Nephron formation adopts a novel spatial topology at cessation of nephrogenesis

Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Australia.
Developmental Biology (Impact Factor: 3.55). 09/2011; 360(1):110-22. DOI: 10.1016/j.ydbio.2011.09.011
Source: PubMed


Nephron number in the mammalian kidney is known to vary dramatically, with postnatal renal function directly influenced by nephron complement. What determines final nephron number is poorly understood but nephron formation in the mouse kidney ceases within the first few days after birth, presumably due to the loss of all remaining nephron progenitors via epithelial differentiation. What initiates this event is not known. Indeed, whether nephron formation occurs in the same way at this time as during embryonic development has also not been examined. In this study, we investigate the key cellular compartments involved in nephron formation; the ureteric tip, cap mesenchyme and early nephrons; from postnatal day (P) 0 to 6 in the mouse. High resolution analyses of gene and protein expression indicate that loss of nephron progenitors precedes loss of ureteric tip identity, but show spatial shifts in the expression of cap mesenchyme genes during this time. In addition, cap mesenchymal volume and rate of proliferation decline prior to birth. Section-based 3D modeling and Optical Projection Tomography revealed a burst of ectopic nephron induction, with the formation of multiple (up to 5) nephrons per ureteric tip evident from P2. While the distal-proximal patterning of these nephrons occurred normally, their spatial relationship with the ureteric compartment was altered. We propose that this phase of nephron formation represents an acceleration of differentiation within the cap mesenchyme due to a displacement of signals within the nephrogenic niche.

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    • "Primers used to evaluate the excision of Dicer exon 24 are listed in Table S6. Section in situ hybridization validation of the microarray was performed as previously described using riboprobes that were PCR-synthesized from wholeembryo cDNA (Rumballe et al. 2011). Sequencing was performed using T7 primers, followed by nucleotide- BLAST to confirm gene specificity. "
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    ABSTRACT: MicroRNAs are small noncoding RNAs that post-transcriptionally regulate mRNA levels. While previous studies have demonstrated that miRNAs are indispensable in the nephron progenitor and ureteric bud lineage, little is understood about stromal miRNAs during kidney development. The renal stroma (marked by expression of FoxD1) gives rise to the renal interstitium, a subset of peritubular capillaries, and multiple supportive vascular cell types including pericytes and the glomerular mesangium. In this study, we generated FoxD1(GC);Dicer(fl/fl) transgenic mice that lack miRNA biogenesis in the FoxD1 lineage. Loss of Dicer activity resulted in multifaceted renal anomalies including perturbed nephrogenesis, expansion of nephron progenitors, decreased renin-expressing cells, fewer smooth muscle afferent arterioles, and progressive mesangial cell loss in mature glomeruli. Although the initial lineage specification of FoxD1(+) stroma was not perturbed, both the glomerular mesangium and renal interstitium exhibited ectopic apoptosis, which was associated with increased expression of Bcl2l11 (Bim) and p53 effector genes (Bax, Trp53inp1, Jun, Cdkn1a, Mmp2, and Arid3a). Using a combination of high-throughput miRNA profiling of the FoxD1(+)-derived cells and mRNA profiling of differentially expressed transcripts in FoxD1(GC);Dicer(fl/fl) kidneys, at least 72 miRNA:mRNA target interactions were identified to be suppressive of the apoptotic program. Together, the results support an indispensable role for stromal miRNAs in the regulation of apoptosis during kidney development.
    10/2015; 3(10). DOI:10.14814/phy2.12537
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    • "Cessation of nephrogenesis is defined as the final round of new nephron formation in which the last wave of NPCs undergoes mesenchyme to epithelial transition. In the mouse, this occurs shortly after birth and is accompanied by a reduction in Cited1+ cap mesenchyme by P2 (Figure 1B) (Hartman et al., 2007; Rumballe et al., 2011). We reasoned that SMAD1/5 signaling might increase during the terminal phase of nephrogenesis, skewing the renewal versus differentiation balance and depleting the cap mesenchyme. "
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    ABSTRACT: FGF, BMP, and WNT balance embryonic nephron progenitor cell (NPC) renewal and differentiation. By modulating these pathways, we have created an in vitro niche in which NPCs from embryonic kidneys or derived from human embryonic stem cells can be propagated. NPC cultures expanded up to one billion-fold in this environment can be induced to form tubules expressing nephron differentiation markers. Single-cell culture reveals phenotypic variability within the early CITED1-expressing NPC compartment, indicating that it is a mixture of cells with varying progenitor potential. Furthermore, we find that the developmental age of NPCs does not correlate with propagation capacity, indicating that cessation of nephrogenesis is related to factors other than an intrinsic clock. This in vitro nephron progenitor niche will have important applications for expansion of cells for engraftment and will facilitate investigation of mechanisms that determine the balance between renewal and differentiation in these cells. Copyright © 2015 Elsevier Inc. All rights reserved.
    Developmental Cell 07/2015; 34(2). DOI:10.1016/j.devcel.2015.06.021 · 9.71 Impact Factor
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    • "Nephron abundance varies amongst individuals and populations, with a demonstrated influence of genetics and maternal nutritional status on nephron number in humans (Benz and Amann, 2010; Hoy et al., 2006; Luyckx and Brenner, 2010; Wlodek et al., 2008). Self-renewal of NPCs ensures a supply of cells for nephrogenesis until the cessation of nephrogenesis at postnatal day 4 in mice and at 35 weeks of gestation in humans (Hartman et al., 2007; Rumballe et al., 2011). The nephron progenitor niche includes the cap mesenchyme (CM), in which the NPCs reside, and provides extrinsic cues, such as nutrients and morphogens that drive cell intrinsic signaling and metabolic pathways, to promote the availability of NPCs for subsequent nephrogenesis. "
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    ABSTRACT: Contrary to its classic role in restraining cell proliferation, we demonstrate here a divergent function of p53 in the maintenance of self-renewal of the nephron progenitor pool in the embryonic mouse kidney. Nephron endowment is regulated by progenitor availability and differentiation potential. Conditional deletion of p53 in nephron progenitor cells (Six2Cre(+);p53(fl/fl)) induces progressive depletion of Cited1(+)/Six2(+) self-renewing progenitors and loss of cap mesenchyme (CM) integrity. The Six2(p53-null) CM is disorganized, with interspersed stromal cells and an absence of a distinct CM-epithelia and CM-stroma interface. Impaired cell adhesion and epithelialization are indicated by decreased E-cadherin and NCAM expression and by ineffective differentiation in response to Wnt induction. The Six2Cre(+);p53(fl/fl) cap has 30% fewer Six2(GFP(+)) cells. Apoptotic index is unchanged, whereas proliferation index is significantly reduced in accordance with cell cycle analysis showing disproportionately fewer Six2Cre(+);p53(fl/fl) cells in the S and G2/M phases compared with Six2Cre(+);p53(+/+) cells. Mutant kidneys are hypoplastic with fewer generations of nascent nephrons. A significant increase in mean arterial pressure is observed in early adulthood in both germline and conditional Six2(p53-null) mice, linking p53-mediated defects in kidney development to hypertension. RNA-Seq analyses of FACS-isolated wild-type and Six2(GFP(+)) CM cells revealed that the top downregulated genes in Six2Cre(+);p53(fl/fl) CM belong to glucose metabolism and adhesion and/or migration pathways. Mutant cells exhibit a ∼50% decrease in ATP levels and a 30% decrease in levels of reactive oxygen species, indicating energy metabolism dysfunction. In summary, our data indicate a novel role for p53 in enabling the metabolic fitness and self-renewal of nephron progenitors. © 2015. Published by The Company of Biologists Ltd.
    Development 04/2015; 142(7):1228-41. DOI:10.1242/dev.111617 · 6.46 Impact Factor
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