scholarly journals Isolation and Culture of Cells from the Nephrogenic Zone of the Embryonic Mouse Kidney

10.3791/2555 ◽  
2011 ◽  
Author(s):  
Aaron C. Brown ◽  
Ulrika Blank ◽  
Derek C. Adams ◽  
Michele J. Karolak ◽  
Jennifer L. Fetting ◽  
...  
Keyword(s):  
Mouse Kidney ◽  
Embryonic Mouse ◽  
Nephrogenic Zone

scholarly journals Colocalization and Redistribution of Dishevelled and Actin during WNT-Induced Mesenchymal Morphogenesis

2000 ◽  
Vol 149 (7) ◽  
pp. 1433-1442 ◽  
Author(s):  
Monica A. Torres ◽  
W. James Nelson
Keyword(s):  
Gene Expression ◽  
Wnt Signaling ◽  
Signaling Pathway ◽  
Wnt Signaling Pathway ◽  
Cell Spreading ◽  
Mesenchymal Cell ◽  
Mouse Kidney ◽  
Cell Morphogenesis ◽  
Embryonic Mouse ◽  
Stimulation Of

Activation of the Wnt signaling pathway is important for induction of gene expression and cell morphogenesis throughout embryonic development. We examined the subcellular localization of dishevelled, the immediate downstream component from the Wnt receptor, in the embryonic mouse kidney. Using immunofluorescence staining, confocal microscopy, and coimmunoprecipitation experiments, we show that dishevelled associates with actin fibers and focal adhesion plaques in metanephric mesenchymal cells. Stimulation of Wnt signaling leads to profound changes in metanephric mesenchymal cell morphology, including disruption of the actin cytoskeleton, increased cell spreading, and increased karyokinesis. Upon activation of Wnt signaling, dishevelled also accumulates in and around the nucleus. Casein kinase Iε colocalizes with dishevelled along actin fibers and in the perinuclear region, whereas axin and GSK-3 are only present around the nucleus. These data indicate a branched Wnt signaling pathway comprising a canonical signal that targets the nucleus and gene expression, and another signal that targets the cytoskeleton and regulates cell morphogenesis.

scholarly journals Three-Dimensional Imaging of Embryonic Mouse Kidney by Two-Photon Microscopy

2001 ◽  
Vol 158 (1) ◽  
pp. 49-55 ◽  
Author(s):  
Carrie L. Phillips ◽  
Lois J. Arend ◽  
Adele J. Filson ◽  
Doug J. Kojetin ◽  
Jeffrey L. Clendenon ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Mouse Kidney ◽  
Three Dimensional Imaging ◽  
Embryonic Mouse ◽  
Two Photon Microscopy ◽  
Two Photon

Ontogeny of calbindin-D28K and calbindin-D9K in the mouse kidney, duodenum, cerebellum and placenta

Development ◽  
1992 ◽  
Vol 116 (2) ◽  
pp. 491-496 ◽  
Author(s):  
D.R. Shamley ◽  
L.A. Opperman ◽  
R. Buffenstein ◽  
F.P. Ross
Keyword(s):  
Vitamin D ◽  
Calcium Binding ◽  
Giant Cells ◽  
Mouse Kidney ◽  
Embryonic Mouse ◽  
Immunohistochemical Assay ◽  
Calbindin D9k ◽  
Early Appearance ◽  
Mesonephric Duct ◽  
Calbindin D28k

The appearance of the calcium-binding proteins (CaBP-D28K and CaBP-D9K) in embryonic mice tissues was determined using a sensitive immunohistochemical assay. CaBP-D28K first appears in myenteric nerve plexuses of the duodenum on day E15, in duodenal villus cells on day E16, in Purkinje cells of the cerebellum on day E19, in cells of the mesonephric duct on day E11 and in the metanephric duct on day E12. CaBP-D9K first appears in enterocytes of the duodenum on day E18, in trophoblastic giant cells (TGC) of the placenta on day E10, and in the metanephric duct on day E15. A differential time of appearance and colocalization of the two CaBPs is demonstrated in the embryonic mouse kidney, suggesting either that vitamin D does not control both CaBPs in the foetus or that the vitamin D control is unequal. The early appearance and location of CaBP-D9K in TGCs may suggest that these cells play an important role in transplacental transfer of calcium.

scholarly journals Integrin-Linked Kinase Mediates Bone Morphogenetic Protein 7-Dependent Renal Epithelial Cell Morphogenesis

2005 ◽  
Vol 25 (9) ◽  
pp. 3648-3657 ◽  
Author(s):  
Chungyee Leung-Hagesteijn ◽  
Ming Chang Hu ◽  
Ahalya S. Mahendra ◽  
Sunny Hartwig ◽  
Henry J. Klamut ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Branching Morphogenesis ◽  
Mouse Kidney ◽  
Dominant Negative ◽  
Cell Morphogenesis ◽  
Renal Epithelial Cell ◽  
Embryonic Mouse ◽  
Morphogenetic Protein ◽  
Culture Model ◽  
Integrin Linked Kinase

ABSTRACT Bone morphogenetic protein 7 (BMP7) stimulates renal branching morphogenesis via p38 mitogen-activated protein kinase (p38MAPK) and activating transcription factor 2 (ATF-2) (M. C. Hu, D. Wasserman, S. Hartwig, and N. D. Rosenblum, J. Biol. Chem. 279:12051-12059, 2004). Here, we demonstrate a novel role for integrin-linked kinase (ILK) in mediating renal epithelial cell morphogenesis in embryonic kidney explants and identify p38MAPK as a target of ILK signaling in a cell culture model of renal epithelial morphogenesis. The spatial and temporal expression of ILK in embryonic mouse kidney cells suggested a role in branching morphogenesis. Adenovirus-mediated expression of ILK stimulated and expression of a dominant negative ILK mutant inhibited ureteric bud branching in embryonic mouse kidney explants. BMP7 increased ILK kinase activity in inner medullary collecting duct 3 (IMCD-3) cells, and adenovirus-mediated expression of ILK increased IMCD-3 cell morphogenesis in a three-dimensional culture model. In contrast, treatment with a small molecule ILK inhibitor or expression of a dominant negative-acting ILK (ILKE359K) inhibited epithelial cell morphogenesis. Further, expression of ILKE359K abrogated BMP7-dependent stimulation. To investigate the role of ILK in BMP7 signaling, we showed that ILK overexpression increased basal and BMP7-induced levels of phospho-p38MAPK and phospho-ATF-2. Consistent with its inhibitory effects on IMCD-3 cell morphogenesis, expression of ILKE359K blocked BMP7-dependent increases in phospho-p38MAPK and phospho-ATF-2. Inhibition of p38MAPK activity with the specific inhibitor, SB203580, failed to inhibit BMP7-dependent stimulation of ILK activity, suggesting that ILK functions upstream of p38MAPK during BMP7 signaling. We conclude that ILK functions in a BMP7/p38MAPK/ATF-2 signaling pathway and stimulates epithelial cell morphogenesis.

scholarly journals Mouse β6 Integrin Sequence, Pattern of Expression, and Role in Kidney Development

2000 ◽  
Vol 11 (12) ◽  
pp. 2297-2305
Author(s):  
LOIS J. AREND ◽  
ANN M. SMART ◽  
JOSIE P. BRIGGS
Keyword(s):  
Mrna Expression ◽  
Reverse Transcription ◽  
Kidney Development ◽  
Mouse Kidney ◽  
Thick Ascending Limb ◽  
Integrin Expression ◽  
Coding Region ◽  
Embryonic Mouse ◽  
Nephron Segments ◽  
Reverse Transcription Pcr

Abstract. Integrins mediate cell-cell and cell-extracellular matrix interactions and play key roles in development. β6 integrin expression has been demonstrated in human fetal kidney at a higher level than in the adult, making β6 integrin a marker of interest for the study of development of the nephron. The aims of this study were to determine the cDNA sequence for the mouse β6 integrin and to characterize β6 integrin expression in the developing mouse kidney. Two embryonic mouse kidney cDNA libraries were screened, and the coding region was sequenced. The mouse β6 nucleotide coding region sequence shows 82% nucleotide identity to the human sequence. The putative amino acid sequence has 89.5% identity to human β6 integrin and contains many conserved domains. By reverse transcription-PCR, β6 integrin mRNA expression is very low at 11 d of gestation in the mouse, increases dramatically by E14 and E17 (20-fold, normalized for increases in β actin), and plateaus by 2 wk of age. β6 integrin expression is induced 15- to 20-fold after 5 d in metanephric explant culture. Reverse transcription-PCR of adult rat microdissected nephron segments demonstrates β6 integrin mRNA expression in proximal tubule, cortical thick ascending limb, distal nephron segments (inner and outer medullary collecting ducts), and macula densa—containing segments. Lectin-peroxidase and in situ colocalization studies demonstrated expression of β6 integrin mRNA in developing proximal tubules and thick ascending limb. Culture of mouse metanephric kidneys with antisense oligonucleotides to β6 integrin resulted in inhibition of ureteric bud branching and complete lack of mesenchyme condensation. These studies demonstrate a high homology between the human and mouse β6 integrin sequence, a different pattern of expression in the developing mouse kidney compared with the primate kidney, and abnormal metanephric development in culture in the absence of β6 integrin. These findings suggest an important role for β6 integrin in normal development of the mouse kidney.

scholarly journals Molecular determinants of WNT9b responsiveness in nephron progenitor cells

2018 ◽  
Author(s):  
Kyle K. Dickinson ◽  
Leah C. Hammond ◽  
Courtney M. Karner ◽  
Nicholas D. Hastie ◽  
Thomas J. Carroll ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Wnt Signaling Pathway ◽  
Mouse Kidney ◽  
Luciferase Reporter ◽  
Metanephric Mesenchyme ◽  
Ureteric Bud ◽  
Embryonic Mouse ◽  
Luciferase Reporter Plasmid ◽  
Sirna Knockdown ◽  
Nephron Progenitor

AbstractPrimed nephron progenitor cells (NPCs) appear in metanephric mesenchyme by Ell.5 and differentiate in response to the inductive WNT9b signal from the ureteric bud. However, the NPC WNT-receptor complex is unknown. We obtained M15 cells from E10.5 mesonephric mesenchyme and systematically analyzed components required for canonical WNT9b-responsiveness. When M15 cells were transfected with a (β-catenin luciferase reporter plasmid, exposure to recombinant WNT9b resulted in minimal luciferase activity. We then analyzed mRNA-expression of WNT-pathway components and identified Fzdl-6 and Lrp6 transcripts but not RSPO1. When M15 cells were treated with recombinant RSPO1 the response to transfected WNT9b was augmented 4.8-fold. Co-transfection of M15 cells with Fzd5 (but no other Fzd family member) further increased the WNT9b signal to 16.8-fold and siRNA knockdown of Fzd5 reduced the signal by 52%. Knockdown of Lrp6 resulted in 60% WNT9b signal reduction. We confirmed Fzd5, Lrp6 and RSPO1 rrtRNA expression in CITED1(+) NPCs from E15.5 embryonic mouse kidney. Thus, while many WNT signaling-pathway components are present by E10.5, optimum responsiveness of Ell.5 cap mesenchyme requires that NPCs acquire RSPO1, FZD5 and LRP6.Summary StatementResponsiveness to the inductive WMT9b signal from ureteric bud is crucial for nephrogenesis. Here we analyze the molecules needed to prime nephron progenitor cells in embryonic mouse kidney.

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