scholarly journals Expression of claudin-8 is induced by aldosterone in renal collecting duct principal cells

2021 ◽  
Author(s):  
Ali Sassi ◽  
Yubao Wang ◽  
Alexandra Chassot ◽  
Isabelle Roth ◽  
Suresh Ramakrishnan ◽  
...  
Keyword(s):  
Tight Junction ◽  
Protein Kinases ◽  
Mineralocorticoid Receptor ◽  
Collecting Duct ◽  
Fine Tuning ◽  
Mrna Levels ◽  
Aldosterone Secretion ◽  
Mineralocorticoid Receptor Antagonist ◽  
Salt Diet ◽  
Principal Cells

Fine tuning of Na+reabsorption takes place along the aldosterone-sensitive distal nephron (ASDN) which includes the collecting duct (CD) where it is mainly regulated by aldosterone. In the CD,Na+ reabsorption is mediated by the epithelial sodium channel (ENaC) and the sodium pump (Na,K-ATPase). Paracellular ion permeability is mainly dependent on tight junction permeability. Claudin-8 is one of the main tight-junction proteins expressed along the ASDN. We have previously shown a coupling between trancellular Na+ reabsorption and paracellular Na+barrier. We hypothesize that aldosterone controls the expression levels of both transcellular Na+transporters and paracellular claudin-8 in a coordinated manner. Here, we show that aldosterone increased mRNA and protein levels as well as lateral membrane localization of claudin-8 in cultured CD principal cells. The increase in claudin-8 mRNA levels in response to aldosterone was prevented by preincubation with 17-hydroxy progesterone, a mineralocorticoid receptor antagonist, and by inhibition of transcription with actinomycin D. We also show that low salt diet which stimulated aldosterone secretion was associated with increased claudin-8 abundance in the mouse kidney. Reciprocally, mice subjected to high salt diet which inhibits aldosterone secretion or treated with spironolactone, a mineralocorticoid receptor (MR)antagonist, displayed decreased claudin-8 expression. Inhibition of glycogen synthase kinase-3 (GSK3), Lyn and Abl signaling pathways prevented the effect of aldosterone on claudin-8 mRNA and protein abundance, suggesting that signaling protein kinases plays a permissive role on the transcriptional activity of the mineralocorticoid receptor.This study shows that signaling via multiple protein kinases working in concert mediates the aldosterone-induced claudin-8 expression in collecting duct.

scholarly journals Aldosterone up-regulates voltage-gated potassium currents and NKCC1 protein membrane fractions

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Parveen Bazard ◽  
Bo Ding ◽  
Harish K. Chittam ◽  
Xiaoxia Zhu ◽  
Thomas A. Parks ◽  
...  
Keyword(s):  
Protein Expression ◽  
Mineralocorticoid Receptor ◽  
Therapeutic Potential ◽  
Potassium Currents ◽  
Chloride Ions ◽  
Mrna Levels ◽  
Mineralocorticoid Receptor Antagonist ◽  
Voltage Gated ◽  
Age Related

Abstract Na+–K+–2Cl− Cotransporter (NKCC1) is a protein that aids in the active transport of sodium, potassium, and chloride ions across cell membranes. It has been shown that long-term systemic treatment with aldosterone (ALD) can enhance NKCC1 protein expression and activity in the aging cochlea resulting in improved hearing. In the present work, we used a cell line with confirmed NKCC1 expression to demonstrate that in vitro application of ALD increased outward voltage-gated potassium currents significantly, and simultaneously upregulated whole lysate and membrane portion NKCC1 protein expression. These ALD-induced changes were blocked by applying the mineralocorticoid receptor antagonist eplerenone. However, application of the NKCC1 inhibitor bumetanide or the potassium channel antagonist Tetraethyl ammonium had no effect. In addition, NKKC1 mRNA levels remained stable, indicating that ALD modulates NKCC1 protein expression via the activation of mineralocorticoid receptors and post-transcriptional modifications. Further, in vitro electrophysiology experiments, with ALD in the presence of NKCC1, K+ channel and mineralocorticoid receptor inhibitors, revealed interactions between NKCC1 and outward K+ channels, mediated by a mineralocorticoid receptor-ALD complex. These results provide evidence of the therapeutic potential of ALD for the prevention/treatment of inner ear disorders such as age-related hearing loss.

scholarly journals Molecular mechanisms of angiotensin II stimulation on aquaporin-2 expression and trafficking

2011 ◽  
Vol 300 (5) ◽  
pp. F1255-F1261 ◽  
Author(s):  
Chunling Li ◽  
Weidong Wang ◽  
Christopher J. Rivard ◽  
Miguel A. Lanaspa ◽  
Sandra Summer ◽  
...  
Keyword(s):  
Protein Expression ◽  
Signaling Pathways ◽  
Collecting Duct ◽  
Mrna Levels ◽  
Camp Accumulation ◽  
Ang Ii ◽  
Principal Cells ◽  
Aquaporin 2 ◽  
Renal Collecting Duct ◽  
Sodium Regulation

ANG II plays a major role in renal water and sodium regulation. In the immortalized mouse renal collecting duct principal cells (mpkCCDcl4) cell line, we treated cells with ANG II and examined aquaporin-2 (AQP2) protein expression, trafficking, and mRNA levels, by immunoblotting, immunofluorescence, and RT-PCR. After 24-h incubation, ANG II-induced AQP2 protein expression was observed at the concentration of 10−10 M and increased in a dose-dependent manner. ANG II (10−7 M) increased AQP2 protein expression and mRNA levels at 0.5, 1, 2, 6, and 24 h. Immunofluorescence studies showed that ANG II increased the apical membrane targeting of AQP2 from 30 min to 6 h. Next, the signaling pathways underlying the ANG II-induced AQP2 expression were investigated. The PKC inhibitor Ro 31–8220 (5 × 10−6 M) and the PKA inhibitor H89 (10−5 M) blocked ANG II-induced AQP2 expression, respectively. Calmodulin inhibitor W-7 markedly reduced ANG II- and/or dDAVP-stimulated AQP2 expression. ANG II (10−9 M) and/or dDAVP (10−10 M) stimulated AQP2 protein levels and cAMP accumulation, which was completely blocked by pretreatment with the vasopressin V2 receptor (V2R) antagonist SR121463B (10−8 M). Pretreatment with the angiotensin AT1 receptor (AT1R) antagonist losartan (3 × 10−6 M) blocked ANG II (10−9 M)-stimulated AQP2 protein expression and cAMP accumulation, and partially blocked dDAVP (10−10 M)- and dDAVP+ANG II-induced AQP2 protein expression and cAMP accumulation. In conclusion, ANG II regulates AQP2 protein, trafficking, and gene expression in renal collecting duct principal cells. ANG II-induced AQP2 expression involves cAMP, PKC, PKA, and calmodulin signaling pathways via V2 and AT1 receptors.

The α1G-subunit of a voltage-dependent Ca2+ channel is localized in rat distal nephron and collecting duct

2000 ◽  
Vol 279 (6) ◽  
pp. F997-F1005 ◽  
Author(s):  
Ditte Andreasen ◽  
Boye L. Jensen ◽  
Pernille B. Hansen ◽  
Tae-Hwan Kwon ◽  
Søren Nielsen ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Rnase Protection Assay ◽  
Confocal Laser ◽  
Pcr Analysis ◽  
Apical Plasma Membrane ◽  
Mrna Levels ◽  
Double Labeling ◽  
Rnase Protection ◽  
Principal Cells ◽  
Voltage Dependent

The molecular type and localization of calcium channels along the nephron are not well understood. In the present study, we assessed the distribution of the recently identified α1G-subunit encoding a voltage-dependent calcium channel with T-type characteristics. Using a RNase protection assay, α1G-mRNA levels in kidney regions were determined as inner medulla ≫ outer medulla ≅ cortex. RT-PCR analysis of microdissected rat nephron segments revealed α1G expression in the distal convoluted tubule (DCT), in the connecting tubule and cortical collecting duct (CT+CCD), and inner medullary collecting duct (IMCD). α1GmRNA was expressed in the IMCD cell line mIMCD-3. Single- and double-labeling immunohistochemistry and confocal laser microscopy on semithin paraffin sections of rat kidneys by using an anti-α1G antibody demonstrated a distinct labeling at the apical plasma membrane domains of DCT cells, CT principal cells, and IMCD principal cells.

scholarly journals Two Mineralocorticoid Receptor–Mediated Mechanisms of Pendrin Activation in Distal Nephrons

2020 ◽  
Vol 31 (4) ◽  
pp. 748-764 ◽  
Author(s):  
Nobuhiro Ayuzawa ◽  
Mitsuhiro Nishimoto ◽  
Kohei Ueda ◽  
Daigoro Hirohama ◽  
Wakako Kawarazaki ◽  
...  
Keyword(s):  
Sodium Chloride ◽  
Angiotensin Ii ◽  
Knockout Mice ◽  
Mineralocorticoid Receptor ◽  
Receptor Activation ◽  
Intercalated Cells ◽  
Renin Angiotensin Aldosterone System ◽  
Salt Diet ◽  
Principal Cells ◽  
Salt Sensitive Hypertension

BackgroundRegulation of sodium chloride transport in the aldosterone-sensitive distal nephron is essential for fluid homeostasis and BP control. The chloride-bicarbonate exchanger pendrin in β-intercalated cells, along with sodium chloride cotransporter (NCC) in distal convoluted tubules, complementarily regulate sodium chloride handling, which is controlled by the renin-angiotensin-aldosterone system.MethodsUsing mice with mineralocorticoid receptor deletion in intercalated cells, we examined the mechanism and roles of pendrin upregulation via mineralocorticoid receptor in two different models of renin-angiotensin-aldosterone system activation. We also used aldosterone-treated NCC knockout mice to examine the role of pendrin regulation in salt-sensitive hypertension.ResultsDeletion of mineralocorticoid receptor in intercalated cells suppressed the increase in renal pendrin expression induced by either exogenous angiotensin II infusion or endogenous angiotensin II upregulation via salt restriction. When fed a low-salt diet, intercalated cell–specific mineralocorticoid receptor knockout mice with suppression of pendrin upregulation showed BP reduction that was attenuated by compensatory activation of NCC. In contrast, upregulation of pendrin induced by aldosterone excess combined with a high-salt diet was scarcely affected by deletion of mineralocorticoid receptor in intercalated cells, but depended instead on hypokalemic alkalosis through the activated mineralocorticoid receptor–epithelial sodium channel cascade in principal cells. In aldosterone-treated NCC knockout mice showing upregulation of pendrin, potassium supplementation corrected alkalosis and inhibited the pendrin upregulation, thereby lowering BP.ConclusionsIn conjunction with NCC, the two pathways of pendrin upregulation, induced by angiotensin II through mineralocorticoid receptor activation in intercalated cells and by alkalosis through mineralocorticoid receptor activation in principal cells, play important roles in fluid homeostasis during salt depletion and salt-sensitive hypertension mediated by aldosterone excess.

Insulin potentiates AVP-induced AQP2 expression in cultured renal collecting duct principal cells

2005 ◽  
Vol 288 (2) ◽  
pp. F334-F344 ◽  
Author(s):  
Mauro Bustamante ◽  
Udo Hasler ◽  
Olga Kotova ◽  
Alexander V. Chibalin ◽  
David Mordasini ◽  
...  
Keyword(s):  
Protein Expression ◽  
Collecting Duct ◽  
Basal Medium ◽  
Mrna Levels ◽  
Kinase Activation ◽  
Principal Cells ◽  
Inhibitor Analysis ◽  
Mrna And Protein Expression ◽  
Renal Collecting Duct ◽  
Aqp2 Gene

In the renal collecting duct (CD), water reabsorption depends on the presence of aquaporin-2 (AQP2) in the apical membrane of principal cells. AQP2 expression and subcellular repartition are under the control of AVP. Some pieces of experimental evidence indicate that additional hormonal factors, including insulin, may also control AQP2 expression and thereby CD water permeability. We have previously shown that AVP induces endogenous AQP2 expression in cultured mouse mpkCCDcl4 CD principal cells ( 23 ). In the present study, we investigated the effect of insulin on AQP2 expression in mpkCCDcl4 cells. Addition of insulin to the basal medium of cells grown on filters slightly increased AQP2 mRNA and protein expression, whereas insulin potentiated the effect of AVP. The potentiation of AVP-induced AQP2 expression by insulin was abolished by actinomycin D, a transcriptional inhibitor. Analysis of AQP2 protein expression under conditions of AVP washout and/or in the presence of chloroquine, a lysosomal degradation inhibitor, revealed that insulin did not significantly alter AQP2 protein degradation. Inhibition of ERK, p38 kinase, and phosphatidylinositol 3′-kinase (PI 3-kinase) activities prevented the insulin-induced stimulation of AQP2 expression, whereas inhibition of PKC has no effect. Taken together, our results indicate that insulin increased AQP2 protein expression mostly through increased AQP2 mRNA levels in cultured mpkCCDcl4 cells. This effect most likely relies on increased AQP2 gene transcription in response to MAPK and PI 3-kinase activation.

scholarly journals Mineralocorticoid receptor antagonizes Dot1a-Af9 complex to increase αENaC transcription

2013 ◽  
Vol 305 (10) ◽  
pp. F1436-F1444 ◽  
Author(s):  
Xi Zhang ◽  
Qiaoling Zhou ◽  
Lihe Chen ◽  
Stefan Berger ◽  
Hongyu Wu ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Mineralocorticoid Receptor ◽  
Collecting Duct ◽  
Mrna Levels ◽  
Salt Wasting ◽  
Molecular Defects ◽  
Binding Partner ◽  
Medullary Collecting Duct ◽  
Pseudohypoaldosteronism Type

Aldosterone is a major regulator of Na+ absorption and acts by activating the mineralocorticoid receptor (MR) to stimulate the epithelial Na+ channel (ENaC). MR −/− mice exhibited pseudohypoaldosteronism type 1 (hyponatremia, hyperkalemia, salt wasting, and high levels of aldosterone) and died around postnatal day 10. However, if and how MR regulates ENaC transcription remain incompletely understood. Our earlier work demonstrated that aldosterone activates αENaC transcription by reducing expression of Dot1a and Af9 and by impairing Dot1a-Af9 interaction. Most recently, we reported identification of a major Af9 binding site in the αENaC promoter and upregulation of αENaC mRNA expression in mouse kidneys lacking Dot1a. Despite these findings, the putative antagonism between the MR/aldosterone and Dot1a-Af9 complexes has never been addressed. The molecular defects leading to PHA-1 in MR −/− mice remain elusive. Here, we report that MR competes with Dot1a to bind Af9. MR/aldosterone and Dot1a-Af9 complexes mutually counterbalance ENaC mRNA expression in inner medullary collecting duct 3 (IMCD3) cells. Real-time RT-quantitative PCR revealed that 5-day-old MR −/− vs. MR +/+ mice had significantly lower αENaC mRNA levels. This change was associated with an increased Af9 binding and H3 K79 hypermethylation in the αENaC promoter. Therefore, this study identified MR as a novel binding partner and regulator of Af9 and a novel mechanism coupling MR-mediated activation with relief of Dot1a-Af9-mediated repression via MR-Af9 interaction. Impaired ENaC expression due to failure to inhibit Dot1a-Af9 may play an important role in the early stages of PHA-1 (before postnatal day 8) in MR −/− mice.

scholarly journals Rho-kinase inhibitor hydroxyfasudil protects against HIV-1 Tat-induced dysfunction of tight junction and neprilysin/Aβ transfer receptor expression in mouse brain microvessels

2021 ◽  
Vol 476 (5) ◽  
pp. 2159-2170
Author(s):  
Qiangtang Chen ◽  
Yu Wu ◽  
Yachun Yu ◽  
Junxiang Wei ◽  
Wen Huang
Keyword(s):  
Tight Junction ◽  
Signaling Pathway ◽  
Mouse Brain ◽  
Kinase Inhibitor ◽  
Rho Kinase ◽  
Receptor Expression ◽  
Mrna Levels ◽  
Brain Microvessels ◽  
Rho Kinase Inhibitor ◽  
Hiv 1

AbstractHIV-1 transactivator protein (Tat) induces tight junction (TJ) dysfunction and amyloid-beta (Aβ) clearance dysfunction, contributing to the development and progression of HIV-1-associated neurocognitive disorder (HAND). The Rho/ROCK signaling pathway has protective effects on neurodegenerative disease. However, the underlying mechanisms of whether Rho/ROCK protects against HIV-1 Tat-caused dysfunction of TJ and neprilysin (NEP)/Aβ transfer receptor expression have not been elucidated. C57BL/6 mice were administered sterile saline (i.p., 100 μL) or Rho-kinase inhibitor hydroxyfasudil (HF) (i.p., 10 mg/kg) or HIV-1 Tat (i.v., 100 μg/kg) or HF 30 min before being exposed to HIV-1 Tat once a day for seven consecutive days. Evans Blue (EB) leakage was detected via spectrophotometer and brain slides in mouse brains. The protein and mRNA levels of zonula occludens-1 (ZO-1), occludin, NEP, receptor for advanced glycation end products (RAGE), and low-density lipoprotein receptor-related protein 1 (LRP1) in mouse brain microvessels were, respectively, analyzed by Western blotting and quantitative real-time polymerase chain reaction (qRT-PCR) analyses. Exposure of the mice to HIV-1 Tat increased the amount of EB leakage, EB fluorescence intensity, blood–brain barrier (BBB) permeability, as well as the RAGE protein and mRNA levels, and decreased the protein and mRNA levels of ZO-1, occludin, NEP, and LRP1 in mouse brain microvessels. However, these effects were weakened by Rho-kinase inhibitor HF. Taken together, these results provide information that the Rho/ROCK signaling pathway is involved in HIV-1 Tat-induced dysfunction of TJ and NEP/Aβ transfer receptor expression in the C57BL/6 mouse brain. These findings shed some light on potentiality of inhibiting Rho/Rock signaling pathway in handling HAND.

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