system. These expression data are annotated using a high-resolution anatomy ontology specific to the developing murine GU system. GUDMAP data are freely accessible at www.gudmap.org via easy-to-use interfaces. This curated, high-resolution dataset serves as a powerful resource for biologists, clinicians and bioinformaticians interested in the developing urogenital system. This paper gives ex les of how the data have been used to address problems in developmental biology and provides a primer for those wishing to use the database in their own research.
Analysis of early nephron patterning reveals a role for distal RV proliferation in fusion to the ureteric tip via a cap mesenchyme-derived connecting segment
Publisher: Elsevier BV
Date: 08-2009
DOI: 10.1016/J.YDBIO.2009.05.578
Abstract: While nephron formation is known to be initiated by a mesenchyme-to-epithelial transition of the cap mesenchyme to form a renal vesicle (RV), the subsequent patterning of the nephron and fusion with the ureteric component of the kidney to form a patent contiguous uriniferous tubule has not been fully characterized. Using dual section in situ hybridization (SISH)/immunohistochemistry (IHC) we have revealed distinct distal roximal patterning of Notch, BMP and Wnt pathway components within the RV stage nephron. Quantitation of mitoses and Cyclin D1 expression indicated that cell proliferation was higher in the distal RV, reflecting the differential developmental programs of the proximal and distal populations. A small number of RV genes were also expressed in the early connecting segment of the nephron. Dual ISH/IHC combined with serial section immunofluorescence and 3D reconstruction revealed that fusion occurs between the late RV and adjacent ureteric tip via a process that involves loss of the intervening ureteric epithelial basement membrane and insertion of cells expressing RV markers into the ureteric tip. Using Six2-eGFPCre x R26R-lacZ mice, we demonstrate that these cells are derived from the cap mesenchyme and not the ureteric epithelium. Hence, both nephron patterning and patency are evident at the late renal vesicle stage.
A protocol for the identification and validation of novel genetic causes of kidney disease.
Publisher: Springer Science and Business Media LLC
Date: 15-09-2015
Novel WT1 exon 9 mutation (D396Y) in a patient with early onset Denys Drash syndrome
Publisher: Hindawi Limited
Date: 04-2000
DOI: 10.1002/(SICI)1098-1004(200004)15:4<389::AID-HUMU29>3.0.CO;2-E
An illustrated anatomical ontology of the developing mouse lower urogenital tract
Publisher: The Company of Biologists
Date: 15-05-2015
DOI: 10.1242/DEV.117903
Abstract: Malformation of the urogenital tract represents a considerable paediatric burden, with many defects affecting the lower urinary tract (LUT), genital tubercle and associated structures. Understanding the molecular basis of such defects frequently draws on murine models. However, human anatomical terms do not always superimpose on the mouse, and the lack of accurate and standardised nomenclature is h ering the utility of such animal models. We previously developed an anatomical ontology for the murine urogenital system. Here, we present a comprehensive update of this ontology pertaining to mouse LUT, genital tubercle and associated reproductive structures (E10.5 to adult). Ontology changes were based on recently published insights into the cellular and gross anatomy of these structures, and on new analyses of epithelial cell types present in the pelvic urethra and regions of the bladder. Ontology changes include new structures, tissue layers and cell types within the LUT, external genitalia and lower reproductive structures. Representative illustrations, detailed text descriptions and molecular markers that selectively label muscle, nerves/ganglia and epithelia of the lower urogenital system are also presented. The revised ontology will be an important tool for researchers studying urogenital development/malformation in mouse models and will improve our capacity to appropriately interpret these with respect to the human situation.
An integrated pipeline for the multidimensional analysis of branching morphogenesis
Publisher: Springer Science and Business Media LLC
Date: 20-11-2014
Abstract: Developmental branching morphogenesis establishes organ architecture, and it is driven by iterative interactions between epithelial and mesenchymal progenitor cell populations. We describe an approach for analyzing this interaction and how it contributes to organ development. After initial in vivo cell labeling with the nucleoside analog 5-ethynyl-2'-deoxyuridine (EdU) and tissue-specific antibodies, optical projection tomography (OPT) and confocal microscopy are used to image the developing organ. These imaging data then inform a second analysis phase that quantifies (using Imaris and Tree Surveyor software), models and integrates these events at a cell and tissue level in 3D space and across developmental time. The protocol establishes a benchmark for assessing the impact of genetic change or fetal environment on organogenesis that does not rely on ex vivo organ culture or section-based reconstruction. By using this approach, examination of two developmental stages for an organ such as the kidney can be undertaken by a postdoctoral-level researcher in 6 weeks, with a full developmental analysis in mouse achievable in 5 months.
Correction: Single cell analysis of the developing mouse kidney provides deeper insight into marker gene expression and ligand-receptor crosstalk (doi:10.1242/dev.178673)
Publisher: The Company of Biologists
Date: 07-2019
DOI: 10.1242/DEV.182162
Enhanced metanephric specification to functional proximal tubule enables toxicity screening and infectious disease modelling in kidney organoids
Publisher: Springer Science and Business Media LLC
Date: 08-10-2022
DOI: 10.1038/S41467-022-33623-Z
Abstract: While pluripotent stem cell-derived kidney organoids are now being used to model renal disease, the proximal nephron remains immature with limited evidence for key functional solute channels. This may reflect early mispatterning of the nephrogenic mesenchyme and/or insufficient maturation. Here we show that enhanced specification to metanephric nephron progenitors results in elongated and radially aligned proximalised nephrons with distinct S1 - S3 proximal tubule cell types. Such PT-enhanced organoids possess improved albumin and organic cation uptake, appropriate KIM-1 upregulation in response to cisplatin, and improved expression of SARS-CoV-2 entry factors resulting in increased viral replication. The striking proximo-distal orientation of nephrons resulted from localized WNT antagonism originating from the organoid stromal core. PT-enhanced organoids represent an improved model to study inherited and acquired proximal tubular disease as well as drug and viral responses.
M2 macrophage polarisation is associated with alveolar formation during postnatal lung development
Publisher: Springer Science and Business Media LLC
Date: 05-04-2013
Abstract: Macrophages are traditionally associated with inflammation and host defence, however a greater understanding of macrophage heterogeneity is revealing their essential roles in non-immune functions such as development, homeostasis and regeneration. In organs including the brain, kidney, mammary gland and pancreas, macrophages reside in large numbers and provide essential regulatory functions that shape organ development and maturation. However, the role of macrophages in lung development and the potential implications of macrophage modulation in the promotion of lung maturation have not yet been ascertained. Embryonic day (E)12.5 mouse lungs were cultured as explants and macrophages associated with branching morphogenesis were visualised by wholemount immunofluorescence microscopy. Postnatal lung development and the correlation with macrophage number and phenotype were examined using Colony-stimulating factor-1 receptor-enhanced green fluorescent protein ( Csf1r -EGFP) reporter mice. Structural histological examination was complemented with whole-body plethysmography assessment of postnatal lung functional maturation over time. Flow cytometry, real-time (q)PCR and immunofluorescence microscopy were performed to characterise macrophage number, phenotype and localisation in the lung during postnatal development. To assess the impact of developmental macrophage modulation, CSF-1 was administered to neonatal mice at postnatal day (P)1, 2 and 3, and lung macrophage number and phenotype were assessed at P5. EGFP transgene expression and in situ hybridisation was performed to assess CSF-1R location in the developing lung. Macrophages in embryonic lungs were abundant and densely located within branch points during branching morphogenesis. During postnatal development, structural and functional maturation of the lung was associated with an increase in lung macrophage number. In particular, the period of alveolarisation from P14-21 was associated with increased number of Csf1r -EGFP+ macrophages and upregulated expression of Arginase 1 (Arg1), Mannose receptor 1 (Mrc1) and Chemokine C-C motif ligand 17 (Ccl1 7), indicative of an M2 or tissue remodelling macrophage phenotype. Administration of CSF-1 to neonatal mice increased trophic macrophages during development and was associated with increased expression of the M2-associated gene Found in inflammatory zone (Fizz)1 and the growth regulator Insulin-like growth factor (Igf)1 . The effects of CSF-1 were identified as macrophage-mediated, as the CSF-1R was found to be exclusively expressed on interstitial myeloid cells. This study identifies the presence of CSF-1R+ M2-polarised macrophages localising to sites of branching morphogenesis and increasing in number during the alveolarisation stage of normal lung development. Improved understanding of the role of macrophages in lung developmental regulation has clinical relevance for addressing neonatal inflammatory perturbation of development and highlights macrophage modulation as a potential intervention to promote lung development.
Simultaneous reprogramming and gene editing of human fibroblasts
Publisher: Springer Science and Business Media LLC
Date: 05-04-2018
Kidney micro-organoids in suspension culture as a scalable source of human pluripotent stem cell-derived kidney cells
Publisher: The Company of Biologists
Date: 03-2019
DOI: 10.1242/DEV.172361
Abstract: Kidney organoids have potential uses in disease modelling, drug screening and regenerative medicine. However, novel cost-effective techniques are needed to enable scaled-up production of kidney cell types in vitro. We describe here a modified suspension culture method for the generation of kidney micro-organoids from human pluripotent stem cells. Optimisation of differentiation conditions allowed the formation of micro-organoids, each containing six to ten nephrons that were surrounded by endothelial and stromal populations. Single cell transcriptional profiling confirmed the presence and transcriptional equivalence of all anticipated renal cell types consistent with a previous organoid culture method. This suspension culture micro-organoid methodology resulted in a three- to fourfold increase in final cell yield compared with static culture, thereby representing an economical approach to the production of kidney cells for various biological applications.
3D organoid-derived human glomeruli for personalised podocyte disease modelling and drug screening.
Publisher: Springer Science and Business Media LLC
Date: 04-12-2018
DOI: 10.1038/S41467-018-07594-Z
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.
Global Quantification of Tissue Dynamics in the Developing Mouse Kidney
Publisher: Elsevier BV
Date: 04-2014
DOI: 10.1016/J.DEVCEL.2014.02.017
Abstract: Although kidneys of equal size can vary 10-fold in nephron number at birth, discovering what regulates such variation has been h ered by a lack of quantitative parameters defining kidney development. Here we report a comprehensive, quantitative, multiscale analysis of mammalian kidney development in which we measure changes in cell number, compartment volumes, and cellular dynamics across the entirety of organogenesis, focusing on two key nephrogenic progenitor populations: the ureteric epithelium and the cap mesenchyme. In doing so, we describe a discontinuous developmental program governed by dynamic changes in interactions between these key cellular populations occurring within a previously unappreciated structurally stereotypic organ architecture. We also illustrate the application of this approach to the detection of a subtle mutant phenotype. This baseline program of kidney morphogenesis provides a framework for assessing genetic and environmental developmental perturbation and will serve as a gold standard for the analysis of other organs.
Replacing renal function using bioengineered tissues
Publisher: Springer Science and Business Media LLC
Date: 17-05-2023
ROBO2 restricts the nephrogenic field and regulates Wolffian duct–nephrogenic cord separation
Publisher: Elsevier BV
Date: 08-2015
DOI: 10.1016/J.YDBIO.2015.05.023
Abstract: ROBO2 plays a key role in regulating ureteric bud (UB) formation in the embryo, with mutations in humans and mice leading to supernumerary kidneys. Previous studies have established that the number and position of UB outgrowths is determined by the domain of metanephric mesenchymal Gdnf expression, which is expanded anteriorly in Robo2 mouse mutants. To clarify how this phenotype arises, we used high-resolution 3D imaging to reveal an increase in the number of nephrogenic cord cells, leading to extension of the metanephric mesenchyme field in Robo2-null mouse embryos. Ex vivo experiments suggested a dependence of this effect on proliferative signals from the Wolffian duct. Loss of Robo2 resulted in a failure of the normal separation of the mesenchyme from the Wolffian duct/ureteric epithelium, suggesting that aberrant juxtaposition of these two compartments in Robo2-null mice exposes the mesenchyme to abnormally high levels of proliferative stimuli. Our data suggest a new model in which SLIT-ROBO signalling acts not by attenuating Gdnf expression or activity, but instead by limiting epithelial/mesenchymal interactions in the nascent metanephros and restricting the extent of the nephrogenic field. These insights illuminate the aetiology of multiplex kidney formation in human in iduals with ROBO2 mutations.
Cellular extrusion bioprinting improves kidney organoid reproducibility and conformation
Publisher: Springer Science and Business Media LLC
Date: 23-11-2021
Generating a self-organizing kidney from pluripotent cells
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 04-2015
Branching morphogenesis in the developing kidney is not impacted by nephron formation or integration
Publisher: eLife Sciences Publications, Ltd
Date: 31-07-2018
DOI: 10.7554/ELIFE.38992
Abstract: Branching morphogenesis of the ureteric bud is integral to kidney development establishing the collecting ducts of the adult organ and driving organ expansion via peripheral interactions with nephron progenitor cells. A recent study suggested that termination of tip branching within the developing kidney involved stochastic exhaustion in response to nephron formation, with such a termination event representing a unifying developmental process evident in many organs. To examine this possibility, we have profiled the impact of nephron formation and maturation on elaboration of the ureteric bud during mouse kidney development. We find a distinct absence of random branch termination events within the kidney or evidence that nephrogenesis impacts the branching program or cell proliferation in either tip or progenitor cell niches. Instead, organogenesis proceeds in a manner indifferent to the development of these structures. Hence, stochastic cessation of branching is not a unifying developmental feature in all branching organs.
Advances in our understanding of genetic kidney disease using kidney organoids
Publisher: Springer Science and Business Media LLC
Date: 07-05-2020
DOI: 10.1007/S00467-019-04259-X
Abstract: A significant proportion of kidney disease presenting in childhood is likely genetic in origin with a growing number of genes implicated in its development. However, many children may have changes in previously undescribed or unrecognised genes. The recent development of methods for generating human kidney organoids from human pluripotent stem cells has the potential to substantially change the rate of diagnosis and the development of new treatments for some forms of genetic kidney disease. In this review, we discuss how accurately a kidney organoid models the human kidney, identifying the strengths and weaknesses of these potentially patient-derived models of renal disease.
Closing the circle: from organoids back to development.
Publisher: The Company of Biologists
Date: 15-03-2016
DOI: 10.1242/DEV.136150
Abstract: Summary: This Editorial looks at the emerging field of in vitro organogenesis and discusses how organoid technology can be applied to better understand developmental processes.
A high-resolution anatomical ontology of the developing murine genitourinary tract
Publisher: Elsevier BV
Date: 06-2007
Generation of kidney organoids from human pluripotent stem cells
Publisher: Springer Science and Business Media LLC
Date: 18-08-2016
An RNA recognition motif in Wilms' tumour protein (WT1) revealed by structural modelling.
Publisher: Springer Science and Business Media LLC
Date: 03-1996
DOI: 10.1038/NG0396-329
Methylation and p16: Suppressing the suppressor
Publisher: Springer Science and Business Media LLC
Date: 07-1995
DOI: 10.1038/NM0795-633
Abstract: The role of point-of-care testing (POCT) out of hospital, especially in home care and ambulatory care settings, is an issue meriting further research. We reviewed studies reporting cardiovascular events as a result of the implementation of B-type natriuretic peptide or N-terminal pro B-type natriuretic peptide POCT (BNP/NT-proBNP POCT) for heart disease patients in the settings. Articles were searched via a PubMed engine until May 30, 2020. In total, six studies were selected. Three studies involving ambulatory care used the POCT to refer patients with suspected heart diseases to a specialist. The other three used the tests in home care to monitor patients with heart failure. In ambulatory care, the randomized controlled trials, in which referrals were made to a specialist, showed that the group using POCT had significantly fewer cardiovascular outcomes, such as hospitalizations and deaths, than the non-use group. In home care, adverse outcomes were predicted from changes in BNP levels. In most studies, the use of BNP/NT-proBNP POCT in home care and ambulatory care settings demonstrated favorable results regarding the cardiovascular outcomes. The utility of POCT in the settings is suggested, while more investigations are required.
Enhanced metanephric specification to functional proximal tubule enables toxicity screening and infectious disease modelling in kidney organoids
Publisher: Cold Spring Harbor Laboratory
Date: 15-10-2021
DOI: 10.1101/2021.10.14.464320
Abstract: While pluripotent stem cell-derived kidney organoids are now being used to model renal disease, the proximal nephron remains immature with limited evidence for key functional solute channels. This may reflect early mispatterning of the nephrogenic mesenchyme and/or insufficient maturation. Here we show that enhanced specification to metanephric nephron progenitors results in elongated and radially aligned proximalised nephrons with distinct S1 - S3 proximal tubule cell types. Such PT-enhanced organoids possess improved albumin and organic cation uptake, appropriate KIM-1 upregulation in response to cisplatin, and improved expression of SARS-CoV-2 entry factors resulting in increased viral replication. The striking proximo-distal orientation of nephrons resulted from localized WNT antagonism originating from the organoid stromal core. PT-enhanced organoids represent an improved model to study inherited and acquired proximal tubular disease as well as drug and viral responses.
In ovo electroporation of Crim1 in the developing chick spinal cord
Publisher: Wiley
Date: 30-12-2003
DOI: 10.1002/DVDY.10204
Abstract: The novel mammalian gene Crim1 encodes a transmembrane bound protein with similarity to the secreted bone morphogenetic protein (BMP) antagonists, vertebrate Chordin, and its Drosophila homologue short gastrulation. Crim1 is expressed in the neural tube in mouse in a restricted pattern, but its function in central nervous system development is largely unknown. We isolated the chicken Crim1 orthologue and analyzed its expression in the developing neural tube. Chicken CRIM1 shares strong homology to human/mouse CRIM1 and C. elegans CRIM1-like proteins. Crim1 is expressed in a similar but not identical pattern to that in the developing spinal cord of mouse, including the notochord, floor plate, motor neurons, and the roof plate. Unlike follistatin, a secreted inhibitor of BMPs, in ovo electroporation of CRIM1, as a full-length transmembrane bound or secreted ectodomain was not sufficient to disrupt early patterning of the neural tube. However, ectodomain CRIM1 overexpression leads to an approximate 50% decrease in populations of specific ventral neuronal populations, including ISL-1(+) motor neurons, CHX-10(+) V1, and EN-1(+) V2 interneurons.
Improving our resolution of kidney morphogenesis across time and space
Publisher: Elsevier BV
Date: 06-2015
DOI: 10.1016/J.GDE.2015.03.001
Abstract: As with many mammalian organs, size and cellular complexity represent considerable challenges to the comprehensive analysis of kidney organogenesis. Traditional analyses in the mouse have revealed early patterning events and spatial cellular relationships. However, an understanding of later events is lacking. The generation of a comprehensive temporospatial atlas of gene expression during kidney development has facilitated advances in lineage definition, as well as selective compartment ablation. Advances in quantitative and dynamic imaging have allowed comprehensive analyses at the level of organ, component tissue and cell across kidney organogenesis. Such approaches will enhance our understanding of the links between kidney development and final postnatal organ function. The final frontier will be translating this understanding to outcomes for renal disease in humans.
A spatially-averaged mathematical model of kidney branching morphogenesis
Publisher: Elsevier BV
Date: 08-2015
DOI: 10.1016/J.JTBI.2015.04.015
Abstract: Kidney development is initiated by the outgrowth of an epithelial ureteric bud into a population of mesenchymal cells. Reciprocal morphogenetic responses between these two populations generate a highly branched epithelial ureteric tree with the mesenchyme differentiating into nephrons, the functional units of the kidney. While we understand some of the mechanisms involved, current knowledge fails to explain the variability of organ sizes and nephron endowment in mice and humans. Here we present a spatially-averaged mathematical model of kidney morphogenesis in which the growth of the two key populations is described by a system of time-dependant ordinary differential equations. We assume that branching is symmetric and is invoked when the number of epithelial cells per tip reaches a threshold value. This process continues until the number of mesenchymal cells falls below a critical value that triggers cessation of branching. The mathematical model and its predictions are validated against experimentally quantified C57Bl6 mouse embryonic kidneys. Numerical simulations are performed to determine how the final number of branches changes as key system parameters are varied (such as the growth rate of tip cells, mesenchyme cells, or component cell population exit rate). Our results predict that the developing kidney responds differently to loss of cap and tip cells. They also indicate that the final number of kidney branches is less sensitive to changes in the growth rate of the ureteric tip cells than to changes in the growth rate of the mesenchymal cells. By inference, increasing the growth rate of mesenchymal cells should maximise branch number. Our model also provides a framework for predicting the branching outcome when ureteric tip or mesenchyme cells change behaviour in response to different genetic or environmental developmental stresses.
Reporter‐based fate mapping in human kidney organoids confirms nephron lineage relationships and reveals synchronous nephron formation
Publisher: EMBO
Date: 12-03-2019
Involvement of Islet-2 in the Slit signaling for axonal branching and defasciculation of the sensory neurons in embryonic zebrafish
Publisher: Elsevier BV
Date: 04-2004
A strategy for generating kidney organoids: Recapitulating the development in human pluripotent stem cells
Publisher: Elsevier BV
Date: 12-2016
Delivering on the promise of human stem-cell research - What are the real barriers?
Publisher: EMBO
Date: 03-11-2006
The Receptor Tyrosine Kinase Regulator Sprouty1 Is a Target of the Tumor Suppressor WT1 and Important for Kidney Development
Publisher: Elsevier BV
Date: 10-2003
Generating kidney tissue from pluripotent stem cells
Publisher: Springer Science and Business Media LLC
Date: 18-07-2016
DOI: 10.1038/CDDISCOVERY.2016.53
Abstract: With the isolation of human pluripotent stem cells came the possibility of generating specific cell types for regenerative medicine. This has required the development of protocols for directed differentiation into many distinct cell types. One of the more complicated tissue types to recreate is the kidney. Here we review recent progress towards the recreation of not only specific kidney cell types but complex kidney organoids, models of the developing human organ, in vitro . We will also discuss potential short and long term applications of these approaches.
Evaluation of biomarkers for in vitro prediction of drug-induced nephrotoxicity: comparison of HK-2, immortalized human proximal tubule epithelial, and primary cultures of human proximal tubular cells.
Publisher: Wiley
Date: 15-05-2015
DOI: 10.1002/PRP2.148
Clinical-Grade Isolated Human Kidney Perivascular Stromal Cells as an Organotypic Cell Source for Kidney Regenerative Medicine
Publisher: Oxford University Press (OUP)
Date: 20-09-2016
Abstract: Mesenchymal stromal cells (MSCs) are immunomodulatory and tissue homeostatic cells that have shown beneficial effects in kidney diseases and transplantation. Perivascular stromal cells (PSCs) identified within several different organs share characteristics of bone marrow-derived MSCs (BM-MSCs). These PSCs may also possess tissue-specific properties and play a role in local tissue homeostasis. We hypothesized that human kidney-derived PSCs (hkPSCs) would elicit improved kidney repair in comparison with BM-MSCs. Here we introduce a novel, clinical-grade isolation method of hkPSCs from cadaveric kidneys by enriching for the perivascular marker, NG2. hkPSCs show strong transcriptional similarities to BM-MSCs but also show organotypic expression signatures, including the HoxD10 and HoxD11 nephrogenic transcription factors. Comparable to BM-MSCs, hkPSCs showed immunosuppressive potential and, when cocultured with endothelial cells, vascular plexus formation was supported, which was specifically in the hkPSCs accompanied by an increased NG2 expression. hkPSCs did not undergo myofibroblast transformation after exposure to transforming growth factor-β, further corroborating their potential regulatory role in tissue homeostasis. This was further supported by the observation that hkPSCs induced accelerated repair in a tubular epithelial wound scratch assay, which was mediated through hepatocyte growth factor release. In vivo, in a neonatal kidney injection model, hkPSCs reintegrated and survived in the interstitial compartment, whereas BM-MSCs did not show this potential. Moreover, hkPSCs gave protection against the development of acute kidney injury in vivo in a model of rhabdomyolysis-mediated nephrotoxicity. Overall, this suggests a superior therapeutic potential for the use of hkPSCs and their secretome in the treatment of kidney diseases.
Generating Kidney from Stem Cells
Publisher: Annual Reviews
Date: 10-02-2019
DOI: 10.1146/ANNUREV-PHYSIOL-020518-114331
Abstract: Human kidney tissue can now be generated via the directed differentiation of human pluripotent stem cells. This advance is anticipated to facilitate the modeling of human kidney diseases, provide platforms for nephrotoxicity screening, enable cellular therapy, and potentially generate tissue for renal replacement. All such applications will rely upon the accuracy and reliability of the model and the capacity for stem cell–derived kidney tissue to recapitulate both normal and diseased states. In this review, we discuss the models available, how well they recapitulate the human kidney, and how far we are from application of these cells for use in cellular therapies.
The Life Cycle of the Nephron Progenitor
Publisher: Elsevier BV
Date: 10-2015
DOI: 10.1016/J.DEVCEL.2015.09.023
Abstract: Although we know that mesenchymal progenitors give rise to nephrons in the kidney, how they balance self-renewal versus differentiation is still unclear. In this issue of Developmental Cell, Chen et al. (2015) show that nephron progenitors age, but not necessarily irreversibly: old progenitors can be "rejuvenated" by a young crowd.
PRODUCTION AND METABOLIC-CLEARANCE OF ANGIOTENSINOGEN IN CONSCIOUS RATS AS MEASURED BY STEADY-STATE ISOTOPE-DILUTION
Publisher: Bioscientifica
Date: 03-1987
Abstract: Previous studies on the hormonal regulation of hepatic angiotensinogen relied on in-vitro liver preparations and on the measurement of changes in plasma concentration. In this study 125 I-labelled angiotensinogen was used to measure simultaneously the production rate (PR) and metabolic clearance rate (MCR) in conscious rats by the constant-rate infusion and single-injection methods. Male rats received daily s.c. injections of isotonic saline (as control), 1 mg corticosterone acetate (CA), 25 μg 17β-oestradiol benzoate (OB) or 20 μg thyroxine (T 4 ) per 100 g body weight. On day 7 of treatment 125 I-labelled angiotensinogen was infused into a jugular vein at a rate of 1 μl/h by osmotic minipumps and blood s les taken 4, 5 and 6 days later. The PR of angiotensinogen increased from 576 ± 28 ( s.e.m. n = 9) to 954 ± 63 ( n = 9), 1010±84 ( n = 9) and 2359±150 ( n = 10) μg/h per kg following treatment with CA, OB and T 4 respectively. In contrast, the PR of rat albumin did not change significantly from 218 ± 8 ( n = 7) mg/h per kg. All three hormones increased MCR from 13 ± 1 ( n = 17) ml/h per kg to 17± 1 ( n = 9), 18 ± 2 ( n = 9) and 27 ± 2 ( n = 9) ml/h per kg for CA, OB and T 4 respectively. Single-injection experiments on five rats showed angiotensinogen to be distributed into three compartments with a half-time of disappearance of 4·4 ± 1 min, 116±11 min and 13·1 ±2·6 h. It was concluded that the production of angiotensinogen in vivo is at least tenfold higher than the reported in-vitro rates, that the clearance of angiotensinogen is under hormonal regulation and that angiotensinogen is distributed between at least three compartments. J. Endocr. (1987) 112, 391–397
Movement through slits: Cellular migration via the Slit family
Publisher: Wiley
Date: 20-12-2002
DOI: 10.1002/BIES.10199
Abstract: First isolated in the fly and now characterised in vertebrates, the Slit proteins have emerged as pivotal components controlling the guidance of axonal growth cones and the directional migration of neuronal precursors. As well as extensive expression during development of the central nervous system (CNS), the Slit proteins exhibit a striking array of expression sites in non-neuronal tissues, including the urogenital system, limb primordia and developing eye. Zebrafish Slit has been shown to mediate mesodermal migration during gastrulation, while Drosophila slit guides the migration of mesodermal cells during myogenesis. This suggests that the actions of these secreted molecules are not simply confined to the sphere of CNS development, but rather act in a more general fashion during development and throughout the lifetime of an organism. This review focuses on the non-neuronal activities of Slit proteins, highlighting a common role for the Slit family in cellular migration.
c-Ha-ras-1 alleles in bladder cancer, Wilms' tumour and malignant melanoma
Publisher: Springer Science and Business Media LLC
Date: 02-1988
DOI: 10.1007/BF00278178
Abstract: Cyclooxygenase-2 (COX-2) is up-regulated in most high-grade gliomas, and high COX-2 expression is associated with aggressive character and poor prognosis. However, the effect of COX-2 in human glioma cell lines is not well known. This study examined the effect of several stimuli, including interleukin-1beta (IL-1beta) and carcinogens, on COX-2 induction in normal astrocyte cells and human glioma cell lines U87MG, A172, and T98G. IL-1beta-induced COX-2 expression strongly at both protein and messenger ribonucleic acid levels in only the U87MG cells of the glioma cell lines. Furthermore, carcinogen induced COX-2 expression. Similar findings were also observed in normal human astrocyte cells. The U87MG glioma cell line is a good model for COX-2 induction in glioma cell lines.
Single-cell analysis reveals congruence between kidney organoids and human fetal kidney
Publisher: Springer Science and Business Media LLC
Date: 23-01-2019
Knockdown of zebrafish crim1 results in a bent tail phenotype with defects in somite and vascular development
Publisher: Elsevier BV
Date: 04-2006
DOI: 10.1016/J.MOD.2006.01.003
Abstract: The Crim1 gene encodes a transmembrane protein containing six cysteine-rich repeats similar to those found in the BMP antagonist, chordin (chd). To investigate its physiological role, zebrafish crim1 was cloned and shown to be both maternally and zygotically expressed during zebrafish development in sites including the vasculature, intermediate cell mass, notochord, and otic vesicle. Bent or hooked tails with U-shaped somites were observed in 85% of morphants from 12 hpf. This was accompanied by a loss of muscle pioneer cells. While morpholino knockdown of crim1 showed some evidence of ventralisation, including expansion of the intermediate cell mass (ICM), reduction in head size bent tails and disruption to the somites and notochord, this did not mimic the classically ventralised phenotype, as assessed by the pattern of expression of the dorsal markers chordin, otx2 and the ventral markers eve1, pax2.1, tal1 and gata1 between 75% epiboly and six-somites. From 24 hpf, morphants displayed an expansion of the ventral mesoderm-derived ICM, as evidenced by expansion of tal1, lmo2 and crim1 itself. Analysis of the crim1 morphant phenotype in Tg(fli:EGFP) fish showed a clear reduction in the endothelial cells forming the intersegmental vessels and a loss of the dorsal longitudinal anastomotic vessel (DLAV). Hence, the primary role of zebrafish crim1 is likely to be the regulation of somitic and vascular development.
Neonatal calyceal dilation and renal fibrosis resulting from loss of Adamts-1 in mouse kidney is due to a developmental dysgenesis
Publisher: Oxford University Press (OUP)
Date: 22-12-2005
DOI: 10.1093/NDT/GFH603
Abstract: A disintegrin and metalloproteinase with thrombospondin motifs 1, Adamts-1, is important for the development and function of the kidney. Mice lacking this protein present with renal lesions comprising enlarged calyces, and reduced cortex and medulla layers. Our current findings are consistent with the defect occurring due to a developmental dysgenesis. We generated Adamts-1 null mice, and further investigated their kidney phenotype in a time course study ranging from E18.5 to 12 months of age. Immunohistochemistry was used to assess the localization of type IV collagen, TGF-beta and F4/80-positive macrophages in the kidneys of Adamts-1 null mice compared to wild-type control animals. The expression of Adamts-1 mRNA was determined in metanephric kidney explants by in situ hybridization. Adamts-1 null mice have a gross kidney defect. At day 18.5 of gestation, the Adamts-1 null kidney has a normal appearance but at birth when the kidney begins to function, the defect becomes evident. During development of the kidney Adamts-1 expression was specifically detected in the developing loops of Henle, as well as in the proximal and distal convoluted tubules. Expression was not detected in the ureter, ureteric bud or its derivatives as had been previously suggested. At 6 months and 1 year of age, the Adamts-1 null mice displayed interstitial fibrosis in the cortical and medullary regions of the kidney. At 1 year of age, the Adamts-1 null mice displayed mild interstitial matrix expansion associated with increased collagen type IV expression, without apparent tubular dilatation, compared to wild-type animals. Immunohistochemical analysis demonstrated TGF-beta protein localized to infiltrating macrophages and glomeruli of Adamts-1 null mice. Adamts-1 is required for the normal development of the kidney. The defect observed in its absence results from a dysgenic malformation affecting the medulla that becomes apparent at birth, once the kidneys start to function.
The BCL‐2 family member BID plays a role during embryonic development in addition to its BH3‐only protein function by acting in parallel to BAX, BAK and BOK
Publisher: EMBO
Date: 27-06-2022
Defining the Molecular Character of the Developing and Adult Kidney Podocyte
Publisher: Public Library of Science (PLoS)
Date: 08-09-2011
GUDMAP
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 04-2008
Growing Kidney Tissue from Stem Cells: How Far from Party Trick to Medical Application?
Publisher: Elsevier BV
Date: 06-2016
ENHANCED EXPRESSION OF INSULIN-LIKE GROWTH FACTOR-II IS NOT A NECESSARY EVENT IN WILMS-TUMOR PROGRESSION
Publisher: Oxford University Press (OUP)
Date: 1987
Abstract: Wilms' tumour (WT) is a paediatric kidney tumour arising from the embryonal metanephrogenic blastema. Recent reports suggest that the expression of messenger RNA (mRNA) for insulin-like growth factor II (IGFII) is elevated in WT. Total cytoplasmic RNA was extracted from 11 sporadic WTs and analysed for IGFII mRNA using dot-blot hybridization. The level of IGFII mRNA expression varied greatly and not all tumours displayed enhanced IGFII expression. Two successive WT xenografts were established in nude mice. The original WT and first passage xenograft showed a blastematous histology, while the second passage xenograft showed epithelioid differentiation and tubule formation. Analysis of the expression patterns of these xenografts showed elevated IGFII expression in the primary undifferentiated tumour and the second differentiated xenograft, while the first undifferentiated xenograft failed to exhibit enhanced IGFII expression. These data show that elevated IGFII mRNA is not an essential component of the progression of WT and that WT tumourigenicity is independent of the level of IGFII expression. Therefore, IGFII overexpression in WT is most likely a tumour epiphenomenon.
WT1: what has the last decade told us?
Publisher: Wiley
Date: 29-03-1999
DOI: 10.1002/(SICI)1521-1878(199903)21:3<191::AID-BIES3>3.0.CO;2-8
The origin of the mammalian kidney: implications for recreating the kidney in vitro
Publisher: The Company of Biologists
Date: 06-2015
DOI: 10.1242/DEV.104802
Abstract: The mammalian kidney, the metanephros, is a mesodermal organ classically regarded as arising from the intermediate mesoderm (IM). Indeed, both the ureteric bud (UB), which gives rise to the ureter and the collecting ducts, and the metanephric mesenchyme (MM), which forms the rest of the kidney, derive from the IM. Based on an understanding of the signalling molecules crucial for IM patterning and kidney morphogenesis, several studies have now generated UB or MM, or both, in vitro via the directed differentiation of human pluripotent stem cells. Although these results support the IM origin of the UB and the MM, they challenge the simplistic view of a common progenitor for these two populations, prompting a reanalysis of early patterning events within the IM. Here, we review our understanding of the origin of the UB and the MM in mouse, and discuss how this impacts on kidney regeneration strategies and furthers our understanding of human development.
Spatial gene expression in the T-stage mouse metanephros
Publisher: Elsevier BV
Date: 10-2006
DOI: 10.1016/J.MODGEP.2006.02.001
Abstract: The E11.5 mouse metanephros is comprised of a T-stage ureteric epithelial tubule sub- ided into tip and trunk cells surrounded by metanephric mesenchyme (MM). Tip cells are induced to undergo branching morphogenesis by the MM. In contrast, signals within the mesenchyme surrounding the trunk prevent ectopic branching of this region. In order to identify novel genes involved in the molecular regulation of branching morphogenesis we compared the gene expression profiles of isolated tip, trunk and MM cells using Compugen mouse long oligo microarrays. We identified genes enriched in the tip epithelium, sim-1, Arg2, Tacstd1, Crlf-1 and BMP7 genes enriched in the trunk epithelium, Innp1, Itm2b, Mkrn1, SPARC, Emu2 and Gsta3 and genes spatially restricted to the mesenchyme surrounding the trunk, CSPG2 and CV-2, with overlapping and complimentary expression to BMP4, respectively. This study has identified genes spatially expressed in regions of the developing kidney involved in branching morphogenesis, nephrogenesis and the development of the collecting duct system, calyces, renal pelvis and ureter.
Identifying the molecular phenotype of renal progenitor cells
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 09-2004
Defining Kidney Biology to Understand Renal Disease
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 04-2014
DOI: 10.2215/CJN.10851013
Modulation of DNA Binding Specificity by Alternative Splicing of the Wilms Tumor wt1 Gene Transcript
Publisher: American Association for the Advancement of Science (AAAS)
Date: 10-07-1992
Abstract: The technique of whole-genome polymerase chain reaction was used to study the DNA binding properties of the product of the wt1 gene. The zinc finger region of this gene is alternatively spliced such that the major transcript encodes a protein with three extra amino acids between the third and fourth fingers. The minor form of the protein binds specifically to DNA. It is now shown that the major form of wt1 messenger RNA encodes a protein that binds to DNA with a specificity that differs from that of the minor form. Therefore, alternative splicing within the DNA binding domain of a transcription factor can generate proteins with distinct DNA binding specificities and probably different physiological targets.
Prenatal hypoxia leads to hypertension, renal renin-angiotensin system activation and exacerbates salt-induced pathology in a sex-specific manner
Publisher: Springer Science and Business Media LLC
Date: 15-08-2017
DOI: 10.1038/S41598-017-08365-4
Abstract: Prenatal hypoxia is associated with growth restriction and adverse cardiovascular outcomes. Here, we describe renal and cardiovascular outcomes in ageing mouse offspring prenatally exposed to hypoxia (12% O 2 ) from embryonic day 14.5 until birth. At 12 months of age, both male and female offspring exposed to prenatal hypoxia had elevated mean arterial pressure. Glomerular number was reduced by 25% in hypoxia-exposed male, but not female, offspring and this was associated with increased urinary albumin excretion, glomerular hypertrophy and renal fibrosis. Hypoxia-exposed offspring of both sexes were more susceptible to salt-induced cardiac fibrosis, however, renal fibrosis was exacerbated by high salt in males only. In male but not female hypoxia-exposed offspring, renal renin mRNA was increased at weaning. By 12 months, renal renin mRNA expression and concentrations were elevated in both sexes. mRNA expression of At 1a R was also elevated in male hypoxia-exposed offspring at 12 months. These results demonstrate that prenatal hypoxia programs elevated blood pressure and exacerbates salt-induced cardiovascular and renal pathology in a sex specific manner. Given sex differences observed in RAS expression and nephron number, future studies may consider RAS blockade as a therapeutic target in this model.
Renal organogenesis What can it tell us about renal repair and regeneration?
Publisher: Informa UK Limited
Date: 10-2011
An interview with Melissa Little
Publisher: The Company of Biologists
Date: 15-03-2016
DOI: 10.1242/DEV.135897
Abstract: Melissa Little is a Senior Principal Research Fellow at the Murdoch Childrens Research Institute in Melbourne, Australia. Her lab has studied kidney development and regeneration for over 20 years, recently making notable advances in the generation of kidney organoids from human iPSCs. We chatted with Melissa about her career, her thoughts on the potential of the organoid and stem cell fields, and what she hopes to achieve during her guest editorship with Development.
Dads and disomy and disease
Publisher: Springer Science and Business Media LLC
Date: 06-1991
DOI: 10.1038/351609A0
Nephron progenitor commitment is a stochastic process influenced by cell migration.
Publisher: eLife Sciences Publications, Ltd
Date: 24-01-2019
DOI: 10.7554/ELIFE.41156
Abstract: Progenitor self-renewal and differentiation is often regulated by spatially restricted cues within a tissue microenvironment. Here, we examine how progenitor cell migration impacts regionally induced commitment within the nephrogenic niche in mice. We identify a subset of cells that express Wnt4, an early marker of nephron commitment, but migrate back into the progenitor population where they accumulate over time. Single cell RNA-seq and computational modelling of returning cells reveals that nephron progenitors can traverse the transcriptional hierarchy between self-renewal and commitment in either direction. This plasticity may enable robust regulation of nephrogenesis as niches remodel and grow during organogenesis.
Allelic imbalance at chromosome 1q21 in Wilms tumor
Publisher: Elsevier BV
Date: 08-1997
DOI: 10.1016/S0165-4608(96)00342-1
Abstract: The Wilms tumor suppressor gene 1, WT1, located on chromosome 11p13 is mutated in only a subset of Wilms tumors. Cytogenetic studies of Wilms tumors show that the most frequent structural anomalies after those affecting chromosome 11p are rearrangements of 1q, suggesting that there is a gene involved in Wilms tumor etiology in this region. The WT1 target sequence +P5 (D1S3309E), isolated using whole-genome polymerase chain reaction (PCR), binds all WT1 isoforms in vitro and has been mapped to 1q21-22. As +P5 may mark a 1q Wilms tumor gene, constitutional and tumor DNA from 33 Wilms tumor patients (36 tumors) was screened for allele imbalance using microsatellite markers from 1p21 to 1q44. Although no gross rearrangements of the +P5 region were found, this study demonstrates allele imbalance for 1q in 12% of patients (5/36 tumors), defining a smallest region of overlap at 1q21. This finding supports a role for 1q21 in Wilms tumorigenesis.
Recreating kidney progenitors from pluripotent cells
Publisher: Springer Science and Business Media LLC
Date: 13-09-2013
Definition and spatial annotation of the dynamic secretome during early kidney development
Publisher: Wiley
Date: 2006
DOI: 10.1002/DVDY.20740
Abstract: The term "secretome" has been defined as a set of secreted proteins (Grimmond et al. [2003] Genome Res 13:1350-1359). The term "secreted protein" encompasses all proteins exported from the cell including growth factors, extracellular proteinases, morphogens, and extracellular matrix molecules. Defining the genes encoding secreted proteins that change in expression during organogenesis, the dynamic secretome, is likely to point to key drivers of morphogenesis. Such secreted proteins are involved in the reciprocal interactions between the ureteric bud (UB) and the metanephric mesenchyme (MM) that occur during organogenesis of the metanephros. Some key metanephric secreted proteins have been identified, but many remain to be determined. In this study, microarray expression profiling of E10.5, E11.5, and E13.5 kidney and consensus bioinformatic analysis were used to define a dynamic secretome of early metanephric development. In situ hybridisation was used to confirm microarray results and clarify spatial expression patterns for these genes. Forty-one secreted factors were dynamically expressed between the E10.5 and E13.5 timeframe profiled, and 25 of these factors had not previously been implicated in kidney development. A text-based anatomical ontology was used to spatially annotate the expression pattern of these genes in cultured metanephric explants.
The kids are OK: It is discrimination, not same-sex parents, that harms children
Publisher: AMPCo
Date: 23-10-2017
DOI: 10.5694/MJA17.00943
The origin and role of the renal stroma
Publisher: The Company of Biologists
Date: 23-09-2021
DOI: 10.1242/DEV.199886
Abstract: The postnatal kidney is predominantly composed of nephron epithelia with the interstitial components representing a small proportion of the final organ, except in the diseased state. This is in stark contrast to the developing organ, which arises from the mesoderm and comprises an expansive stromal population with distinct regional gene expression. In many organs, the identity and ultimate function of an epithelium is tightly regulated by the surrounding stroma during development. However, although the presence of a renal stromal stem cell population has been demonstrated, the focus has been on understanding the process of nephrogenesis whereas the role of distinct stromal components during kidney morphogenesis is less clear. In this Review, we consider what is known about the role of the stroma of the developing kidney in nephrogenesis, where these cells come from as well as their heterogeneity, and reflect on how this information may improve human kidney organoid models.
Plasticity of distal nephron epithelia from human kidney organoids enables the induction of ureteric tip and stalk
Publisher: Elsevier BV
Date: 04-2020
Direct transcriptional reprogramming to nephron progenitors
Publisher: Elsevier BV
Date: 10-2015
DOI: 10.1016/J.GDE.2015.06.001
Abstract: The direct reprogramming of one cell fate to another represents an attractive option for the generation of specific endpoints for cellular therapy. This appears to require both the reactivation of critical transcription factor regulatory networks and chromatin remodelling. The direct reprogramming of mature renal epithelial cell lines to a nephron progenitor state has been reported. However, our limited knowledge of the optimal growth conditions to maintain this state remains a challenge for their therapeutic application. Here we examine whether nephron progenitors as an endpoint of direct reprogramming have been suitably defined and whether alternative options for reprogramming to kidney exist.
Stem cell options for kidney disease
Publisher: Wiley
Date: 04-12-2009
DOI: 10.1002/PATH.2477
Abstract: Chronic kidney disease (CKD) is increasing at the rate of 6-8% per annum in the US alone. At present, dialysis and transplantation remain the only treatment options. However, there is hope that stem cells and regenerative medicine may provide additional regenerative options for kidney disease. Such new treatments might involve induction of repair using endogenous or exogenous stem cells or the reprogramming of the organ to reinitiate development. This review addresses the current state of understanding with respect to the ability of non-renal stem cell sources to influence renal repair, the existence of endogenous renal stem cells and the biology of normal renal repair in response to damage. It also examines the remaining challenges and asks the question of whether there is one solution for all forms of renal disease.
(Re)Building a Kidney
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 17-01-2017
Abstract: (Re)Building a Kidney is a National Institute of Diabetes and Digestive and Kidney Diseases-led consortium to optimize approaches for the isolation, expansion, and differentiation of appropriate kidney cell types and the integration of these cells into complex structures that replicate human kidney function. The ultimate goals of the consortium are two-fold: to develop and implement strategies for in vitro engineering of replacement kidney tissue, and to devise strategies to stimulate regeneration of nephrons in situ to restore failing kidney function. Projects within the consortium will answer fundamental questions regarding human gene expression in the developing kidney, essential signaling crosstalk between distinct cell types of the developing kidney, how to derive the many cell types of the kidney through directed differentiation of human pluripotent stem cells, which bioengineering or scaffolding strategies have the most potential for kidney tissue formation, and basic parameters of the regenerative response to injury. As these projects progress, the consortium will incorporate systematic investigations in physiologic function of in vitro and in vivo differentiated kidney tissue, strategies for engraftment in experimental animals, and development of therapeutic approaches to activate innate reparative responses.
An In Vitro Differentiation Protocol for Human Embryonic Bipotential Gonad and Testis Cell Development
Publisher: Elsevier BV
Date: 12-2020
Directing stem cells to kidney: From development to regeneration
Publisher: Elsevier BV
Date: 07-2017
Patterning a Ureter Is All in the Stroma
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 21-08-2020
Advances in predictive in vitro models of drug-induced nephrotoxicity
Publisher: Springer Science and Business Media LLC
Date: 06-04-2018
GENERATION OF HOMOZYGOSITY AT THE C-HA-RAS-1 LOCUS ON CHROMOSOME-11P IN AN ADRENAL ADENOMA FROM AN ADULT WITH WIEDEMANN-BECKWITH SYNDROME
Publisher: Elsevier BV
Date: 1988
DOI: 10.1016/0165-4608(88)90100-8
Abstract: Generation of homozygosity for the human c-Ha-ras-1 locus on the short arm of chromosome #11 (11p) has been demonstrated for an adrenal adenoma from an adult with Wiedemann-Beckwith syndrome (WBS). This is the first demonstration of loss of somatic heterozygosity for a locus on 11p in an adrenal neoplasm and is the first instance where a tumor of any type, from a patient with WBS, shows loss of heterozygosity in this region of the genome. Generation of homozygosity in an adenoma, rather than a carcinoma, demonstrates that this mechanism is an early event in tumorigenesis rather than a late event associated with tumor progression.
Redirection of renal mesenchyme to stromal and chondrocytic fates in the presence of TGF-Β2
Publisher: Elsevier BV
Date: 04-2010
DOI: 10.1016/J.DIFF.2010.01.004
Abstract: Many members of the transforming growth factor-beta (TGF-beta) superfamily have been shown to be important regulators of metanephric development. In this study, we characterized the effect of TGF-beta2 on metanephric development. Rat and mouse metanephroi cultured in the presence of exogenous TGF-beta2 for up to 15 days were small, and contained rudimentary ureteric branches and few glomeruli. These metanephroi were mostly comprised of mesenchymal cells, with two cell populations (designated Type 1 and Type 2 cells) evident. Type 1 cells were only observed when TGF-beta2 was added from the commencement of culture, they resembled chondroblasts and were Alcian Blue and Col IIB positive. Type 2 cells were observed whenever TGF-beta2 was added to the media, formed a band at the periphery of the explants consisting of 5-10 layers of spindle-shaped cells, and were alpha-smooth muscle actin positive. Molecular and RNA in situ hybridization analysis of metanephroi cultured in the presence of TGF-beta2 for 6 days demonstrated that Type 1 and 2 cells were negative for Pax2, WT1, GDNF and FoxD1. Gene expression profiling demonstrated an upregulation of chondrocyte, myogenic and stromal genes, some of which were identified as markers of Type 1 and Type 2 cells. In addition, TGF-beta2 was capable of maintaining the survival of mouse isolated metanephric mesenchyme (iMM) in the absence of serum or inductive signals from the ureteric epithelium. TGF-beta2 also induced the differentiation of iMM into Type 1 and 2 cells. The presence of chondrocytes and muscle in these cultures is reminiscent of the cell types found in some Wilms' tumors. These studies demonstrate that TGF-beta2 is capable of differentiating metanephric mesenchyme away from a renal cell fate.
Temporal and spatial transcriptional programs in murine kidney development
Publisher: American Physiological Society
Date: 17-10-2005
DOI: 10.1152/PHYSIOLGENOMICS.00043.2005
Abstract: We have performed a systematic temporal and spatial expression profiling of the developing mouse kidney using Compugen long-oligonucleotide microarrays. The activity of 18,000 genes was monitored at 24-h intervals from 10.5-day-postcoitum (dpc) metanephric mesenchyme (MM) through to neonatal kidney, and a cohort of 3,600 dynamically expressed genes was identified. Early metanephric development was further surveyed by directly comparing RNA from 10.5 vs. 11.5 vs. 13.5dpc kidneys. These data showed high concordance with the previously published dynamic profile of rat kidney development (Stuart RO, Bush KT, and Nigam SK. Proc Natl Acad Sci USA 98: 5649–5654, 2001) and our own temporal data. Cluster analyses were used to identify gene ontological terms, functional annotations, and pathways associated with temporal expression profiles. Genetic network analysis was also used to identify biological networks that have maximal transcriptional activity during early metanephric development, highlighting the involvement of proliferation and differentiation. Differential gene expression was validated using whole mount and section in situ hybridization of staged embryonic kidneys. Two spatial profiling experiments were also undertaken. MM (10.5dpc) was compared with adjacent intermediate mesenchyme to further define metanephric commitment. To define the genes involved in branching and in the induction of nephrogenesis, expression profiling was performed on ureteric bud (GFP+) FACS sorted from HoxB7-GFP transgenic mice at 15.5dpc vs. the GFP− mesenchymal derivatives. Comparisons between temporal and spatial data enhanced the ability to predict function for genes and networks. This study provides the most comprehensive temporal and spatial survey of kidney development to date, and the compilation of these transcriptional surveys provides important insights into metanephric development that can now be functionally tested.
Parietal Epithelial Cells Regenerate Podocytes
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 02-2009
Polarity, cell division, and out-of-equilibrium dynamics control the growth of epithelial structures
Publisher: Rockefeller University Press
Date: 21-10-2013
Abstract: The growth of a well-formed epithelial structure is governed by mechanical constraints, cellular apico-basal polarity, and spatially controlled cell ision. Here we compared the predictions of a mathematical model of epithelial growth with the morphological analysis of 3D epithelial structures. In both in vitro cyst models and in developing epithelial structures in vivo, epithelial growth could take place close to or far from mechanical equilibrium, and was determined by the hierarchy of time-scales of cell ision, cell–cell rearrangements, and lumen dynamics. Equilibrium properties could be inferred by the analysis of cell–cell contact topologies, and the nonequilibrium phenotype was altered by inhibiting ROCK activity. The occurrence of an aberrant multilumen phenotype was linked to fast nonequilibrium growth, even when geometric control of cell ision was correctly enforced. We predicted and verified experimentally that slowing down cell ision partially rescued a multilumen phenotype induced by altered polarity. These results improve our understanding of the development of epithelial organs and, ultimately, of carcinogenesis.
Shining a Light on Alport Syndrome
Publisher: Elsevier BV
Date: 05-2018
DOI: 10.1016/J.CHEMBIOL.2018.05.001
Abstract: In this issue of Cell Chemical Biology, Omachi et al. (2018) present a split Nanoluciferase system to identify successful protein trimerization in Alport syndrome. This elegant proof of concept suggests opportunities for drug screening for Alport syndrome and may be transferable to the study of other diseases affecting protein-protein interactions.
Bayesian inference of agent-based models: a tool for studying kidney branching morphogenesis
Publisher: Springer Science and Business Media LLC
Date: 02-2018
High throughput single cell RNA-seq of developing mouse kidney and human kidney organoids reveals a roadmap for recreating the kidney
Publisher: Cold Spring Harbor Laboratory
Date: 16-12-2017
DOI: 10.1101/235499
Abstract: Recent 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 in idual 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 in idual 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.
An integrated cell, tissue and whole organ profile of kidney morphogenesis
Publisher: Elsevier BV
Date: 07-2017
Renal Subcapsular Transplantation of PSC-Derived Kidney Organoids Induces Neo-vasculogenesis and Significant Glomerular and Tubular Maturation In Vivo
Publisher: Elsevier BV
Date: 03-2018
Crim1 is required for maintenance of the ocular lens epithelium
Publisher: Elsevier BV
Date: 05-2018
DOI: 10.1016/J.EXER.2018.02.012
Abstract: The development and growth of the vertebrate ocular lens is dependent on the regulated proliferation of an anterior monolayer of epithelial cells, and their subsequent differentiation into elongate fiber cells. The growth factor rich ocular media that bathes the lens mediates these cellular processes, and their respective intracellular signaling pathways are in turn regulated to ensure that the proper lens architecture is maintained. Recent studies have proposed that Cysteine Rich Motor Neuron 1 (Crim1), a transmembrane protein involved in organogenesis of many tissues, might influence cell adhesion, polarity and proliferation in the lens by regulating integrin-signaling. Here, we characterise the lens and eyes of the Crim1
Evidence that WT1 mutations in Denys — Drash syndrome patients may act in a dominant-negative fashion
Publisher: Oxford University Press (OUP)
Date: 1993
DOI: 10.1093/HMG/2.3.259
Abstract: The triad of nephropathy, partial gonadal dysgenesis and Wilms' tumour (WT) is known as Denys-Drash syndrome (DDS). The WT predisposition gene WT1, which plays a vital role in both genital and renal development, is known to be mutated in DDS patients. The WT1 mutations in these patients are constitutional point mutations clustered in the zinc finger (ZF) encoding exons, particularly the exons encoding ZF2 and ZF3. The predicted functional alteration in WT1 is thought to underlie DDS aetiology either by abolishing binding of the WT1 ZF domain to its normal target DNA binding site(s), perhaps blocking the binding of the wild type WT1 present (dominant negative mutation), and/or by conferring the ability to recognise novel but inappropriate DNA binding sites (dominant mutation). We report here on the analysis of WT1 in a further five cases of DDS. In each case a constitutional point mutation was detected in either ZF2 or ZF3. Three of these mutations are novel, with two affecting the conserved histidine and cysteine residues crucial for ZF tertiary structure. The protein product of the third is predicted to lack ZF2, 3 and 4 as a result of a chain termination mutation, and is presumably incapable of binding DNA. However since the DDS phenotype is only elicited by mutations which lead to loss or alteration of ZF function (presumably DNA binding) while the N-terminal upstream portion of the gene remains intact, we suggest that a dominant negative mechanism is at work here.
Defining and redefining the nephron progenitor population
Publisher: Springer Science and Business Media LLC
Date: 09-2011
Characterisation of Crim1 expression in the developing mouse urogenital tract reveals a sexually dimorphic gonadal expression pattern
Publisher: Wiley
Date: 2000
DOI: 10.1002/1097-0177(2000)9999:9999<::AID-DVDY1072>3.0.CO;2-I
Patient-iPSC-Derived Kidney Organoids Show Functional Validation of a Ciliopathic Renal Phenotype and Reveal Underlying Pathogenetic Mechanisms.
Publisher: Elsevier BV
Date: 05-2018
Colony-stimulating factor-1 promotes kidney growth and repair via alteration of macrophage responses
Publisher: Elsevier BV
Date: 09-2011
Regrow or Repair: An Update on Potential Regenerative Therapies for the Kidney
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 2022
Abstract: Fifteen years ago, this journal published a review outlining future options for regenerating the kidney. At that time, stem cell populations were being identified in multiple tissues, the concept of stem cell recruitment to a site of injury was of great interest, and the possibility of postnatal renal stem cells was growing in momentum. Since that time, we have seen the advent of human induced pluripotent stem cells, substantial advances in our capacity to both sequence and edit the genome, global and spatial transcriptional analysis down to the single-cell level, and a pandemic that has challenged our delivery of health care to all. This article will look back over this period of time to see how our view of kidney development, disease, repair, and regeneration has changed and envision a future for kidney regeneration and repair over the next 15 years.
Understanding kidney morphogenesis to guide renal tissue regeneration
Publisher: Springer Science and Business Media LLC
Date: 30-08-2016
Abstract: The treatment of renal failure has seen little change in the past 70 years. Patients with end-stage renal disease (ESRD) are treated with renal replacement therapy, including dialysis or organ transplantation. The growing imbalance between the availability of donor organs and prevalence of ESRD is pushing an increasing number of patients to undergo dialysis. Although the prospect of new treatment options for patients through regenerative medicine has long been suggested, advances in the generation of human kidney cell types through the directed differentiation of human pluripotent stem cells over the past 2 years have brought this prospect closer to delivery. These advances are the result of careful research into mammalian embryogenesis. By understanding the decision points made within the embryo to pattern the kidney, it is now possible to recreate self-organizing kidney tissues in vitro. In this Review, we describe the key decision points in kidney development and how these decisions have been mimicked experimentally. Recreation of human nephrons from human pluripotent stem cells opens the door to patient-derived disease models and personalized drug and toxicity screening. In the long term, we hope that these efforts will also result in the generation of bioengineered organs for the treatment of kidney disease.
Report on the first Asia-Pacific Kidney Development Workshop.
Publisher: Wiley
Date: 25-02-2013
DOI: 10.1111/NEP.12026
Self-organisation after embryonic kidney dissociation is driven via selective adhesion of ureteric epithelial cells.
Publisher: The Company of Biologists
Date: 2017
DOI: 10.1242/DEV.140228
Abstract: Human pluripotent stem cells, after directed differentiation in vitro, can spontaneously generate complex tissues via self-organisation of the component cells. Self-organisation can also reform embryonic organ structure after tissue disruption. It has previously been demonstrated that dissociated embryonic kidneys can recreate component epithelial and mesenchymal relationships sufficient to allow continued kidney morphogenesis. Here we investigate the timing and underlying mechanisms driving self-organisation after dissociation of the embryonic kidney using time-lapse imaging, high-resolution confocal analyses and mathematical modelling. Organotypic self-organisation sufficient for nephron initiation was observed within a 24 hour period. This involved cell movement with structure emerging after the clustering of ureteric epithelial cells, a process consistent with models of random cell movement with preferential cell adhesion. Ureteric epithelialisation rapidly followed the formation of ureteric cell clusters with the reformation of nephron forming niches representing a later event. Disruption of P-cadherin interactions was seen to impair this ureteric epithelial cell clustering without affecting epithelial maturation. This understanding may facilitate improved regulation of patterning within organoids and facilitate kidney engineering approaches guided by cell-cell self-organisation.
Multivariate patterning of human pluripotent cells under perfusion reveals critical roles of induced paracrine factors in kidney organoid development
Publisher: American Association for the Advancement of Science (AAAS)
Date: 10-01-2020
Abstract: Multivariate patterning of perfused pluripotent cells reveals critical roles of induced paracrine factors in kidney organoids.
Differential gene expression in the developing mouse ureter
Publisher: Elsevier BV
Date: 06-2006
DOI: 10.1016/J.MODGEP.2005.10.008
Abstract: In many instances, kidney dysgenesis results as a secondary consequence to defects in the development of the ureter. Through the use of mouse genetics a number of genes associated with such malformations have been identified, however, the cause of many other abnormalities remain unknown. In order to identify novel genes involved in ureter development we compared gene expression in embryonic day (E) 12.5, E15.5 and postnatal day (P) 75 ureters using the Compugen mouse long oligo microarrays. A total of 248 genes were dynamically upregulated and 208 downregulated between E12.5 and P75. At E12.5, when the mouse ureter is comprised of a simple cuboidal epithelium surrounded by ureteric mesenchyme, genes previously reported to be expressed in the ureteric mesenchyme, foxC1 and foxC2 were upregulated. By E15.5 the epithelial layer develops into urothelium, impermeable to urine, and smooth muscle develops for the peristaltic movement of urine towards the bladder. The development of these two cell types coincided with the upregulation of UPIIIa, RAB27b and PPARgamma reported to be expressed in the urothelium, and several muscle genes, Acta1, Tnnt2, Myocd, and Tpm2. In situ hybridization identified several novel genes with spatial expression within the smooth muscle, Acta1 ureteric mesenchyme and smooth muscle, Thbs2 and Col5a2 and urothelium, Kcnj8 and Adh1. This study marks the first known report defining global gene expression of the developing mouse ureter and will provide insight into the molecular mechanisms underlying kidney and lower urinary tract malformations.
Subfractionation of differentiating human embryonic stem cell populations allows the isolation of a mesodermal population enriched for intermediate mesoderm and putative renal progenitors
Publisher: Mary Ann Liebert Inc
Date: 10-2010
Use of in situ hybridization to examine gene expression in the embryonic, neonatal, and adult urogenital system.
Publisher: Humana Press
Date: 2012
DOI: 10.1007/978-1-61779-851-1_20
Abstract: Studies into the molecular basis of morphogenesis frequently begin with investigations into gene expression across time and cell type in that organ. One of the most anatomically informative approaches to such studies is the use of in situ hybridization, either of intact or histologically sectioned tissues. Here, we describe the optimization of this approach for use in the temporal and spatial analysis of gene expression in the urogenital system, from embryonic development to the postnatal period. The methods described are applicable for high throughput analysis of large gene sets. As such, ISH has become a powerful technique for gene expression profiling and is valuable for the validation of profiling analyses performed using other approaches such as microarrays.
DNA binding capacity of the WT1 protein is abolished by Denys—Drash syndrome WT1 point mutations
Publisher: Oxford University Press (OUP)
Date: 1995
DOI: 10.1093/HMG/4.3.351
Abstract: Constitutional point mutations in the zinc finger (ZF) region of the Wilms' tumour suppressor gene 1 (WT1) lead to Denys-Drash syndrome (DDS). Patients with this syndrome display renal failure, Wilms' tumour (WT) and pseudohermaphroditism. DDS WT1 mutations fall into three major categories: (a) missense mutations altering amino acids which directly interact with the DNA target (b) substitution of amino acids involved in zinc complexing and (c) nonsense mutations leading to the removal of at least two zinc fingers. We have expressed the WT1 zinc fingers as glutathione-S-transferase fusion proteins, with the lysine-threonine-serine (KTS) alternate splice between ZF3 and ZF4 either present or absent. WT1 fusion constructs with all three classes of DDS mutation were also created. Wild-type and mutant fusion proteins were assayed for their DNA-binding affinity using four previously identified WT1 DNA targets: an EGR1 consensus site murine insulin-like growth factor 2 promoter 2 (IGF2P2) a (TCC)n motif from the PDGFA-chain promoter and +P5, a genomic fragment isolated by its affinity for WT1 + KTS. WT1-KTS bound all four targets, but WT1 + KTS only bound +P5. All three classes of DDS mutation investigated, with or without KTS, abolished binding to all four targets. This provides evidence that DDS mutations act either as dominant-negative antimorphs, or elicit their effect through disturbed isoform dosage balance.
Lin28 and let-7 regulate the timing of cessation of murine nephrogenesis
Publisher: Springer Science and Business Media LLC
Date: 11-01-2019
DOI: 10.1038/S41467-018-08127-4
Abstract: In humans and in mice the formation of nephrons during embryonic development reaches completion near the end of gestation, after which no new nephrons are formed. The final nephron complement can vary 10-fold, with reduced nephron number predisposing in iduals to hypertension, renal, and cardiovascular diseases in later life. While the heterochronic genes lin28 and let-7 are well-established regulators of developmental timing in invertebrates, their role in mammalian organogenesis is not fully understood. Here we report that the Lin28b/ let-7 axis controls the duration of kidney development in mice. Suppression of let-7 miRNAs, directly or via the transient overexpression of LIN28B , can prolong nephrogenesis and enhance kidney function potentially via upregulation of the Igf2 / H19 locus. In contrast, kidney-specific loss of Lin28b impairs renal development. Our study reveals mechanisms regulating persistence of nephrogenic mesenchyme and provides a rationale for therapies aimed at increasing nephron mass.
A Cas9 Variant for Efficient Generation of Indel-Free Knockin or Gene-Corrected Human Pluripotent Stem Cells
Publisher: Elsevier BV
Date: 09-2016
Reprogramming the kidney: a novel approach for regeneration
Publisher: Elsevier BV
Date: 07-2012
DOI: 10.1038/KI.2012.68
Abstract: Nuclear reprogramming has reshaped stem cell science and created new avenues for cell-based therapies. The ability to bestow any given phenotype upon adult cells regardless of their origin is an exciting possibility. How can this powerful tool be harnessed for the treatment of kidney disease? Many approaches, including induced pluripotent stem cell (iPSC) production, direct lineage conversion, and reprogramming to a kidney progenitor, are now possible. Indeed, the generation of iPSC lines from adult kidney-derived cells has been successfully achieved. This, however, is just the beginning of the challenge. This review will discuss the fundamental concepts of transcription factor-based reprogramming in its various forms, highlighting recent advances in the field and how these are applicable to the kidney. The relative merits of each approach will be discussed in the context of what is a realistic and feasible strategy for kidney regeneration via reprogramming.
Transcriptional evaluation of the developmental accuracy, reproducibility and robustness of kidney organoids derived from human pluripotent stem cells
Publisher: Cold Spring Harbor Laboratory
Date: 22-12-2017
DOI: 10.1101/238428
Abstract: We have previously reported a protocol for the directed differentiation of human induced pluripotent stem cells to kidney organoids comprised of nephrons, proximal and distal epithelium, vasculature and surrounding interstitial elements. The utility of this protocol for applications such as disease modelling will rely implicitly on the developmental accuracy of the model, technical robustness of the protocol and transferability between iPSC lines. Here we report extensive transcriptional analyses of the sources of variation across the timecourse of differentiation from pluripotency to complete kidney organoid, focussing on repeated differentiations to day 18 organoid. In idual organoids generated within the same differentiation experiment show Spearman’s correlation coefficients of .99. The greatest source of variation was seen between experimental batch, with the enrichment for genes that also varied temporally between day 10 and day 25 organoids implicating nephron maturation as contributing to transcriptional variance between in idual differentiation experiments. A morphological analysis revealed a transition from renal vesicle to capillary loop stage nephrons across the same time period. Distinct iPSC clones were also shown to display congruent transcriptional programs with inter-experimental and inter-clonal variation most strongly associated with nephron patterning. Even epithelial cells isolated from organoids showed transcriptional alignment with total organoids of the same day of differentiation. This data provides a framework for managing experimental variation, thereby increasing the utility of this approach for personalised medicine and functional genomics.
Recessive NOS1AP variants impair actin remodeling and cause glomerulopathy in humans and mice
Publisher: American Association for the Advancement of Science (AAAS)
Date: 2021
Abstract: NOS1AP variants impair actin remodeling in podocytes and cause glomerular disease in humans, kidney organoids, and mice.
Dissociation of Embryonic Kidney Followed by Re-aggregation as a Method for Chimeric Analysis
Publisher: Humana Press
Date: 2012
DOI: 10.1007/978-1-61779-851-1_12
Abstract: This chapter presents three methods for re-constructing mouse foetal kidney tissue from simple suspensions of cells. These techniques are very useful for a number of purposes: (1) they allow the production of fine-grained chimaeras in which cell autonomy of mutations can be tested, (2) they provide an environment that allows the renal differentiation potential of stem cells to be assessed, and (3) they are an excellent system in which to study the mechanisms of self-organization. Each of the methods described here begins with disaggregation of embryonic mouse kidneys, followed by re-aggregation and culture the main differences are in the culture methods, each of which has advantages for particular purposes.
LOSS OF ALLELES ON THE SHORT ARM OF CHROMOSOME-11 IN A HEPATOBLASTOMA FROM A CHILD WITH BECKWITH-WIEDEMANN SYNDROME
Publisher: Springer Science and Business Media LLC
Date: 06-1988
DOI: 10.1007/BF00280564
Abstract: Prospective memory is a complex cognitive construct ubiquitous in everyday life that is thought to sometimes rely on executive skills commonly affected by Parkinson's disease (PD). The present study investigated the effect of PD on prospective memory tasks with varying demand on executive control processes, namely on the amount of strategic attentional monitoring required for intention retrieval. In iduals with PD but without dementia and healthy adults performed laboratory event-based prospective memory tasks that varied in whether strategic attentional monitoring (nonfocal condition) or spontaneous processes (focal condition) were primarily involved in intention retrieval. Participants also completed a questionnaire rating their frequency of prospective memory failures in everyday life for both self-cued and environment-cued tasks. PD participants performed worse than non-PD participants in the nonfocal, but not focal, condition of the laboratory task. They also reported more everyday failures than non-PD participants for self-cued, but not environment-cued, prospective memory tasks. Thus, nondemented in iduals with PD are preferentially impaired on prospective memory tasks for which higher levels of executive control are needed to support intention retrieval. This pattern is consistent across laboratory and reported real-world performance.
The Renal Papilla: An Enigma in Damage and Repair
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 12-2011
Characterisation and trophic functions of murine embryonic macrophages based upon the use of a Csf1r-EGFP transgene reporter
Publisher: Elsevier BV
Date: 08-2007
DOI: 10.1016/J.YDBIO.2007.05.027
Abstract: All solid organs contain resident monocyte-derived cells that appear early in organogenesis and persist throughout life. These cells are critical for normal development in some organs. Here we report the use of a previously described transgenic line, with EGFP driven by the macrophage-restricted Csf1r (c-fms) promoter, to image macrophage production and infiltration accompanying organogenesis in many tissues. Using microarray analysis of FACS-isolated EGFP-positive cells, we show that fetal kidney, lung and brain macrophages show similar gene expression profiles irrespective of their tissue of origin. EGFP-positive cells appeared in the renal interstitium from 12 days post coitum, prior to nephrogenesis, and maintain a close apposition to renal tubules postnatally. CSF-1 added to embryonic kidney explants increased overall renal growth and ureteric bud branching. Expression profiling of tissue macrophages and of CSF-1-treated explants showed evidence of the alternate, pro-proliferative (M2) activation profile, including expression of macrophage mannose receptor (CD206), macrophage scavenger receptor 2 (Msr2), C1q, CD163, selenoprotein P, CCL24 and TREM2. This response has been associated with the trophic role of tumour-associated macrophages. These findings suggest a trophic role of macrophages in embryonic kidney development, which may continue to play a similar role in postnatal repair.
Evaluation of variability in human kidney organoids.
Publisher: Springer Science and Business Media LLC
Date: 20-12-2018
A zinc finger truncation of murine WT1 results in the characteristic urogenital abnormalities of Denys–Drash syndrome
Publisher: Proceedings of the National Academy of Sciences
Date: 16-03-1999
Abstract: The Wilms tumor-suppressor gene, WT1 , plays a key role in urogenital development, and WT1 dysfunction is implicated in both neoplastic (Wilms tumor, mesothelioma, leukemias, and breast cancer) and nonneoplastic (glomerulosclerosis) disease. The analysis of diseases linked specifically with WT1 mutations, such as Denys–Drash syndrome (DDS), can provide valuable insight concerning the role of WT1 in development and disease. DDS is a rare childhood disease characterized by a nephropathy involving mesangial sclerosis, XY pseudohermaphroditism, and/or Wilms tumor (WT). DDS patients are constitutionally heterozygous for exonic point mutations in WT1 , which include mutations predicted to truncate the protein within the C-terminal zinc finger (ZF) region. We report that heterozygosity for a targeted murine Wt1 allele, Wt1 tmT396 , which truncates ZF3 at codon 396, induces mesangial sclerosis characteristic of DDS in adult heterozygous and chimeric mice. Male genital defects also were evident and there was a single case of Wilms tumor in which the transcript of the nontargeted allele showed an exon 9 skipping event, implying a causal link between Wt1 dysfunction and Wilms tumorigenesis in mice. However, the mutant WT1 tmT396 protein accounted for only 5% of WT1 in both heterozygous embryonic stem cells and the WT. This has implications regarding the mechanism by which the mutant allele exerts its effect.
Assignment<footref rid=foot01><sup>1</sup></footref> of the human slit homologue SLIT2 to human chromosome band 4p15.2
Publisher: S. Karger AG
Date: 1999
DOI: 10.1159/000015351
Anlaysis of complementary expression profiles following WT1 induction versus repression reveals the cholesterol/fatty acid synthetic pathways as a possible major target of WT1
Publisher: Springer Science and Business Media LLC
Date: 15-03-2004
Tracing the life of the kidney tubule - Re-establishing dogma and redirecting the options
Publisher: Elsevier BV
Date: 03-2008
DOI: 10.1016/J.STEM.2008.02.007
Abstract: Renal pathology suggests that tubular repair results from tubular proliferation. In contrast, recent studies propose that postnatal kidney repair may involve renal stem cells. In this issue of Cell Stem Cell, Humphreys et al. (2008) use lineage tracing to genetically assess contribution of adult nontubular cells (potentially stem cells) to repair of damaged renal tubules.
Nephron formation adopts a novel spatial topology at cessation of nephrogenesis
Publisher: Elsevier BV
Date: 12-2011
Double-hit compression behavior of TWIP steels
Publisher: Elsevier BV
Date: 12-2011
Wnt11 directs nephron progenitor polarity and motile behavior ultimately determining nephron endowment
Publisher: eLife Sciences Publications, Ltd
Date: 05-12-2018
DOI: 10.7554/ELIFE.40392
Abstract: A normal endowment of nephrons in the mammalian kidney requires a balance of nephron progenitor self-renewal and differentiation throughout development. Here, we provide evidence for a novel action of ureteric branch tip-derived Wnt11 in progenitor cell organization and interactions within the nephrogenic niche, ultimately determining nephron endowment. In Wnt11 mutants, nephron progenitors dispersed from their restricted niche, intermixing with interstitial progenitors. Nephron progenitor differentiation was accelerated, kidneys were significantly smaller, and the nephron progenitor pool was prematurely exhausted, halving the final nephron count. Interestingly, RNA-seq revealed no significant differences in gene expression. Live imaging of nephron progenitors showed that in the absence of Wnt11 they lose stable attachments to the ureteric branch tips, continuously detaching and reattaching. Further, the polarized distribution of several markers within nephron progenitors is disrupted. Together these data highlight the importance of Wnt11 signaling in directing nephron progenitor behavior which determines a normal nephrogenic program.
Luminal Mitosis Drives Epithelial Cell Dispersal within the Branching Ureteric Bud
Publisher: Elsevier BV
Date: 11-2013
Recurrent Nephrotic Plasma Activates Pro-fibrotic Signalling Pathways Downstream of Protease-activated Receptor 1
Publisher: Research Square Platform LLC
Date: 12-01-2022
DOI: 10.21203/RS.3.RS-1226619/V1
Abstract: Recurrence of steroid-resistant nephrotic syndrome (SRNS) is thought to be due to an unknown “circulating factor”, the identity of which has so far remained elusive. Our previous work suggests a signaling role for protease-activated receptor-1 (PAR-1), leading to impaired podocyte function. Here, we show that relapse nephrotic plasma (NP), but not paired remission plasma, induced a pro-fibrotic response. This change was inhibited by PAR-1 inhibitors, but not by TGF-β1 inhibition. Four PAR-1 inhibitors demonstrated distinct antagonistic properties. The phosphorylation of VASP and JNK in a 3D spheroid model (GlomSpheres) and kidney organoids corroborated the finding from a 2D ciPods model. Functionally, relapse NP induced podocyte motility, and podocyte loss from spheroids both of which were also selectively rescued by PAR-1 inhibitors. Also, it induced the loss of podocyte-specific markers in kidney organoids. We propose that the circulating factor acts as a pro-fibrotic effector by activating PAR-1, leading to increased podocyte injury.
Recapitulating kidney development: Progress and challenges
Publisher: Elsevier BV
Date: 07-2019
Identification of novel markers of mouse fetal ovary development
Publisher: Public Library of Science (PLoS)
Date: 26-07-2012
Epigenetics and developmental programming of adult onset diseases
Publisher: Springer Science and Business Media LLC
Date: 02-02-2012
DOI: 10.1007/S00467-012-2108-X
Abstract: Maternal perturbations or sub-optimal conditions during development are now recognized as contributing to the onset of many diseases manifesting in adulthood. This "developmental programming" of disease has been explored using animal models allowing insights into the potential mechanisms involved. Impaired renal development, resulting in a low nephron number, has been identified as a common outcome that is likely to contribute to the development of hypertension in the offspring as adults. Changes in other organs and systems, including the heart and the hypothalamic–pituitary–adrenal axis, have also been found. Evidence has recently emerged suggesting that epigenetic changes may occur as a result of developmental programming and result in permanent changes in the expression patterns of particular genes. Such epigenetic modifications may be responsible not only for an increased susceptibility to disease for an in idual, but indirectly for the establishment of a disease state in a subsequent generation. Further research in this field, particularly examination as to whether epigenetic changes to genes affecting kidney development do occur, are essential to understanding the underlying mechanisms of developmental programming of disease.
Modelling cell turnover in a complex tissue during development
Publisher: Elsevier BV
Date: 12-2013
DOI: 10.1016/J.JTBI.2013.08.033
Abstract: The growth of organs results from proliferation within distinct cellular compartments. Organ development also involves transitions between cell types and variations in cell cycle duration as development progresses, and is regulated by a balance between entry into the compartment, proliferation of cells within the compartment, acquisition of quiescence and exit from that cell state via differentiation or death. While it is important to understand how environmental or genetic alterations can perturb such development, most approaches employed to date are descriptive rather than quantitative. This is because the identification and quantification of such parameters, while tractable in vitro, is challenging in the context of a complex tissue in vivo. Here we present a new framework for determining cell turnover in developing organs in vivo that combines cumulative cell-labelling and quantification of distinct cell-cycle phases without assuming homogeneity of behaviour within that compartment. A mathematical model is given that allows the calculation of cell cycle length in the context of a specific biological ex le and assesses the uncertainty of this calculation due to incomplete knowledge of cell cycle dynamics. This includes the development of a two population model to quantify possible heterogeneity of cell cycle length within a compartment and estimate the aggregate proliferation rate. These models are demonstrated on data collected from a progenitor cell compartment within the developing mouse kidney, the cap mesenchyme. This tissue was labelled by cumulative infusion, volumetrically quantified across time, and temporally analysed for the proportion of cells undergoing proliferation. By combining the cell cycle length predicted by the model with measurements of total cell population and mitotic rate, this approach facilitates the quantification of exit from this compartment without the need for a direct marker of that event. As a method specifically designed with assumptions appropriate to developing organs we believe this approach will be applicable to a range of developmental systems, facilitating estimations of cell cycle length and compartment behaviour that extend beyond simple comparisons of mitotic rates between normal and perturbed states.
Prolonged prenatal hypoxia selectively disrupts collecting duct patterning and postnatal function in male mouse offspring
Publisher: Wiley
Date: 05-07-2018
DOI: 10.1113/JP275918
Proximal tubule overexpression of a locally acting IGF isoform, Igf-1Ea, increases inflammation after ischemic injury
Publisher: Elsevier BV
Date: 02-2012
DOI: 10.1016/J.GHIR.2011.11.002
Abstract: IGF-1 is an important regulator of postnatal growth in mammals. In mice, a non-circulating, locally acting isoform of IGF-1, IGF-1Ea, has been documented as a central regulator of muscle regeneration and has been shown to improve repair in the heart and skin. In this study, we examine whether local production of IGF1-Ea protein improves tubular repair after renal ischemia reperfusion injury. Transgenic mice in which the proximal-tubule specific promoter Sglt2 was driving the expression of an Igf-1Ea transgene. These animals were treated with an ischemic-reperfusion injury and the response at 24h and 5days compared with wildtype littermates. Transgenic mice demonstrated rapid and enhanced renal injury in comparison to wild type mice. Five days after injury the wild type and low expressing Igf-1Ea transgenic mice showed significant tubular recovery, while high expressing Igf-1Ea transgenic mice displayed significant tubular damage. This marked injury was accompanied by a two-fold increase in the number of F4/80 positive macrophages and a three-fold increase in the number of Gr1-positive neutrophils in the kidney. At the molecular level, Igf-1Ea expression resulted in significant up-regulation of proinflammatory cytokines such as TNF-α and Ccl2. Expression of Nfatc1 was also delayed, suggesting reduced tubular proliferation after kidney injury. These data indicate that, unlike the muscle, heart and skin, elevated levels of IGF-1Ea in the proximal tubules exacerbates ischemia reperfusion injury resulting in increased recruitment of macrophages and neutrophils and delays repair in a renal setting.
Zinc finger point mutations within the WT1 gene in Wilms tumor patients.
Publisher: Proceedings of the National Academy of Sciences
Date: 06-1992
Abstract: A proposed Wilms tumor gene, WT1, which encodes a zinc finger protein, has previously been isolated from human chromosome 11p13. Chemical mismatch cleavage analysis was used to identify point mutations in the zinc finger region of this gene in a series of 32 Wilms tumors. Two exonic single base changes were detected. In zinc finger 3 of a bilateral Wilms tumor patient, a constitutional de novo C----T base change was found changing an arginine to a stop codon. One tumor from this patient showed allele loss leading to 11p hemizygosity of the abnormal allele. In zinc finger 2 of a sporadic Wilms tumor patient, a C----T base change resulted in an arginine to cysteine amino acid change. To our knowledge, a WT1 gene missense mutation has not been detected previously in a Wilms tumor. By comparison with a recent NMR and x-ray crystallographic analysis of an analogous zinc finger gene, early growth response gene 1 (EGR1), this amino acid change in WT1 occurs at a residue predicted to be critical for DNA binding capacity and site specificity. The detection of one nonsense point mutation and one missense WT1 gene point mutation adds to the accumulating evidence implicating this gene in a proportion of Wilms tumor patients.
Returning to kidney development to deliver synthetic kidneys
Publisher: Elsevier BV
Date: 06-2020
Reprogramming Somatic Cells to a Kidney Fate
Publisher: Elsevier BV
Date: 07-2014
DOI: 10.1016/J.SEMNEPHROL.2014.06.012
Abstract: Recent years have challenged the view that adult somatic cells reach a state of terminal differentiation. Although the ultimate ex le of this, somatic cell nuclear transfer, has not proven feasible in human beings, dedifferentiation of mature cell types to a more primitive state, direct reprogramming from one mature state to another, and the reprogramming of any adult cell type to a pluripotent state via enforced expression of key transcription factors now all have been shown. The implications of these findings for kidney disease include the re-creation of key renal cell types from more readily available and expandable somatic cell sources. The feasibility of such an approach recently was shown with the dedifferentiation of proximal tubule cells to nephrogenic mesenchyme. In this review, we examine the technical and clinical challenges that remain to such an approach and how new reprogramming approaches also may be useful for kidney disease.
Bioprinted pluripotent stem cell-derived kidney organoids provide opportunities for high content screening
Publisher: Cold Spring Harbor Laboratory
Date: 23-12-2018
DOI: 10.1101/505396
Abstract: Recent advances in the directed differentiation of human pluripotent stem cells to kidney brings with it the prospect of drug screening and disease modelling using patient-derived stem cell lines. Development of such an approach for high content screening will require substantial quality control and improvements in throughput. Here we demonstrate the use of the NovoGen MMX 3D bioprinter for the generation of highly reproducible kidney organoids from as few as 4,000 cells. Histological and immunohistochemical analyses confirmed the presence of renal epithelium, glomeruli, stroma and endothelium, while single cell RNAseq revealed equivalence to the cell clusters present within previously described organoids. The process is highly reproducible, rapid and transferable between cell lines, including genetically engineered reporter lines. We also demonstrate the capacity to bioprint organoids in a 96-well format and screen for response to doxorubicin toxicity as a proof of concept for high content compound screening.
Kidney Organoids Generated Using an Allelic Series of NPHS2 Point Variants Reveal Distinct Intracellular Podocin Mistrafficking
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 27-09-2022
Abstract: Missense variants of NPHS2 that cause mistrafficking of the encoded protein, PODOCIN, have been associated with steroid-resistant nephrotic syndrome. However, most studies have overexpressed such variants in 2D nonpodocyte cells. This study describes the generation and characterization of human kidney organoids representing an allelic series of homozygous NPHS2 missense variants. The strategy revealed a previously unappreciated reduction in variant PODOCIN protein, variant-specific subcellular localization, and specific effects on NEPHRIN association. All variants showed apoptosis in the absence of endoplasmic reticulum stress. Engineering endogenous NPHS2 variants to model in 3D human organoids provides a more accurate view of the pathobiology and a toolkit to screen compounds for reduction of variant protein degradation and mistrafficking. NPHS2 variants are the most common cause of steroid-resistant nephrotic syndrome in children month old. Missense NPHS2 variants were reported to cause mistrafficking of the encoded protein, PODOCIN, but this conclusion was on the basis of overexpression in some nonpodocyte cell lines. We generated a series of human induced pluripotent stem cell (iPSC) lines bearing pathogenic missense variants of NPHS2 , encoding the protein changes p.G92C, p.P118L, p.R138Q, p.R168H, and p.R291W, and control lines. iPSC lines were also generated from a patient with steroid-resistant nephrotic syndrome (p.R168H homozygote) and a healthy heterozygous parent. All lines were differentiated into kidney organoids. Immunofluorescence assessed PODOCIN expression and subcellular localization. Podocytes were transcriptionally profiled and PODOCIN-NEPHRIN interaction interrogated. All variant lines revealed reduced levels of PODOCIN protein in the absence of reduced transcription. Although wild-type PODOCIN localized to the membrane, distinct variant proteins displayed unique patterns of subcellular protein trafficking, some unreported. P118L and R138Q were preferentially retained in the endoplasmic reticulum (ER) R168H and R291W accumulated in the Golgi. Podocyte profiling demonstrated minimal disease-associated transcriptional change. All variants displayed podocyte-specific apoptosis, which was not linked to ER stress. NEPHRIN-PODOCIN colocalization elucidated the variant-specific effect on NEPHRIN association and hence NEPHRIN trafficking. Specific variants of endogenous NPHS2 result in distinct subcellular PODOCIN localization within organoid podocytes. Understanding the effect of each variant on protein levels and localization and the effect on NEPHRIN provides additional insight into the pathobiology of NPHS2 variants. This article contains a podcast at edirect.mp3/edia odcast/JASN/2023_01_05_JASN2022060707.mp3
Directing human embryonic stem cell differentiation towards a renal lineage generates a self-organizing kidney
Publisher: Springer Science and Business Media LLC
Date: 15-12-2013
DOI: 10.1038/NCB2894
Abstract: With the prevalence of end-stage renal disease rising 8% per annum globally, there is an urgent need for renal regenerative strategies. The kidney is a mesodermal organ that differentiates from the intermediate mesoderm (IM) through the formation of a ureteric bud (UB) and the interaction between this bud and the adjacent IM-derived metanephric mesenchyme (MM). The nephrons arise from a nephron progenitor population derived from the MM (ref. ). The IM itself is derived from the posterior primitive streak. Although the developmental origin of the kidney is well understood, nephron formation in the human kidney is completed before birth. Hence, there is no postnatal stem cell able to replace lost nephrons. In this study, we have successfully directed the differentiation of human embryonic stem cells (hESCs) through posterior primitive streak and IM under fully chemically defined monolayer culture conditions using growth factors used during normal embryogenesis. This differentiation protocol results in the synchronous induction of UB and MM that forms a self-organizing structure, including nephron formation, in vitro. Such hESC-derived components show broad renal potential ex vivo, illustrating the potential for pluripotent-stem-cell-based renal regeneration.
Organoids: a Special Issue
Publisher: The Company of Biologists
Date: 15-03-2017
DOI: 10.1242/DEV.150292
Abstract: Summary: This Editorial provides an overview of the entire contents of the Special Issue, highlighting some of the important findings and major themes therein.
Two N-Terminal Self-Association Domains Are Required for the Dominant Negative Transcriptional Activity of WT1 Denys-Drash Mutant Proteins
Publisher: Elsevier BV
Date: 04-1997
Abstract: Patients with Denys-Drash syndrome (DDS) have been shown to be constitutionally heterozygous for mutations of the WT1 gene. Almost all DDS mutations inactivate or remove the DNA-binding zinc finger region of WT1 and the resulting mutant proteins appear to act in a dominant negative manner. This may occur via WT1 self-association, which has been shown to involve the first 180 amino acids. By creating a series of N-terminal deletions, we have further investigated WT1 self-association using a yeast di-hybrid system and an in vitro protein binding assay. Our results suggest that there are two distinct domains within the N-terminal region facilitating self-association, residing from amino acids 1-45 and 157-253. Co-transfection of WT1 with progressively shorter N-terminal constructs demonstrates that both of these sites are required for a dominant negative activity as assessed by activation of a reporter construct.
Diving Deep into the Adult Mouse Kidney
Publisher: Elsevier BV
Date: 11-2019
Identification of molecular compartments and genetic circuitry in the developing mammalian kidney
Publisher: The Company of Biologists
Date: 15-05-2012
DOI: 10.1242/DEV.074005
Abstract: Lengthy developmental programs generate cell ersity within an organotypic framework, enabling the later physiological actions of each organ system. Cell identity, cell ersity and cell function are determined by cell type-specific transcriptional programs consequently, transcriptional regulatory factors are useful markers of emerging cellular complexity, and their expression patterns provide insights into the regulatory mechanisms at play. We performed a comprehensive genome-scale in situ expression screen of 921 transcriptional regulators in the developing mammalian urogenital system. Focusing on the kidney, analysis of regional-specific expression patterns identified novel markers and cell types associated with development and patterning of the urinary system. Furthermore, promoter analysis of synexpressed genes predicts transcriptional control mechanisms that regulate cell differentiation. The annotated informational resource (www.gudmap.org) will facilitate functional analysis of the mammalian kidney and provides useful information for the generation of novel genetic tools to manipulate emerging cell populations.
Expression of the vertebrate Slit gene family and their putative receptors, the Robo genes, in the developing murine kidney
Publisher: Elsevier BV
Date: 06-2000
DOI: 10.1016/S0925-4773(00)00313-0
Abstract: The slit (sli) gene, encoding a secreted glycoprotein, has been demonstrated to play a vital role in axonal guidance in Drosophila melanogaster by acting as a signalling ligand for the robo receptor (Rothberg, J.M., Jacobs, J.R., Goodman, C.S., Artavanis-Tsakonas, S., 1990. slit: an extracellular protein necessary for development of midline glia and commissural axon pathways contains both EGF and LRR domains. Genes Dev. 4, 2169-2187 Kidd, T., Bland, K.S., Goodman, C. S., 1999. Slit is the midline repellent for the robo receptor in Drosophila. Cell 96, 785-794). Multiple homologs of both sli and robo have been identified in vertebrates and are thought to play similar roles to their fly counterparts in neural development (Brose, K., Bland, K.S., Wang, K.H., Arnott, D., Henzel, W., Goodman, C.S., Tessier-Lavigne, M., Kidd, T., 1999. Slit proteins bind Robo receptors and have an evolutionarily conserved role in repulsive axon guidance. Cell 96, 795-806). Slit2 has been shown to bind Robo1, mediating both neuronal and axonal guidance in the developing central nervous system (CNS), (Brose et al., 1999 Hu, H., 1999. Chemorepulsion of neuronal migration by Slit2 in the developing mammalian forebrain. Neuron 23, 703-711). Importantly, both gene families display distinct expression patterns outside the CNS (Holmes, G.P., Negus, K., Burridge, L., Raman, S., Algar, E., Yamada, T., Little, M.H., 1998. Distinct but overlapping expression patterns of two vertebrate slit homologs implies functional roles in CNS development and organogenesis. Mech. Dev. 79, 57-72 Yuan, W., Zhou, L., Chen, J.H., Wu, J.Y., Rao, Y., Ornitz, D.M., 1999. The mouse SLIT family: secreted ligands for ROBO expressed in patterns that suggest a role in morphogenesis and axon guidance. Dev. Biol. 212, 290-306). Using in situ hybridization on metanephric explant cultures and urogenital tract sections, the expression patterns of Slit1, 2, 3 and Robo1 and 2 were investigated during murine metanephric development. Slit1 was expressed in the metanephric mesenchyme (MM) surrounding the invading ureteric tree (UT). Slit2 was expressed at the tips of the UT and both Slit2 and Slit3 were expressed at the far proximal end of the comma shaped and S-shaped bodies. Expression of Robo1 was initially diffuse throughout the MM, then upregulated in the pretubular aggregates, and maintained at the distal end of the comma and S-shaped bodies. Robo2 was detected in the induced MM surrounding the arborizing UT tips and later in the proximal end of the S-shaped bodies. Coincident expression of Robo1 with Slit1 in the metanephric mesenchyme and Robo2, Slit2 and Slit3 in the far proximal end of the S-shaped bodies was observed during metanephric development.
Neonatal vascularization and oxygen tension regulate appropriate perinatal renal medulla/papilla maturation
Publisher: Wiley
Date: 17-03-2016
DOI: 10.1002/PATH.4690
Abstract: Congenital medullary dysplasia with obstructive nephropathy is a common congenital disorder observed in paediatric patients and represents the foremost cause of renal failure. However, the molecular processes regulating normal papillary outgrowth during the postnatal period are unclear. In this study, transcriptional profiling of the renal medulla across postnatal development revealed enrichment of non-canonical Wnt signalling, vascular development, and planar cell polarity genes, all of which may contribute to perinatal medulla apilla maturation. These pathways were investigated in a model of papillary hypoplasia with functional obstruction, the Crim1(KST264/KST264) transgenic mouse. Postnatal elongation of the renal papilla via convergent extension was unaffected in the Crim1(KST264/KST264) hypoplastic renal papilla. In contrast, these mice displayed a disorganized papillary vascular network, tissue hypoxia, and elevated Vegfa expression. In addition, we demonstrate the involvement of accompanying systemic hypoxia arising from placental insufficiency, in appropriate papillary maturation. In conclusion, this study highlights the requirement for normal vascular development in collecting duct patterning, development of appropriate nephron architecture, and perinatal papillary maturation, such that disturbances contribute to obstructive nephropathy.
Review article: Potential cellular therapies for renal disease: Can we translate results from animal studies to the human condition?
Publisher: Wiley
Date: 26-08-2009
DOI: 10.1111/J.1440-1797.2009.01144.X
Abstract: The incidence of chronic kidney disease is increasing worldwide, prompting considerable research into potential regenerative therapies. These have included studies to determine whether an endogenous renal stem cell exists in the postnatal kidney and whether non-renal adult stem cells, such as mesenchymal stem cell, can ameliorate renal damage. Such stem cells will either need to be recruited to the damaged kidney to repair the damage in situ or be differentiated into the desired cell type and delivered into the damaged kidney to subsequently elicit repair without maldifferentiation. To date, these studies have largely been performed using experimental and genetic models of renal damage in rodents. The translation of such research into a therapy applicable to human disease faces many challenges. In this review, we examine which animal models have been used to evaluate potential cellular therapies and how valid these are to human chronic kidney disease.
Renal Structural and Functional Repair in a Mouse Model of Reversal of Ureteral Obstruction
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 12-2005
The Wilms' tumour suppressor protein, WT1, undergoes CRM1-independent nucleocytoplasmic shuttling
Publisher: Wiley
Date: 15-10-2003
DOI: 10.1016/S0014-5793(03)01144-X
Abstract: The Wilms' tumour suppressor gene (WT1) encodes a zinc finger-containing nuclear protein essential for kidney and urogenital development. Initially considered a transcription factor, there is mounting evidence that WT1 has a role in post-transcriptional processing. Using the interspecies heterokaryon assay, we have demonstrated that WT1 can undergo nucleocytoplasmic shuttling. We have also mapped the region responsible for nuclear export to residues 182-324. Our data add further complexity to the role of WT1 in transcriptional and post-transcriptional regulation.
Three-dimensional visualization of testis cord morphogenesis, a novel tubulogenic mechanism in development
Publisher: Wiley
Date: 05-2009
DOI: 10.1002/DVDY.21925
Haploinsufficiency for the Six2 gene increases nephron progenitor proliferation promoting branching and nephron number.
Publisher: Elsevier BV
Date: 03-2018
Determining lineage relationships in kidney development and disease
Publisher: Springer Science and Business Media LLC
Date: 30-09-2022
DOI: 10.1038/S41581-021-00485-5
Abstract: The lineage relationships of cells provide information about the origins of component cell types during development and repair as well as the source of aberrant cells during disease. Genetic approaches to lineage tracing applied in the mouse have revealed much about how the mammalian kidney forms, including the identification of key progenitors for the nephrons and stromal compartments. Inducible Cre systems have also facilitated lineage tracing studies in the postnatal animal that illustrate the changes in cellular fate that can occur during kidney injury. With the advent of single-cell transcriptional profiling and trajectory analyses, predictions of cellular relationships across development are now being made in model systems, such as the mouse, as well as in human fetal kidney. Importantly, these approaches provide predictions of lineage relationships rather than definitive evidence. Although genetic approaches to the study of lineage have not previously been possible in a human setting, the application of CRISPR-Cas9 gene editing of pluripotent stem cells is beginning to teach us about human lineage relationships.
Stromal Protein Ecm1 Regulates Ureteric Bud Patterning and Branching
Publisher: Public Library of Science (PLoS)
Date: 31-12-2013
Stromal cells in tissue homeostasis: balancing regeneration and fibrosis
Publisher: Springer Science and Business Media LLC
Date: 13-08-2013
Abstract: The ageing population and the increasing prevalence of noncommunicable diseases such as diabetes and hypertension have led to an increased prevalence of chronic kidney disease. The generation of de novo kidney tissue from embryonic tissue and stem cells using tissue engineering approaches is being explored as an alternative to renal replacement therapy for treating the disease. It is, however, becoming clear that resident cells can not only induce fibrotic repair, but can also restore damaged kidney tissue. Mobilizing this innate capacity of the kidney to regenerate is of particular interest in the prevention of irreversible kidney failure. A novel concept is that the interaction of interstitial stromal cells with the local immune system may regulate tissue homeostasis and the balance between tissue repair and fibrosis. Mesenchymal stromal cells (MSCs), in particular, may enhance the intrinsic reparative capabilities of the kidney. This Perspectives article considers the innate regenerative potential of the kidney in the context of ongoing studies of MSC therapy.
Expression of Crim1 during murine ocular development
Publisher: Elsevier BV
Date: 06-2000
DOI: 10.1016/S0925-4773(00)00292-6
Abstract: Crim1 (cysteine-rich motor neuron 1), a novel gene encoding a putative transmembrane protein, has recently been isolated and characterized (Kolle, G., Georgas, K., Holmes, G.P., Little, M.H., Yamada, T., 2000. CRIM1, a novel gene encoding a cysteine-rich repeat protein, is developmentally regulated and implicated in vertebrate CNS development and organogenesis. Mech. Dev. 90, 181-193). Crim1 contains an IGF-binding protein motif and multiple cysteine-rich repeats, analogous to those of chordin and short gastrulation (sog) proteins that associate with TGFbeta superfamily members, namely Bone Morphogenic Protein (BMP). High levels of Crim1 have been detected in the brain, spinal chord and lens. As members of the IGF and TGFbeta growth factor families have been shown to influence the behaviour of lens cells (Chamberlain, C.G., McAvoy, J. W., 1997. Fibre differentiation and polarity in the mammalian lens: a key role for FGF. Prog. Ret. Eye Res. 16, 443-478 de Iongh R.U., Lovicu, F.J., Overbeek, P.A., Schneider, M.D., McAvoy J.W., 1999. TGF-beta signalling is essential for terminal differentiation of lens fibre cells. Invest. Ophthalmol. Vis. Sci. 40, S561), to further understand the role of Crim1 in the lens, its expression during ocular morphogenesis and growth is investigated. Using in situ hybridisation, the expression patterns of Crim1 are determined in murine eyes from embryonic day 9.5 through to postnatal day 21. Low levels of transcripts for Crim1 are first detected in the lens placode. By the lens pit stage, Crim1 is markedly upregulated with high levels persisting throughout embryonic and foetal development. Crim1 is expressed in both lens epithelial and fibre cells. As lens fibres mature in the nucleus, Crim1 is downregulated but strong expression is maintained in the lens epithelium and in the young fibre cells of the lens cortex. Crim1 is also detected in other developing ocular tissues including corneal and conjunctival epithelia, corneal endothelium, retinal pigmented epithelium, ciliary and iridial retinae and ganglion cells. During postnatal development Crim1 expression is restricted to the lens, with strongest expression in the epithelium and in the early differentiating secondary fibres. Thus, strong expression of Crim1 is a distinctive feature of the lens during morphogenesis and postnatal growth.
Development of the Human Fetal Kidney from Mid to Late Gestation in Male and Female Infants
Publisher: Elsevier BV
Date: 12-0012
Identification of Anchor Genes during Kidney Development Defines Ontological Relationships, Molecular Subcompartments and Regulatory Pathways
Publisher: Public Library of Science (PLoS)
Date: 28-02-2011
RNA binding by the Wilms tumor suppressor zinc finger proteins.
Publisher: Proceedings of the National Academy of Sciences
Date: 23-07-1996
Abstract: The Wilms tumor suppressor gene WT1 is implicated in the ontogeny of genito-urinary abnormalities, including Denys-Drash syndrome and Wilms tumor of the kidney. WT1 encodes Kruppel-type zinc finger proteins that can regulate the expression of several growth-related genes, apparently by binding to specific DNA sites located within 5' untranslated leader regions as well as 5' promoter sequences. Both WT1 and a closely related early growth response factor, EGR1, can bind the same DNA sequences from the mouse gene encoding insulin-like growth factor 2 (Igf-2). We report that WT1, but not EGR1, can bind specific Igf-2 exonic RNA sequences, and that the zinc fingers are required for this interaction. WT1 zinc finger 1, which is not represented in EGR1, plays a more significant role in RNA binding than zinc finger 4, which does have a counterpart in EGR1. Furthermore, the normal subnuclear localization of WT1 proteins is shown to be RNase, but not DNase, sensitive. Therefore, WT1 might, like the Kruppel-type zinc finger protein TFIIIA, regulate gene expression by both transcriptional and posttranscriptional mechanisms.
Angioblast-mesenchyme induction of early kidney development is mediated by Wt1 and Vegfa
Publisher: The Company of Biologists
Date: 15-12-2005
DOI: 10.1242/DEV.02095
Abstract: Most studies on kidney development have considered the interaction of the metanephric mesenchyme and the ureteric bud to be the major inductive event that maintains tubular differentiation and branching morphogenesis. The mesenchyme produces Gdnf, which stimulates branching, and the ureteric bud stimulates continued growth of the mesenchyme and differentiation of nephrons from the induced mesenchyme. Null mutation of the Wt1 gene eliminates outgrowth of the ureteric bud, but Gdnf has been identified as a target of Pax2, but not of Wt1. Using a novel system for microinjecting and electroporating plasmid expression constructs into murine organ cultures, it has been demonstrated that Vegfa expression in the mesenchyme is regulated by Wt1. Previous studies had identified a population of Flk1-expressing cells in the periphery of the induced mesenchyme, and adjacent to the stalk of the ureteric bud, and that Vegfa was able to stimulate growth of kidneys in organ culture. Here it is demonstrated that signaling through Flk1 is required to maintain expression of Pax2 in the mesenchyme of the early kidney, and for Pax2 to stimulate expression of Gdnf. However, once Gdnf stimulates branching of the ureteric bud, the Flk1-dependent angioblast signal is no longer required to maintain branching morphogenesis and induction of nephrons. Thus,this work demonstrates the presence of a second set of inductive events,involving the mesenchymal and angioblast populations, whereby Wt1-stimulated expression of Vegfa elicits an as-yet-unidentified signal from the angioblasts, which is required to stimulate the expression of Pax2 and Gdnf,which in turn elicits an inductive signal from the ureteric bud.
Mutations in DZIP1L, which encodes a ciliary-transition-zone protein, cause autosomal recessive polycystic kidney disease.
Publisher: Springer Science and Business Media LLC
Date: 22-05-2017
DOI: 10.1038/NG.3871
Hamartin regulates cessation of mouse nephrogenesis independently of Mtor
Publisher: Proceedings of the National Academy of Sciences
Date: 21-05-2018
Abstract: Mammals form the final numbers of nephrons, the functional units of their kidneys, before or within a few days after birth. This initial endowment declines throughout life without the ability to replace nephrons lost to injury. Therefore, humans with low nephron numbers (such as premature infants) have higher rates of kidney disease in adulthood. Here we report that partial reduction in hamartin activity within mouse renal progenitors can lead to a significant increase in nephron numbers in newborns. Nephrogenesis was prolonged by at least a day and correlated with higher engraftment rates in the developing niche in a stem cell engraftment assay. We provide evidence that this phenotype is independent of the inhibitory effect of hamartin on the Mtor complex.
Kidney organoids from human iPS cells contain multiple lineages and model human nephrogenesis
Publisher: Springer Science and Business Media LLC
Date: 07-10-2015
DOI: 10.1038/NATURE15695
Abstract: The human kidney contains up to 2 million epithelial nephrons responsible for blood filtration. Regenerating the kidney requires the induction of the more than 20 distinct cell types required for excretion and the regulation of pH, and electrolyte and fluid balance. We have previously described the simultaneous induction of progenitors for both collecting duct and nephrons via the directed differentiation of human pluripotent stem cells. Paradoxically, although both are of intermediate mesoderm in origin, collecting duct and nephrons have distinct temporospatial origins. Here we identify the developmental mechanism regulating the preferential induction of collecting duct versus kidney mesenchyme progenitors. Using this knowledge, we have generated kidney organoids that contain nephrons associated with a collecting duct network surrounded by renal interstitium and endothelial cells. Within these organoids, in idual nephrons segment into distal and proximal tubules, early loops of Henle, and glomeruli containing podocytes elaborating foot processes and undergoing vascularization. When transcription profiles of kidney organoids were compared to human fetal tissues, they showed highest congruence with first trimester human kidney. Furthermore, the proximal tubules endocytose dextran and differentially apoptose in response to cisplatin, a nephrotoxicant. Such kidney organoids represent powerful models of the human organ for future applications, including nephrotoxicity screening, disease modelling and as a source of cells for therapy.
Mammalian Kidney Development: Principles, Progress, and Projections
Publisher: Cold Spring Harbor Laboratory
Date: 05-2012
Regenerative medicine in kidney disease
Publisher: Elsevier BV
Date: 08-2016
DOI: 10.1016/J.KINT.2016.03.030
Abstract: The treatment of renal failure has changed little in decades. Organ transplantation and dialysis continue to represent the only therapeutic options available. However, decades of fundamental research into the response of the kidney to acute injury and the processes driving progression to chronic kidney disease are beginning to open doors to new options. Similarly, continued investigations into the cellular and molecular basis of normal kidney development, together with major advances in stem cell biology, are now delivering options in regenerative medicine not possible as recently as a decade ago. In this review, we will discuss advances in regenerative medicine as it may be applied to the kidney. This will cover cellular therapies focused on ameliorating injury and improving repair as well as advancements in the generation of new renal tissue from stem rogenitor cells.
Is There Such a Thing as a Renal Stem Cell?
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 10-2009
Genetically engineering endothelial niche in human kidney organoids enables multilineage maturation, vascularization and de novo cell types
Publisher: Cold Spring Harbor Laboratory
Date: 30-05-2023
DOI: 10.1101/2023.05.30.542848
Abstract: Vascularization plays a critical role in organ maturation and cell type development. Drug discovery, organ mimicry, and ultimately transplantation in a clinical setting thereby hinges on achieving robust vascularization of in vitro engineered organs. Here, focusing on human kidney organoids, we overcome this hurdle by combining an inducible ETS translocation variant 2 ( ETV2 ) human induced pluripotent stem cell (iPSC) line, which directs endothelial fate, with a non-transgenic iPSC line in suspension organoid culture. The resulting human kidney organoids show extensive vascularization by endothelial cells with an identity most closely related to endogenous kidney endothelia. Vascularized organoids also show increased maturation of nephron structures including more mature podocytes with improved marker expression, foot process interdigitation, an associated fenestrated endothelium, and the presence of renin + cells. The creation of an engineered vascular niche capable of improving kidney organoid maturation and cell type complexity is a significant step forward in the path to clinical translation. Furthermore, this approach is orthogonal to native tissue differentiation paths, hence readily adaptable to other organoid systems and thus has the potential for a broad impact on basic and translational organoid studies. Developing therapies for patients with kidney diseases relies on a morphologically and physiologically representative in vitro model. Human kidney organoids are an attractive model to recapitulate kidney physiology, however, they are limited by the absence of a vascular network and mature cell populations. In this work, we have generated a genetically inducible endothelial niche that, when combined with an established kidney organoid protocol, induces the maturation of a robust endothelial cell network, induces a more mature podocyte population, and induces the emergence a functional renin population. This advance significantly increases the clinical relevance of human kidney organoids for etiological studies of kidney disease and future regenerative medicine strategies. Genetically engineered endothelial niche induces mature cell populations in human kidney organoids
Overexpression of a Slit Homologue Impairs Convergent Extension of the Mesoderm and Causes Cyclopia in Embryonic Zebrafish
Publisher: Elsevier BV
Date: 02-2001
DNA Methyltransferase 1 Controls Nephron Progenitor Cell Renewal and Differentiation
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 05-12-2018
Abstract: Nephron number is a major determinant of long-term renal function and cardiovascular risk. Observational studies suggest that maternal nutritional and metabolic factors during gestation contribute to the high variability of nephron endowment. However, the underlying molecular mechanisms have been unclear. We used mouse models, including DNA methyltransferase ( Dnmt1, Dnmt3a, and Dnmt3b ) knockout mice, optical projection tomography, three-dimensional reconstructions of the nephrogenic niche, and transcriptome and DNA methylation analysis to characterize the role of DNA methylation for kidney development. We demonstrate that DNA hypomethylation is a key feature of nutritional kidney growth restriction in vitro and in vivo, and that DNA methyltransferases Dnmt1 and Dnmt3a are highly enriched in the nephrogenic zone of the developing kidneys. Deletion of Dnmt1 in nephron progenitor cells (in contrast to deletion of Dnmt3a or Dnm3b ) mimics nutritional models of kidney growth restriction and results in a substantial reduction of nephron number as well as renal hypoplasia at birth. In Dnmt1 -deficient mice, optical projection tomography and three-dimensional reconstructions uncovered a significant reduction of stem cell niches and progenitor cells. RNA sequencing analysis revealed that global DNA hypomethylation interferes in the progenitor cell regulatory network, leading to downregulation of genes crucial for initiation of nephrogenesis, Wt1 and its target Wnt4. Derepression of germline genes, protocadherins, Rhox genes, and endogenous retroviral elements resulted in the upregulation of IFN targets and inhibitors of cell cycle progression. These findings establish DNA methylation as a key regulatory event of prenatal renal programming, which possibly represents a fundamental link between maternal nutritional factors during gestation and reduced nephron number.
Analysed cap mesenchyme track data from live imaging of mouse kidney development
Publisher: Elsevier BV
Date: 12-2016
Isolation of clonogenic, long-term self renewing embryonic renal stem cells
Publisher: Elsevier BV
Date: 07-2010
DOI: 10.1016/J.SCR.2010.03.003
Abstract: A tissue stem cell should exhibit long-term self-renewal, clonogenicity and a capacity to differentiate into the tissue of origin. Such a postnatal renal stem cell has not been formally identified. The metanephric mesenchyme (MM) of the developing kidney gives rise to both the renal interstitium and the nephrons and is regarded as the progenitor population of the developing kidney. However, isolated MM does not self renew and requires immortalization for survival in culture. Here we report the isolation and sustained culture of long-term repopulating, clonal progenitors from the embryonic kidney as free floating nephrospheres. Such cells displayed clonal self renewal for in excess of twenty passages when cultured with bFGF and thrombin, showed broad mesodermal multipotentiality, but retained expression of key renal transcription factors (Wt1, Sall1, Eya1, Six1, Six2, Osr1 and Hoxa11). While these cells did display limited capacity to contribute to developing embryonic kidney explants, nephrospheres did not display in vitro renal epithelial capacity. Nephrospheres could be cultured from both Sall1(+) and Sall1(-) fractions of embryonic kidney, suggesting that they were derived from the MM as a whole and not specifically the MM-derived cap mesenchyme committed to nephron formation. This embryonic renal stem cell population was not able to be isolated from postnatal kidney confirming that while the embryonic MM represents a mulitpotent stem cell population, this does not persist after birth.
Atlas of Gene Expression in the Developing Kidney at Microanatomic Resolution
Publisher: Elsevier BV
Date: 11-2008
DevKidCC allows for robust classification and direct comparisons of kidney organoid datasets
Publisher: Springer Science and Business Media LLC
Date: 22-02-2022
DOI: 10.1186/S13073-022-01023-Z
Abstract: While single-cell transcriptional profiling has greatly increased our capacity to interrogate biology, accurate cell classification within and between datasets is a key challenge. This is particularly so in pluripotent stem cell-derived organoids which represent a model of a developmental system. Here, clustering algorithms and selected marker genes can fail to accurately classify cellular identity while variation in analyses makes it difficult to meaningfully compare datasets. Kidney organoids provide a valuable resource to understand kidney development and disease. However, direct comparison of relative cellular composition between protocols has proved challenging. Hence, an unbiased approach for classifying cell identity is required. The R package, scPred , was trained on multiple single cell RNA-seq datasets of human fetal kidney. A hierarchical model classified cellular subtypes into nephron, stroma and ureteric epithelial elements. This model, provided in the R package DevKidCC ( github.com/KidneyRegeneration/DevKidCC ), was then used to predict relative cell identity within published kidney organoid datasets generated using distinct cell lines and differentiation protocols, interrogating the impact of such variations. The package contains custom functions for the display of differential gene expression within cellular subtypes. DevKidCC was used to directly compare between distinct kidney organoid protocols, identifying differences in relative proportions of cell types at all hierarchical levels of the model and highlighting variations in stromal and unassigned cell types, nephron progenitor prevalence and relative maturation of in idual epithelial segments. Of note, DevKidCC was able to distinguish distal nephron from ureteric epithelium, cell types with overlapping profiles that have previously confounded analyses. When applied to a variation in protocol via the addition of retinoic acid, DevKidCC identified a consequential depletion of nephron progenitors. The application of DevKidCC to kidney organoids reproducibly classifies component cellular identity within distinct single-cell datasets. The application of the tool is summarised in an interactive Shiny application, as are ex les of the utility of in-built functions for data presentation. This tool will enable the consistent and rapid comparison of kidney organoid protocols, driving improvements in patterning to kidney endpoints and validating new approaches.
Related Organisations
Novo Nordisk Foundation Centre For Stem Cell Medicine (reNEW)
Location: Netherlands
University Of Queensland Institute For Molecular Bioscience
Location: Australia
Western General Hospital
Location: United Kingdom of Great Britain and Northern Ireland
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