Did you mean: pronephri
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Nature Biotechnology Feb 2023Directed differentiation of human pluripotent stem cells (hPSCs) into functional ureteric and collecting duct (CD) epithelia is essential to kidney regenerative...
Directed differentiation of human pluripotent stem cells (hPSCs) into functional ureteric and collecting duct (CD) epithelia is essential to kidney regenerative medicine. Here we describe highly efficient, serum-free differentiation of hPSCs into ureteric bud (UB) organoids and functional CD cells. The hPSCs are first induced into pronephric progenitor cells at 90% efficiency and then aggregated into spheres with a molecular signature similar to the nephric duct. In a three-dimensional matrix, the spheres form UB organoids that exhibit branching morphogenesis similar to the fetal UB and correct distal tip localization of RET expression. Organoid-derived cells incorporate into the UB tips of the progenitor niche in chimeric fetal kidney explant culture. At later stages, the UB organoids differentiate into CD organoids, which contain >95% CD cell types as estimated by single-cell RNA sequencing. The CD epithelia demonstrate renal electrophysiologic functions, with ENaC-mediated vectorial sodium transport by principal cells and V-type ATPase proton pump activity by FOXI1-induced intercalated cells.
Topics: Humans; Kidney; Ureter; Cell Differentiation; Organoids; Pluripotent Stem Cells; Morphogenesis; Forkhead Transcription Factors
PubMed: 36038632
DOI: 10.1038/s41587-022-01429-5 -
Developmental Dynamics : An Official... Jan 2003Initiation of excretory system development in all vertebrates requires (1) delamination of the pronephric and pronephric duct rudiments from intermediate mesoderm at the... (Review)
Review
Initiation of excretory system development in all vertebrates requires (1) delamination of the pronephric and pronephric duct rudiments from intermediate mesoderm at the ventral border of anterior somites, and (2) extension of the pronephric duct to the cloaca. Pronephric duct extension is the central event in nephric system development; the pronephric duct differentiates into the tubule that carries nephric filtrate out of the body and induces terminal differentiation of adult kidneys. Early studies concluded that pronephric ducts formed by means of in situ segregation of pronephric duct tissue from lateral mesoderm ventral to the forming somites; more recent studies highlight caudal migration of the pronephric duct as the major morphogenetic mechanism. The purpose of this review is to provide the historical background on studies of the mechanisms of amphibian pronephric duct extension, to review evidence showing that different amphibians perform pronephric duct morphogenesis in different ways, and to suggest future studies that may help illuminate the molecular basis of the mechanisms that have evolved in amphibians to extend the pronephric duct to the cloaca.
Topics: Ambystoma; Animals; Cell Movement; Embryo, Nonmammalian; In Situ Hybridization; Kidney; Models, Biological; Morphogenesis; Time Factors; Wolffian Ducts; Xenopus laevis
PubMed: 12508219
DOI: 10.1002/dvdy.10205 -
Cell Discovery Mar 2022Primary cilia are antenna-like subcellular structures to act as signaling platforms to regulate many cellular processes and embryonic development. mA RNA modification...
Primary cilia are antenna-like subcellular structures to act as signaling platforms to regulate many cellular processes and embryonic development. mA RNA modification plays key roles in RNA metabolism and gene expression; however, the physiological function of mA modification remains largely unknown. Here we find that the mA demethylase ALKBH3 significantly inhibits ciliogenesis in mammalian cells by its demethylation activity. Mechanistically, ALKBH3 removes mA sites on mRNA of Aurora A, a master suppressor of ciliogenesis. Depletion of ALKBH3 enhances Aurora A mRNA decay and inhibits its translation. Moreover, alkbh3 morphants exhibit ciliary defects, including curved body, pericardial edema, abnormal otoliths, and dilation in pronephric ducts in zebrafish embryos, which are significantly rescued by wild-type alkbh3, but not by its catalytically inactive mutant. The ciliary defects caused by ALKBH3 depletion in both vertebrate cells and embryos are also significantly reversed by ectopic expression of Aurora A mRNA. Together, our data indicate that ALKBH3-dependent mA demethylation has a crucial role in the regulation of Aurora A mRNA, which is essential for ciliogenesis and cilia-associated developmental events in vertebrates.
PubMed: 35277482
DOI: 10.1038/s41421-022-00385-3 -
Disease Models & Mechanisms Dec 2022Meckel syndrome, nephronophthisis, Joubert syndrome and Bardet-Biedl syndrome are caused by mutations in proteins that localize to the ciliary transition zone (TZ). The...
Meckel syndrome, nephronophthisis, Joubert syndrome and Bardet-Biedl syndrome are caused by mutations in proteins that localize to the ciliary transition zone (TZ). The phenotypically distinct syndromes suggest that these TZ proteins have differing functions. However, mutations in a single TZ gene can result in multiple syndromes, suggesting that the phenotype is influenced by modifier genes. We performed a comprehensive analysis of ten zebrafish TZ mutants, including mks1, tmem216, tmem67, rpgrip1l, cc2d2a, b9d2, cep290, tctn1, nphp1 and nphp4, as well as mutants in ift88 and ift172. Our data indicate that variations in phenotypes exist between different TZ mutants, supporting different tissue-specific functions of these TZ genes. Further, we observed phenotypic variations within progeny of a single TZ mutant, reminiscent of multiple disease syndromes being associated with mutations in one gene. In some mutants, the dynamics of the phenotype became complex with transitory phenotypes that are corrected over time. We also demonstrated that multiple-guide-derived CRISPR/Cas9 F0 'crispant' embryos recapitulate zygotic null phenotypes, and rapidly identified ciliary phenotypes in 11 cilia-associated gene candidates (ankfn1, ccdc65, cfap57, fhad1, nme7, pacrg, saxo2, c1orf194, ttc26, zmynd12 and cfap52).
Topics: Animals; Cilia; Zebrafish; Penetrance; Syndrome; Polycystic Kidney Diseases; Biological Variation, Population; Zebrafish Proteins; Vesicular Transport Proteins
PubMed: 36533556
DOI: 10.1242/dmm.049568 -
Journal of the American Society of... Mar 2021Galloway-Mowat syndrome (GAMOS) is characterized by neurodevelopmental defects and a progressive nephropathy, which typically manifests as steroid-resistant nephrotic...
BACKGROUND
Galloway-Mowat syndrome (GAMOS) is characterized by neurodevelopmental defects and a progressive nephropathy, which typically manifests as steroid-resistant nephrotic syndrome. The prognosis of GAMOS is poor, and the majority of children progress to renal failure. The discovery of monogenic causes of GAMOS has uncovered molecular pathways involved in the pathogenesis of disease.
METHODS
Homozygosity mapping, whole-exome sequencing, and linkage analysis were used to identify mutations in four families with a GAMOS-like phenotype, and high-throughput PCR technology was applied to 91 individuals with GAMOS and 816 individuals with isolated nephrotic syndrome. and studies determined the functional significance of the mutations identified.
RESULTS
Three biallelic variants of the transcriptional regulator were detected in six families with proteinuric kidney disease. Four families with a variant in the protein's zinc-finger (ZNF) domain have additional GAMOS-like features, including brain anomalies, cardiac defects, and skeletal defects. All variants destabilize the PRDM15 protein, and the ZNF variant additionally interferes with transcriptional activation. Morpholino oligonucleotide-mediated knockdown of Prdm15 in embryos disrupted pronephric development. Human wild-type RNA rescued the disruption, but the three variants did not. Finally, CRISPR-mediated knockout of in human podocytes led to dysregulation of several renal developmental genes.
CONCLUSIONS
Variants in can cause either isolated nephrotic syndrome or a GAMOS-type syndrome on an allelic basis. PRDM15 regulates multiple developmental kidney genes, and is likely to play an essential role in renal development in humans.
Topics: Amino Acid Sequence; Amino Acid Substitution; Animals; Cell Line; Child, Preschool; DNA-Binding Proteins; Female; Gene Expression Regulation, Developmental; Gene Knockdown Techniques; Gene Knockout Techniques; Hernia, Hiatal; High-Throughput Nucleotide Sequencing; Humans; Infant; Infant, Newborn; Male; Microcephaly; Models, Molecular; Mutation, Missense; Nephrosis; Nephrotic Syndrome; Podocytes; Polymorphism, Single Nucleotide; Pronephros; Protein Stability; Transcription Factors; Xenopus laevis; Zinc Fingers
PubMed: 33593823
DOI: 10.1681/ASN.2020040490 -
Scientific Reports Oct 2023CLIC5 belongs to a family of ion channels with six members reported so far. In vertebrates, the CLIC5 gene encodes two different isoforms, CLIC5A and CLIC5B. In addition...
CLIC5 belongs to a family of ion channels with six members reported so far. In vertebrates, the CLIC5 gene encodes two different isoforms, CLIC5A and CLIC5B. In addition to its ion channel activity, there is evidence for further functions of CLIC5A, such as the remodeling of the actin cytoskeleton during the formation of a functional glomerulus in the vertebrate kidney. However, its specific role is still incompletely understood and a specific functional role for CLIC5B has not been described yet. Here we report our findings on the differential expression and functions of Clic5a and Clic5b during zebrafish kidney development. Whole-mount in situ hybridization studies revealed specific expression of clic5a in the eye and pronephric glomerulus, and clic5b is expressed in the gut, liver and the pronephric tubules. Clic5 immunostainings revealed that Clic5b is localized in the cilia. Whereas knockdown of Clic5a resulted in leakiness of the glomerular filtration barrier, Clic5b deficient embryos displayed defective ciliogenesis, leading to ciliopathy-associated phenotypes such as ventral body curvature, otolith deposition defects, altered left-right asymmetry and formation of hydrocephalus and pronephric cysts. In addition, Clic5 deficiency resulted in dysregulation of cilia-dependent Wnt signalling pathway components. Mechanistically, we identified a Clic5-dependent activation of the membrane-cytoskeletal linker proteins Ezrin/Radixin/Moesin (ERM) in the pronephric tubules of zebrafish. In conclusion, our in vivo data demonstrates a novel role for Clic5 in regulating essential ciliary functions and identified Clic5 as a positive regulator of ERM phosphorylation.
Topics: Animals; Actin Cytoskeleton; Chloride Channels; Chlorides; Cilia; Kidney Glomerulus; Microfilament Proteins; Zebrafish
PubMed: 37848494
DOI: 10.1038/s41598-023-44235-y -
Scientific Reports Feb 2017The human ubiquitous protein cystinosin is responsible for transporting the disulphide amino acid cystine from the lysosomal compartment into the cytosol. In humans,...
The human ubiquitous protein cystinosin is responsible for transporting the disulphide amino acid cystine from the lysosomal compartment into the cytosol. In humans, Pathogenic mutations of CTNS lead to defective cystinosin function, intralysosomal cystine accumulation and the development of cystinosis. Kidneys are initially affected with generalized proximal tubular dysfunction (renal Fanconi syndrome), then the disease rapidly affects glomeruli and progresses towards end stage renal failure and multiple organ dysfunction. Animal models of cystinosis are limited, with only a Ctns knockout mouse reported, showing cystine accumulation and late signs of tubular dysfunction but lacking the glomerular phenotype. We established and characterized a mutant zebrafish model with a homozygous nonsense mutation (c.706 C > T; p.Q236X) in exon 8 of ctns. Cystinotic mutant larvae showed cystine accumulation, delayed development, and signs of pronephric glomerular and tubular dysfunction mimicking the early phenotype of human cystinotic patients. Furthermore, cystinotic larvae showed a significantly increased rate of apoptosis that could be ameliorated with cysteamine, the human cystine depleting therapy. Our data demonstrate that, ctns gene is essential for zebrafish pronephric podocyte and proximal tubular function and that the ctns-mutant can be used for studying the disease pathogenic mechanisms and for testing novel therapies for cystinosis.
Topics: Amino Acid Sequence; Amino Acid Transport Systems, Neutral; Animals; Apoptosis; Cystine; Cystinosis; Disease Models, Animal; Gene Knockout Techniques; Glomerular Filtration Rate; Humans; Kidney Glomerulus; Kidney Tubules, Proximal; Locomotion; Lysosomes; Mutation; Phenotype; Podocytes; Zebrafish
PubMed: 28198397
DOI: 10.1038/srep42583 -
Kidney International Jan 2023The kidney is an essential organ that ensures bodily fluid homeostasis and removes soluble waste products from the organism. Nephrons, the functional units of the...
The kidney is an essential organ that ensures bodily fluid homeostasis and removes soluble waste products from the organism. Nephrons, the functional units of the kidney, comprise a blood filter, the glomerulus or glomus, and an epithelial tubule that processes the filtrate from the blood or coelom and selectively reabsorbs solutes, such as sugars, proteins, ions, and water, leaving waste products to be eliminated in the urine. Genes coding for transporters are segmentally expressed, enabling the nephron to sequentially process the filtrate. The Xenopus embryonic kidney, the pronephros, which consists of a single large nephron, has served as a valuable model to identify genes involved in nephron formation and patterning. Therefore, the developmental patterning program that generates these segments is of great interest. Prior work has defined the gene expression profiles of Xenopus nephron segments via in situ hybridization strategies, but a comprehensive understanding of the cellular makeup of the pronephric kidney remains incomplete. Here, we carried out single-cell mRNA sequencing of the functional Xenopus pronephric nephron and evaluated its cellular composition through comparative analyses with previous Xenopus studies and single-cell mRNA sequencing of the adult mouse kidney. This study reconstructs the cellular makeup of the pronephric kidney and identifies conserved cells, segments, and associated gene expression profiles. Thus, our data highlight significant conservation in podocytes, proximal and distal tubule cells, and divergence in cellular composition underlying the capacity of each nephron to remove wastes in the form of urine, while emphasizing the Xenopus pronephros as a model for physiology and disease.
Topics: Animals; Mice; Gene Expression Regulation, Developmental; Kidney; Kidney Glomerulus; Nephrons; RNA, Messenger; Xenopus laevis
PubMed: 36055600
DOI: 10.1016/j.kint.2022.07.027 -
Pediatric Nephrology (Berlin, Germany) Apr 2014The main functions of the kidney are to excrete metabolic waste products and actively reabsorb essential molecules such as amino acids, ions, glucose and water. In... (Review)
Review
The main functions of the kidney are to excrete metabolic waste products and actively reabsorb essential molecules such as amino acids, ions, glucose and water. In humans, a wide range of genetic disorders exist characterized by wasting of metabolically important compounds. At the cellular level, more than 20 highly specialized renal epithelial cell types located in different segments of the nephron contribute to the reabsorption process. In particular, proximal tubular cells play a crucial role and are uniquely adapted to maximize reabsorption efficiency. They accommodate high numbers of transporters and channels by increasing the apical surface area in contact with the primary filtrate by forming a brush border as well as undergoing hypertrophy and hyperplasia. This adaptation is evolutionarily conserved and is detected in the primitive pronephric kidney of fish and amphibians as well as the metanephric kidney of higher vertebrates. Surprisingly, signaling pathways regulating these three processes have remained largely unknown. Here we summarize recent studies that highlight the early phases of kidney development as a critical juncture in establishing proximal tubule size.
Topics: Animals; Humans; Nephrons
PubMed: 23974984
DOI: 10.1007/s00467-013-2581-x -
Cells Sep 2015The nephron is the basic structural and functional unit of the vertebrate kidney. To ensure kidney functions, the nephrons possess a highly segmental organization where... (Review)
Review
The nephron is the basic structural and functional unit of the vertebrate kidney. To ensure kidney functions, the nephrons possess a highly segmental organization where each segment is specialized for the secretion and reabsorption of particular solutes. During embryogenesis, nephron progenitors undergo a mesenchymal-to-epithelial transition (MET) and acquire different segment-specific cell fates along the proximo-distal axis of the nephron. Even if the morphological changes occurring during nephrogenesis are characterized, the regulatory networks driving nephron segmentation are still poorly understood. Interestingly, several studies have shown that the pronephric nephrons in Xenopus and zebrafish are segmented in a similar fashion as the mouse metanephric nephrons. Here we review functional and molecular aspects of nephron segmentation with a particular interest on the signaling molecules and transcription factors recently implicated in kidney development in these three different vertebrate model organisms. A complete understanding of the mechanisms underlying nephrogenesis in different model organisms will provide novel insights on the etiology of several human renal diseases.
PubMed: 26378582
DOI: 10.3390/cells4030483