scholarly journals Mutagenesis of the Cleavage Site of Pro Renin Receptor Abrogates Angiotensin II-Induced Hypertension in Mice

Hypertension ◽  
2021 ◽  
Author(s):  
Fei Wang ◽  
Yanting Chen ◽  
Chang-Jiang Zou ◽  
Renfei Luo ◽  
Tianxin Yang
Keyword(s):  
Angiotensin Ii ◽  
Sodium Channel ◽  
Cleavage Site ◽  
Collecting Duct ◽  
Ussing Chamber ◽  
Renin Angiotensin System ◽  
Epithelial Sodium Channel ◽  
Wild Type ◽  
Developmental Abnormalities ◽  
Induced Hypertension

It is well demonstrated that activation of renal PRR ([pro]renin receptor) contributes to AngII (angiotensin II)-induced hypertension. Relatively, less is known for the function of sPRR (soluble PRR), the extracellular domain of PRR, primarily generated by S1P (site-1 protease) and furin. Moreover, the relationship between PRR/sPRR and the renin-angiotensin system (RAS) has been debated. In the present study, we used CRISPR/Cas9 strategy to generate mice with mutagenesis of the overlapping cleavage site for both proteases in PRR (termed as PRR R279V/L282V ) to examine the phenotype during AngII infusion with particular emphasis on circulating and intrarenal renin status. PRR R279V/L282V mice exhibited a reduction of sPRR level in plasma by ≈53% and in the kidney by ≈82%, were fertile, and had no gross developmental abnormalities. At basal condition, PRR R279V/L282V mice had drastically suppressed renin levels from plasma, urine, and the kidney as compared to wild-type controls. The hypertensive response of PRR R279V/L282V to AngII infusion was blunted in parallel with attenuated response of intrarenal renin and renal medullary α-epithelial sodium channel expression. By using Ussing chamber technique, primary collecting duct cells from PRR R279V/L282V mice exhibited blunted response of epithelial sodium channel activity to AngII as compared to wild-type cells. Together, these results represent strong evidence favoring sPRR as a mediator of AngII-induced hypertension and a master regulator of renin expression. Therefore, PRR should be considered as an integrative member of the RAS.

Abstract MP46: Mutagenesis Of PRR Cleavage Site In Mice Attenuates Angiotensin II-induced Hypertension And Suppresses Renin Expression

Hypertension ◽  
2021 ◽  
Vol 78 (Suppl_1) ◽  
Author(s):  
Fei Wang ◽  
Yanting Chen ◽  
Chang-jiang Zou ◽  
Renfei Luo ◽  
Tianxin Yang
Keyword(s):  
Angiotensin Ii ◽  
Cell Line ◽  
Cleavage Site ◽  
Collecting Duct ◽  
Renin Angiotensin System ◽  
Cell Types ◽  
Mouse Cell ◽  
Mrna Levels ◽  
Direct Role ◽  
Induced Hypertension

It is well demonstrated that activation of renal (pro)renin receptor (PRR) contributes to AngII-induced hypertension. Relatively, less is known about involvement of soluble PRR (sPRR), the extracellular domain of PRR, primarily generated by site-1 protease(S1P) and furin. Moreover, the relationship between PRR/sPRR and the renin-angiotensin system (RAS) has been debated. In the present study, we used CRISPR/Cas9 strategy to generate mice with mutagenesis of the overlapping cleavage site for both proteases in PRR (termed as PRR R279V/L282V ) to examine the phenotype during angiotensin II (AngII) infusion with particular emphasis on circulating and intrarenal renin status. PRR R279V/L282V mice exhibited a reduction of sPRR level in plasma by ~53% and in the kidney by ~82%, were fertile, and had no gross developmental abnormalities. At basal condition, PRR R279V/L282V mice had drastically suppressed renin levels from plasma (plasma total prorenin/renin content [ng/ml]: 9.3 ± 0.7 in PRR R279V/L282V mice vs. 12.8 ± 0.9 in WT mice, n = 5, p < 0.05), urine (urinary total prorenin/renin excretion [ng/24h]: 0.54 ± 0.11 in PRR R279V/L282V mice vs. 1.05 ± 0.15 in WT mice, n = 5, p < 0.05), and the kidney as compared to wild-type controls (WT). By telemetry, the hypertensive response of PRR R279V/L282V to AngII infusion (300 ng/min/kg) was blunted (MAP [mmHg] in Day 10: 125.9 ± 4.6 in PRR R279V/L282V mice vs. 138.5 ± 2.8 in WT mice, n = 8 for each group, p < 0.05) in parallel with attenuated response of intrarenal renin and renal medullary α-ENaC expression. We further examined the direct role of sPRR in renin regulation in As4.1 cells, a renin-expressing cell line isolated from mouse renal tumor and M-1 cells, a mouse cell line derived from the collecting duct. The exposure to sPRR-His in both cell types consistently elevated renin activity, and renin expression at both protein and mRNA levels, all of which were sensitive to inhibition by ICG-001, a β-catenin signaling inhibitor. Together, these results represent strong evidence favoring sPRR as a mediator of AngII-induced hypertension and a master regulator of renin expression at systemic and local levels. Therefore, PRR should be considered as an integrative member of the RAS.

scholarly journals Site-1 Protease-Derived Soluble (Pro)Renin Receptor Contributes to Angiotensin II–Induced Hypertension in Mice

Hypertension ◽  
2020 ◽  
Author(s):  
Ye Feng ◽  
Kexin Peng ◽  
Renfei Luo ◽  
Fei Wang ◽  
Tianxin Yang
Keyword(s):  
Blood Pressure ◽  
Angiotensin Ii ◽  
Collecting Duct ◽  
Short Circuit ◽  
Ussing Chamber ◽  
Short Circuit Current ◽  
Duct Cell ◽  
Ang Ii ◽  
Cortical Collecting Duct ◽  
Induced Hypertension

Activation of PRR ([pro]renin receptor) contributes to enhancement of intrarenal RAS and renal medullary α-ENaC and thus elevated blood pressure during Ang II (angiotensin II) infusion. The goal of the present study was to test whether such action of PRR was mediated by sPRR (soluble PRR), generated by S1P (site-1 protease), a newly identified PRR cleavage protease. F1 B6129SF1/J mice were infused for 6 days with control or Ang II at 300 ng/kg per day alone or in combination with S1P inhibitor PF-429242 (PF), and blood pressure was monitored by radiotelemetry. S1P inhibition significantly attenuated Ang II–induced hypertension accompanied with suppressed urinary and renal medullary renin levels and expression of renal medullary but not renal cortical α-ENaC expression. The effects of S1P inhibition were all reversed by supplement with histidine-tagged sPRR termed as sPRR-His. Ussing chamber technique was performed to determine amiloride-sensitive short-circuit current, an index of ENaC activity in confluent mouse cortical collecting duct cell line cells exposed for 24 hours to Ang II, Ang II + PF, or Ang II + PF + sPRR-His. Ang II–induced ENaC activity was blocked by PF, which was reversed by sPRR-His. Together, these results support that S1P-derived sPRR mediates Ang II–induced hypertension through enhancement of intrarenal renin level and activation of ENaC.

scholarly journals Plasminogen Deficiency and Amiloride Mitigate Angiotensin II–Induced Hypertension in Type 1 Diabetic Mice Suggesting Effects Through the Epithelial Sodium Channel

2020 ◽  
Vol 9 (23) ◽  
Author(s):  
Henrik Andersen ◽  
Maria Høj Hansen ◽  
Kristian B. Buhl ◽  
Mette Stæhr ◽  
Ulla G. Friis ◽  
...  
Keyword(s):  
Diabetes Mellitus ◽  
Blood Pressure ◽  
Angiotensin Ii ◽  
Sodium Channel ◽  
Arterial Blood Pressure ◽  
Epithelial Sodium Channel ◽  
Diabetic Mice ◽  
Wild Type ◽  
Arterial Blood ◽  
Prorenin Receptor

Background Diabetic nephropathy is a common diabetes mellitus complication associated with hypertension, proteinuria, and excretion of urinary plasmin that activates the epithelial sodium channel, ENaC, in vitro . Here we hypothesized that the deletion of plasminogen and amiloride treatment protect against hypertension in diabetes mellitus. Methods and Results Male plasminogen knockout (plasminogen‐deficient [Plg −/− ]) and wild‐type mice were rendered diabetic with streptozotocin. Arterial blood pressure was recorded continuously by indwelling catheters before and during 10 days of angiotensin II infusion (ANGII; 30–60 ng/kg per minute). The effect of amiloride infusion (2 mg/kg per day, 4 days) was tested in wild‐type, diabetic ANGII‐treated mice. Streptozotocin increased plasma and urine glucose concentrations and 24‐hour urine albumin and plasminogen excretion. Diabetic Plg −/− mice displayed larger baseline albuminuria and absence of urine plasminogen. Baseline mean arterial blood pressure did not differ between groups. Although ANGII elevated blood pressure in wild‐type, diabetic wild‐type, and Plg −/− control mice, ANGII did not change blood pressure in diabetic Plg −/− mice. Compared with ANGII infusion alone, wild‐type ANGII‐infused diabetic mice showed blood pressure reduction upon amiloride treatment. There was no difference in plasma renin, ANGII, aldosterone, tissue prorenin receptor, renal inflammation, and fibrosis between groups. Urine from wild‐type mice evoked larger amiloride‐sensitive current than urine from Plg −/− mice with or without diabetes mellitus. Full‐length γ‐ENaC and α‐ENaC subunit abundances were not changed in kidney homogenates, but the 70 kDa γ‐ENaC cleavage product was increased in diabetic versus nondiabetic mice. Conclusions Plasmin promotes hypertension in diabetes mellitus with albuminuria likely through the epithelial sodium channel.

Renal NOXA1/NOX1 Signaling Regulates Epithelial Sodium Channel and Sodium Retention in Angiotensin II-induced Hypertension

2021 ◽  
Author(s):  
Aleksandr Vendrov ◽  
Mark D. Stevenson ◽  
Andrey Lozhkin ◽  
Takayuki Hayami ◽  
Nathan A Holland ◽  
...  
Keyword(s):  
Angiotensin Ii ◽  
Sodium Channel ◽  
Epithelial Sodium Channel ◽  
Sodium Retention ◽  
Induced Hypertension

Abstract P236: Site-1 Protease-derived Soluble (Pro) Renin Receptor Mediates Angiotensin II-induced Hypertension via Activation of Intrarenal Renin System

Hypertension ◽  
2017 ◽  
Vol 70 (suppl_1) ◽  
Author(s):  
Kexin Peng ◽  
Fei Wang ◽  
Chuanming Xu ◽  
Xiaohan Lu ◽  
Tianxin Yang
Keyword(s):  
Angiotensin Ii ◽  
Collecting Duct ◽  
Renin Angiotensin System ◽  
Ang Ii ◽  
Blood Pressure Lowering Effect ◽  
Angiotensin System ◽  
Induced Hypertension ◽  
Pressure Lowering ◽  
Cleavage Enzyme ◽  
Renin Content

We have previously shown that collecting duct (CD)-specific deletion of (pro)renin receptor (PRR) attenuates angiotensin II (Ang II)-induced hypertension, accompanied with reduced soluble PRR (sPRR) that exerts antidiuretic action. Recent preliminary and published results demonstrated site-1portease (S1P) but not furin or ADMA19 as the predominant PPR cleavage enzyme. In the present study, we evaluated involvement of S1P-derived sPRR in Ang II-induced hypertension. By radiotelemetry, CD PRR KO mice exhibited reduced MAP on day 7 of Ang II infusion at 300 ng/kg/min as compared with floxed mice (MAP: 118±5 vs. 137±3 mmHg, N=5, p<0.05). Administration of sPRR-His, a histidine-tagged sPRR, at 120 μg/kg/d via i.v. infusion to CD PRR KO mice for additional 7 days largely restored the sensitivity to Ang II (MAP: 139±6 mmHg in sPRR-His +Ang II group vs. 116±5 mmHg in Ang II group, N = 4, p<0.05). The i.v. infusion was achieved via placement of a catheter in jugular vein with the other end connected to mimipump. In C57/BL6 mice, administration of a S1P inhibitor PF429242 (PF) via mini pump infusion at 30 mg/kg/d for 7 days attenuated Ang II-induced increases in MAP (day 7: 125±5 in Ang II+ PF group vs. 142±3 in Ang II group; N=6, p<0.05), urinary sPRR excretion (27±4 vs. 63±9 pg/24h; N=6, p<0.05). In parallel, urinary renin levels were elevated by Ang II, which was blunted by PF (renin activity: 0.17±0.03 in Ang II+PF vs. 0.80±0.081 in Ang II vs. 0.12±0.016 ng/24h in Control, N=6, p<0.01; active renin content: 15.2±2.7 vs. 236.0±23.2 vs. 5.6±1.3 ng/24h, N=6, p<0.01; prorenin content: 9.6±3.1 vs. 27.8±6.1 vs. 6.2±1.8 ng/24h, N=6, p<0.05; total renin content: 24.8±5.2 vs. 263.8±27.0 vs. 11.8±3.0 ng/24h, N=6, p<0.01). An intravenous infusion of sPRR-His counteracted the blood pressure-lowering effect of PF in Ang II-infused mice (MAP: 147±3 in PF+sPRR vs. 126±4 mmHg in PF; N=4, p<0.05). Together, these results suggest that S1P-derived sPRR contributes to Ang II-induced hypertension through activation of intrarenal renin-angiotensin system.

Abstract 023: Collecting Duct (Pro)Renin Receptor Mediates Angiotensin II-induced Hypertension

Hypertension ◽  
2015 ◽  
Vol 66 (suppl_1) ◽  
Author(s):  
Fei Wang ◽  
Kexin Peng ◽  
Xiaohan Lu ◽  
Kevin Yang ◽  
Mi Liu ◽  
...  
Keyword(s):  
Angiotensin Ii ◽  
Direct Action ◽  
Collecting Duct ◽  
Second Phase ◽  
Na Transport ◽  
Dual Effect ◽  
Induced Hypertension ◽  
Nanomolar Range ◽  
Local Generation ◽  
Specific Deletion

Within the kidney, (pro)renin receptor (PRR) is predominantly expressed in the intercalated cells (IC) of collecting duct (CD) where its expression is induced by angiotensin II (AngII). Here we examined the function of PRR in the CD by analyzing mice with CD-specific deletion of PRR (CD PRR KO) using AQP2-Cre which has recently been shown to target both IC and principal cells (PC). Radiotelemetry demonstrated that the null mice were largely resistant to AngII-induced hypertension (MAP on day 7: Floxed/AngII 137.4 ± 3.5 vs. KO/AngII 121.2 ± 1.1 mmHg, p<0.05, n=4), accompanied with reduced urinary soluble PRR (sPRR) and aldosterone levels. Electrophysiology analysis demonstrated that within minutes activation of PRR by 10 nM prorenin induced a transient increase in amiloride-sensitive Na+ transport in cultured mpkCCD cells (Ieq: 1.85 ± 0.17 vs. 1.30 ± 0.06 μA/cm2, p<0.05). Interestingly, this was followed by a second phase of ENaC activation after 6 h, which reached the plateau activation at 24 h, accompanied with increased aldosterone release as assessed by ELISA (14.41 ± 0.92 vs. 5.45 ± 0.28 pg/ ml/μg protein, p<0.05). The chronic but not acute phase of ENaC activation was abolished by eplerenone. Both phases of ENaC activation depended on Nox4-derived reactive oxygen species (ROS). Immunostaining using an antibody against sPRR (the N terminus) showed exclusive labeling in the principle cells (PC) whereas the labeling with the C-terminal antibody was exclusively found in IC. A recombinant histidine-tagged sPRR, termed sPRR-His, in the nanomolar range induced a similar dual effect on ENaC activation as prorenin. Intravenous infusion of sPRR-His in CD PRR KO mice for 5 days completely restored the hypertensive response to AngII (MAP: 135.5 ± 7.5 vs. 116.7 ± 5.7 mmHg, p<0.05). We conclude that: 1) CD PRR mediates AngII-induced hypertension; 2) PRR activation in the CD leads to increased ENaC activity acutely through the direct action of ROS and chronically through local generation of aldosterone; 3) sPRR derived from IC may act in a paracrine fashion to stimulate Na+ transport in PC.

scholarly journals Interaction between Epithelial Sodium Channel γ-Subunit and Claudin-8 Modulates Paracellular Sodium Permeability in Renal Collecting Duct

2020 ◽  
Vol 31 (5) ◽  
pp. 1009-1023 ◽  
Author(s):  
Ali Sassi ◽  
Yubao Wang ◽  
Alexandra Chassot ◽  
Olga Komarynets ◽  
Isabelle Roth ◽  
...  
Keyword(s):  
Sodium Channel ◽  
Knockout Mice ◽  
Collecting Duct ◽  
Epithelial Sodium Channel ◽  
Paracellular Permeability ◽  
Sodium Reabsorption ◽  
Kidney Tubule ◽  
Sodium Permeability ◽  
Tubular Lumen ◽  
Renal Collecting Duct

BackgroundWater and solute transport across epithelia can occur via the transcellular or paracellular pathways. Tight junctions play a key role in mediating paracellular ion reabsorption in the kidney. In the renal collecting duct, which is a typical absorptive tight epithelium, coordination between transcellular sodium reabsorption and paracellular permeability may prevent the backflow of reabsorbed sodium to the tubular lumen along a steep electrochemical gradient.MethodsTo investigate whether transcellular sodium transport controls tight-junction composition and paracellular permeability via modulating expression of the transmembrane protein claudin-8, we used cultured mouse cortical collecting duct cells to see how overexpression or silencing of epithelial sodium channel (ENaC) subunits and claudin-8 affect paracellular permeability. We also used conditional kidney tubule–specific knockout mice lacking ENaC subunits to assess the ENaC’s effect on claudin-8 expression.ResultsOverexpression or silencing of the ENaC γ-subunit was associated with parallel and specific changes in claudin-8 abundance. Increased claudin-8 abundance was associated with a reduction in paracellular permeability to sodium, whereas decreased claudin-8 abundance was associated with the opposite effect. Claudin-8 overexpression and silencing reproduced these functional effects on paracellular ion permeability. Conditional kidney tubule–specific ENaC γ-subunit knockout mice displayed decreased claudin-8 expression, confirming the cell culture experiments' findings. Importantly, ENaC β-subunit or α-subunit silencing or kidney tubule–specific β-ENaC or α-ENaC knockout mice did not alter claudin-8 abundance.ConclusionsOur data reveal the specific coupling between ENaC γ-subunit and claudin-8 expression. This coupling may play an important role in preventing the backflow of reabsorbed solutes and water to the tubular lumen, as well as in coupling paracellular and transcellular sodium permeability.

scholarly journals Proximal Tubule-Specific Deletion of Angiotensin II Type 1a Receptors in the Kidney Attenuates Circulating and Intratubular Angiotensin II–Induced Hypertension in PT- Agtr1a −/− Mice

Hypertension ◽  
2021 ◽  
Author(s):  
Xiao Chun Li ◽  
Ana Paula Oliveira Leite ◽  
Xiaowen Zheng ◽  
Chunling Zhao ◽  
Xu Chen ◽  
...  
Keyword(s):  
Blood Pressure ◽  
Angiotensin Ii ◽  
Fusion Protein ◽  
Proximal Tubule ◽  
Proximal Tubules ◽  
Normal Blood ◽  
Ang Ii ◽  
Normal Blood Pressure ◽  
Wild Type ◽  
Induced Hypertension

The present study used a novel mouse model with proximal tubule-specific knockout of AT 1a receptors in the kidney, PT- Agtr1a −/− , to test the hypothesis that intratubular Ang II (angiotensin II) and AT 1a receptors in the proximal tubules are required for maintaining normal blood pressure and the development of Ang II–induced hypertension. Twenty-six groups (n=6–15 per group) of adult male wild-type, global Agtr1a −/− , and PT- Agtr1a −/− mice were infused with Ang II (1.5 mg/kg per day, IP), or overexpressed an intracellular Ang II fusion protein in the proximal tubules for 2 weeks. Basal telemetry blood pressure were ≈15±3 mm Hg lower in PT- Agtr1a −/− than wild-type mice and ≈13±3 mm Hg higher than Agtr1a −/− mice ( P <0.01). Basal glomerular filtration was ≈23.9% higher ( P <0.01), whereas fractional proximal tubule Na + reabsorption was lower in PT- Agtr1a −/− mice ( P <0.01). Deletion of AT 1a receptors in the proximal tubules augmented the pressure-natriuresis response ( P <0.01) and natriuretic responses to salt loading or Ang III infusion ( P <0.01). Ang II induced hypertension in wild-type, PT- Agtr1a −/− and PT- Nhe3 −/− mice, but the pressor response was ≈16±2 mm Hg lower in PT- Agtr1a −/− and PT- Nhe3 −/− mice ( P <0.01). Deletion of AT 1a receptors or NHE3 (Na + /H + exchanger 3) in the proximal tubules attenuated ≈50% of Ang II–induced hypertension in wild-type mice ( P <0.01), but blocked intracellular Ang II fusion protein-induced hypertension in PT- Agtr1a −/− mice ( P <0.01). Taken together, the results of the present study provide new insights into the critical role of intratubular Ang II/AT 1 (AT 1a )/NHE3 pathways in the proximal tubules in normal blood pressure control and the development of Ang II–induced hypertension.

Functional interaction of COMMD3 and COMMD9 with the epithelial sodium channel

2013 ◽  
Vol 305 (1) ◽  
pp. F80-F89 ◽  
Author(s):  
Yong Feng Liu ◽  
Marianne Swart ◽  
Ying Ke ◽  
Kevin Ly ◽  
Fiona J. McDonald
Keyword(s):  
Cell Surface ◽  
Sodium Channel ◽  
Collecting Duct ◽  
Extracellular Fluid ◽  
Copper Metabolism ◽  
Epithelial Sodium Channel ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Endogenous Regulators ◽  
Collecting Duct Cells

The epithelial sodium channel (ENaC) plays an important role in controlling Na+ homeostasis, extracellular fluid volume, and blood pressure. Copper metabolism Murr1 domain-containing protein 1 (COMMD1) interacts with ENaC and downregulates ENaC. COMMD1 belongs to the COMMD family consisting of COMMD1–10, and all COMMD family members share a C-terminal COMM domain. Here, we report that COMMD2–10 also interacts with ENaC, and COMMD3 and COMMD9 were selected for further study. Amiloride-sensitive current in mammalian epithelia expressing ENaC was significantly reduced by COMMD3 or COMMD9, and ENaC expression at the cell surface was significantly decreased in the presence of COMMD3 or COMMD9. COMMD3 and COMMD9 retained their ability to reduce current when COMMD1 was knocked down. COMMD3 and COMMD9 were widely expressed in kidney and were colocalized with ENaC in renal collecting duct cells. These data suggest that COMMD3 and COMMD9 may be endogenous regulators of ENaC to regulate Na+ transport through altering ENaC cell surface expression.

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