Node retraction during patterning of the urinary collecting duct system.
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ABSTRACT:
SUBMITTER: Lindstrom NO
PROVIDER: S-EPMC4299504 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundGlandular organs require the development of a correctly patterned epithelial tree. These arise by iterative branching: early branches have a stereotyped anatomy, while subsequent branching is more flexible, branches spacing out to avoid entanglement. Previous studies have suggested different genetic programs are responsible for these two classes of branches.ResultsHere, working with the urinary collecting duct tree of mouse kidneys, we show that the transition from the initial, stereotyped, wide branching to narrower later branching is independent from previous branching events but depends instead on the proximity of other branch tips. A simple computer model suggests that a repelling molecule secreted by branches can in principle generate a well-spaced tree that switches automatically from wide initial branch angles to narrower subsequent ones, and that co-cultured trees would distort their normal shapes rather than colliding. We confirm this collision-avoidance experimentally using organ cultures, and identify BMP7 as the repelling molecule.ConclusionsWe propose that self-avoidance, an intrinsically error-correcting mechanism, may be an important patterning mechanism in collecting duct branching, operating along with already-known mesenchyme-derived paracrine factors.
| S-EPMC4448276 | biostudies-literature
Project description:Gene expression analysis of mouse kidney after conditional inactivation of transcription factor Tfcp2l1
2017-06-19 | GSE85325 | GEO
Project description:The collecting duct of the mammalian kidney is important for the regulation of extracellular volume, osmolarity, and pH. There are two major structurally and functionally distinct cell types: principal cells and intercalated cells. The former regulates Na+ and water homeostasis, while the latter participates in acid-base homeostasis. In vivo lineage tracing using Cre recombinase or its derivatives such as CreGFP and CreERT2 is a powerful new technique to identify stem/progenitor cells in their native environment and to decipher the origins of the tissue that they give rise to. Recent studies using this technique in mice have revealed multiple renal progenitor cell populations that differentiate into various nephron segments and collecting duct. In particular, emerging evidence suggests that like principal cells, most of intercalated cells originate from the progenitor cells expressing water channel Aquaporin 2. Mutations or malfunctions of the channels, pumps, and transporters expressed in the collecting duct system cause various human diseases. For example, gain-of-function mutations in ENaC cause Liddle's syndrome, while loss-of-function mutations in ENaC lead to Pseudohypoaldosteronism type 1. Mutations in either AE1 or V-ATPase B1 result in distal renal tubular acidosis. Patients with disrupted AQP2 or AVPR2 develop nephrogenic diabetes insipidus. A better understanding of the function and development of the collecting duct system may facilitate the discovery of new therapeutic strategies for treating kidney disease.
| S-EPMC6907014 | biostudies-literature
Project description:BackgroundAntagonists of the V1a vasopressin receptor (V1aR) are emerging as a strategy for slowing progression of CKD. Physiologically, V1aR signaling has been linked with acid-base homeostasis, but more detailed information is needed about renal V1aR distribution and function.MethodsWe used a new anti-V1aR antibody and high-resolution microscopy to investigate Va1R distribution in rodent and human kidneys. To investigate whether V1aR activation promotes urinary H+ secretion, we used a V1aR agonist or antagonist to evaluate V1aR function in vasopressin-deficient Brattleboro rats, bladder-catheterized mice, isolated collecting ducts, and cultured inner medullary collecting duct (IMCD) cells.ResultsLocalization of V1aR in rodent and human kidneys produced a basolateral signal in type A intercalated cells (A-ICs) and a perinuclear to subapical signal in type B intercalated cells of connecting tubules and collecting ducts. Treating vasopressin-deficient Brattleboro rats with a V1aR agonist decreased urinary pH and tripled net acid excretion; we observed a similar response in C57BL/6J mice. In contrast, V1aR antagonist did not affect urinary pH in normal or acid-loaded mice. In ex vivo settings, basolateral treatment of isolated perfused medullary collecting ducts with the V1aR agonist or vasopressin increased intracellular calcium levels in ICs and decreased luminal pH, suggesting V1aR-dependent calcium release and stimulation of proton-secreting proteins. Basolateral treatment of IMCD cells with the V1aR agonist increased apical abundance of vacuolar H+-ATPase in A-ICs.ConclusionsOur results show that activation of V1aR contributes to urinary acidification via H+ secretion by A-ICs, which may have clinical implications for pharmacologic targeting of V1aR.
| S-EPMC6551786 | biostudies-literature
Project description:It is well-established that the kidney collecting duct (CD) plays a central role in regulation of systemic water homeostasis. Aquaporin 2 (AQP2)-dependent water reabsorption in the CD critically depends on the arginine vasopressin (AVP) antidiuretic input and the presence of a favorable osmotic gradient at the apical plasma membrane with tubular lumen being hypotonic compared to the cytosol. This osmotic difference creates a mechanical force leading to an increase in [Ca2+]i in CD cells. The significance of the osmosensitive [Ca2+]i signaling for renal water transport and urinary concentration remain unknown. To examine molecular mechanism and physiological relevance of osmosensitivity in the CD, we implemented simultaneous direct measurements of [Ca2+]i dynamics and the rate of cell swelling as a readout of the AQP2-dependent water reabsorption in freshly isolated split-opened CDs of wild type and genetically manipulated animals and combined this with immunofluorescent detection of AVP-induced AQP2 trafficking and assessment of systemic water balance. We identified the critical role of the Ca2+-permeable TRPC3 channel in osmosensitivity and water permeability in the CD. We further demonstrated that TRPC3 -/- mice exhibit impaired urinary concentration, larger urinary volume and a greater weight loss in response to water deprivation despite increased AVP levels and AQP2 abundance. TRPC3 deletion interfered with AQP2 translocation to the plasma membrane in response to water deprivation. In summary, we provide compelling multicomponent evidence in support of a critical contribution of TRPC3 in the CD for osmosensitivity and renal water handling.
| S-EPMC6919627 | biostudies-literature
Project description:Urinary exosomes and microvesicles (EMV) are promising biomarkers for renal diseases. Although the density of EMV is very low in urine, large quantity of urine can be easily obtained. In order to analyze urinary EMV mRNA, a unique filter device to adsorb urinary EMV from 10 mL urine was developed, which is far more convenient than the standard ultracentrifugation protocol. The filter part of the device is detachable and aligned to a 96-well microplate format, therefore multiple samples can be processed simultaneously in a high throughput manner following the isolation step. For EMV mRNA quantification, the EMV on the filter is lysed directly by adding lysis buffer and transferred to an oligo(dT)-immobilized microplate for mRNA isolation followed by cDNA synthesis and real-time PCR. Under the optimized assay condition, our method provided comparable or even superior results to the standard ultracentrifugation method in terms of mRNA assay sensitivity, linearity, intra-assay reproducibility, and ease of use. The assay system was applied to quantification of kidney-specific mRNAs such as NPHN and PDCN (glomerular filtration), SLC12A1 (tubular absorption), UMOD and ALB (tubular secretion), and AQP2 (collecting duct water absorption). 12-hour urine samples were collected from four healthy subjects for two weeks, and day-to-day and individual-to-individual variations were investigated. Kidney-specific genes as well as control genes (GAPDH, ACTB, etc.) were successfully detected and confirmed their stable expressions through the two-week study period. In conclusion, this method is readily available to clinical studies of kidney diseases.
| S-EPMC4183527 | biostudies-literature
Project description:Hepatocyte nuclear factor-1β (HNF-1β) is an essential transcription factor that regulates tissue-specific gene expression in the kidney, liver, pancreas and genitourinary tract. In humans, mutations of HNF-1β cause renal cysts and diabetes (RCAD) and congenital anomalies of the kidney and urinary tract (CAKUT). Inactivation of Hnf-1β in tubular epithelial cells throughout the nephron leads to early-onset cyst formation and postnatal lethality. Here, we used Pkhd1/Cre mice to delete Hnf-1β specifically in renal collecting ducts (CD). Hnf-1β mutant mice survived long-term and developed slowly progressive cystic kidney disease, renal fibrosis, and hydronephrosis. Compared with wild-type littermates, Hnf-1β mutant mice had higher urine volume, lower urine osmolality, and higher water intake. Differences were seen at baseline, following 24h water restriction, and following administration of dDAVP. Circulating ADH levels were similar in wild type and mutant mice. Polyuria, polydipsia, and decreased urine osmolality were present prior to the onset of cyst formation and hydronephrosis. These findings indicated that Hnf-1β mutant mice have a primary defect in urinary concentrating ability. Studies using in vitro hypertonicity response experiments identified NR1H4 (FXR), a transcription factor previously shown to regulate water homeostasis, as a novel HNF-1β target. mIMCD3 cells exposed to hypertonic medium robustly upregulated Fxr mRNA levels; this upregulation was lost in Hnf-1β mutant cells. HNF-1β was bound to the Fxr promoter region in vivo, and Fxr mRNA was significantly downregulated in mutant mice. FXR protein localized to CD in wild-type mice and was almost undetectable in the cyst epithelium of mutant mice. These findings highlight a new and critical role for HNF-1β in urinary concentration and hypertonicity by regulating the transcription of FXR in the CD.
2017-06-13 | GSE85976 | GEO
Project description:To investigate the role of inner medullary collecting duct (IMCD) urea transporters in the renal concentrating mechanism, we deleted 3 kb of the UT-A urea transporter gene containing a single 140-bp exon (exon 10). Deletion of this segment selectively disrupted expression of the two known IMCD isoforms of UT-A, namely UT-A1 and UT-A3, producing UT-A1/3(-/-) mice. In isolated perfused IMCDs from UT-A1/3(-/-) mice, there was a complete absence of phloretin-sensitive or vasopressin-stimulated urea transport. On a normal protein intake (20% protein diet), UT-A1/3(-/-) mice had significantly greater fluid consumption and urine flow and a reduced maximal urinary osmolality relative to wild-type controls. These differences in urinary concentrating capacity were nearly eliminated when urea excretion was decreased by dietary protein restriction (4% by weight), consistent with the 1958 Berliner hypothesis stating that the chief role of IMCD urea transport in the concentrating mechanism is the prevention of urea-induced osmotic diuresis. Analysis of inner medullary tissue after water restriction revealed marked depletion of urea in UT-A1/3(-/-) mice, confirming the concept that phloretin-sensitive IMCD urea transporters play a central role in medullary urea accumulation. However, there were no significant differences in mean inner medullary Na(+) or Cl(-) concentrations between UT-A1/3(-/-) mice and wild-type controls, indicating that the processes that concentrate NaCl were intact. Thus, these results do not corroborate the predictions of passive medullary concentrating models stating that NaCl accumulation in the inner medulla depends on rapid vasopressin-regulated urea transport across the IMCD epithelium.
| S-EPMC409942 | biostudies-literature
Project description:BACKGROUND:Retinoic acid is the bioactive derivative of vitamin A, which plays an indispensible role in kidney development by activating retinoic acid receptors. Although the location, concentration and roles of endogenous retinoic acid in post-natal kidneys are poorly defined, there is accumulating evidence linking post-natal vitamin A deficiency to impaired renal concentrating and acidifying capacity associated with increased susceptibility to urolithiasis, renal inflammation and scarring. The aim of this study is to examine the presence and the detailed localization of endogenous retinoic acid activity in neonatal, young and adult mouse kidneys, to establish a fundamental ground for further research into potential target genes, as well as physiological and pathophysiological roles of endogenous retinoic acid in the post-natal kidneys. METHODOLOGY/PRINCIPAL FINDINGS:RARE-hsp68-lacZ transgenic mice were employed as a reporter for endogenous retinoic acid activity that was determined by X-gal assay and immunostaining of the reporter gene product, ?-galactosidase. Double immunostaining was performed for ?-galactosidase and markers of kidney tubules to localize retinoic acid activity. Distinct pattern of retinoic acid activity was observed in kidneys, which is higher in neonatal and 1- to 3-week-old mice than that in 5- and 8-week-old mice. The activity was present specifically in the principal cells and the intercalated cells of the collecting duct system in all age groups, but was absent from the glomeruli, proximal tubules, thin limbs of Henle's loop and distal tubules. CONCLUSIONS/SIGNIFICANCE:Endogenous retinoic acid activity exists in principal cells and intercalated cells of the mouse collecting duct system after birth and persists into adulthood. This observation provides novel insights into potential roles for endogenous retinoic acid beyond nephrogenesis and warrants further studies to investigate target genes and functions of endogenous retinoic acid in the kidney after birth, particularly in the collecting duct system.
| S-EPMC3033902 | biostudies-literature
Project description:An elaborate system of ducts collects urine from all nephrons, and this structure is known as the urinary collecting system (UCS). This study focused on how the UCS is formed during human embryogenesis. Fifty human embryos between the Carnegie stage (CS) 14 and CS23 were selected from the Kyoto Collection at the Congenital Anomaly Research Center of Kyoto University, Japan. Metanephroses, including the UCS, were segmented on serial digital virtual histological sections. Three-dimensional images were computationally reconstructed for morphological and quantitative analyses. A CS timeline was plotted. It consisted of the 3-D structural morphogenesis of UCS and quantification of the total amount of end-branching, average and maximum numbers of generations, deviation in the metanephros, differentiation of the urothelial epithelium in the renal pelvis, and timing of the rapid expansion of the renal pelvis. The first UCS branching generation occurred by CS16. The average branching generation reached a maximum of 8.74 ± 1.60 and was already the twelfth in CS23. The total end-branching number squared between the start and the end of the embryonic period. UCS would reach the fifteenth branching generation soon after CS23. The number of nephrons per UCS end-branch was low (0.21 ± 0.14 at CS19, 1.34 ± 0.49 at CS23), indicating that the bifid branching occurred rapidly and that the formation of nephrons followed after. The renal pelvis expanded mainly in CS23, which was earlier than that reported in a previous study. The number of nephrons connected to the UCS in the expanded group (246.0 ± 13.2) was significantly larger than that of the pre-expanded group (130.8 ± 80.1) (P < 0.05). The urothelial epithelium differentiated from the zeroth to the third generations at CS23. Differentiation may have continued up until the tenth generation to allow for renal pelvis expansion. The branching speed was not uniform. There were significantly more branching generations in the polar- than in the interpolar regions (P < 0.05). Branching speed reflects the growth orientation required to form the metanephros. Further study will be necessary to understand the renal pelvis expansion mechanism in CS23. Our CS-based timeline enabled us to map UCS formation and predict functional renal capacity after differentiation and growth.
| S-EPMC6128595 | biostudies-literature