Abstract 16868: Conditional Renal Tubule Specific Deletion of KDM6a Gene Causes Hypertension in Mice

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Xiaobin Han ◽  
Leah Akinseye ◽  
Kai Chen ◽  
Zhongjie Sun
Keyword(s):  
Blood Pressure ◽  
Kidney Function ◽  
Renal Tubule ◽  
Erk Signaling ◽  
Thick Ascending Limb ◽  
Artery Cannulation ◽  
Histone 3 ◽  
H3k27me3 Level ◽  
Specific Deletion

Objective: Aging-associated downregulation of Klotho is partly due to epigenetic upregulation of H3K27me3 in the kidney. The purpose of this study is to investigate the potential role of KDM6a, a demethylase of histone 3 lysine (K) 27 trimethylation (H3K27me3), in the regulation of blood pressure and explore the related molecular pathways that may influence the kidney function during aging. Methods and Results: Wild type and Ksp-Cre/KDM6a-floxed mice were divided into vehicle control and tamoxifen (10 mg/kg/mouse/7days) treated cohorts (WT mice: KDM6a floxed or Ksp-Cre only and Ksp-Cre/KDM6a -loxed mice). Blood pressure (BP) was measured by tail cuff method and confirmed by carotid artery cannulation at the end of study. Deletion of the KDM6a gene (KDM6a-cKO) significantly increased BP at day 7. BP remained elevated throughout the study. We found that the H3K27me3 level was significantly increased in the kidneys of KDM6a-cKO mice compared to control mice confirming that KDM6a is an important H3K27 demethylase in the control of methylation levels in the kidney. Elevation of BP was likely due to upregulation of the Na + :K + :2Cl - co-transporter (NKCC2) in the thick ascending limb of Henle's loop and NaCl co-transporter (NCC) in the distal convoluted tubule confirmed by qPCR and immunohistochemistry. Accordingly, we found that the urine sodium level was decreased in KDM6a-cKO mice compared to KDM6a-con mice. We showed that expression of aquaporin 2 (AQP2) was increased in the kidney of KDM6a-cKO mice, suggesting that AQP2 may also contribute to increased BP through modulation of body water homeostasis. Furthermore, we demonstrated that expression of Klotho was downregulated by 50%, which blocked FGFR1/Klotho/ERK signaling in the kidney of KDM6a-cKO mice. Notably, expression of aging markers including p53, p21, and p16 was significantly increased in the kidney of KDM6a-cKO mice, indicating that deletion of KDM6a in the renal tubule results in kidney aging. Conclusion: KDM6A is essential to the maintenance of normal kidney function and blood pressure. Renal-specific KDM6A knockout-induced hypertension is likely attributed to increased H3K27me3 levels and the resultant dowregulation of Klotho gene expression which impairs the FGFR1/Klotho/ERK signaling.

MO092THE ROLE OF CALCIUM IN UROMODULIN EXPRESSION AND SECRETION FROM RENAL MEDULLARY EPITHELIAL CELLS OF HYPERTENSIVE AND NORMOTENSIVE RATS

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
Philipp Boder ◽  
Sheon Mary ◽  
Lesley Graham ◽  
Christian Delles
Keyword(s):  
Blood Pressure ◽  
Epithelial Cells ◽  
Mrna Level ◽  
Extracellular Calcium ◽  
Female Rats ◽  
Thick Ascending Limb ◽  
Molecular Signaling ◽  
Extracellular Secretion ◽  
Wistar Kyoto

Abstract Background and Aims Uromodulin (UMOD) is the most abundantly secreted protein found within the urine, primarily produced by medullary thick ascending limb (mTAL) epithelial cells of the kidneys. There is accruing genetic evidence implicating UMOD in blood pressure regulation and consequently hypertension. The molecular signaling induced by calcium in the kidney and its influence on blood pressure are not well understood. The aim of this study was to investigate the potential role of extracellular calcium and the calcium-sensing receptor (CaSR) in mTAL on UMOD production and secretion in TAL cells with the hope of defining novel clinical targets for the treatment of hypertension. Method Kidneys were harvested from normotensive Wistar-Kyoto (WKY) and stroke-prone spontaneously hypertensive (SHRSP) female rats. To determine the effect of extracellular calcium on UMOD secretion, mTAL tubules were incubated in media with and without 1mM calcium, nifedipine (10µM), NPS2143 (1 or 5 µM) and spermine (2mM). Extracellular and intracellular UMOD protein levels were detected by Western blot. Gene expression of Umod was determined by qRT-PCR. Results Calcium increased mTAL tubule UMOD secretion in WKY and SHRSP. Nifedipine slightly decreased UMOD secretion in WKY without calcium. In both strains, NPS2143 increased calcium-induced UMOD secretion, with an enhanced effect in SHRSP. Stimulation of CaSR with spermine decreased UMOD secretion in WKY. Analysis of intracellular UMOD levels in these conditions demonstrated increased accumulation when extracellular secretion was low, and vice versa. Incubation of primary mTAL cells with calcium confirmed increased localisation of UMOD at the membrane compared to the cytosol, without any major differences in cell morphology. The Umod mRNA level changes were not statistically significant among conditions. Conclusion Trafficking of UMOD in the mTAL is influenced by the type of CaSR ligand and the biased nature of G-protein coupled CaSR signalling. Unravelling the signalling events post-calcium will be necessary for identification of key regulators of UMOD secretion and provide new sites for therapeutic intervention in hypertension.

Abstract 215: Adam17 Regulates Renal Function

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_4) ◽  
Author(s):  
Bo Zhang ◽  
Kai Chen ◽  
Zhongjie Sun
Keyword(s):  
Renal Function ◽  
Kidney Function ◽  
Renal Tubule ◽  
Renal Diseases ◽  
Normal Kidney ◽  
Normal Kidney Function ◽  
Membrane Bound ◽  
Tubule Cells ◽  
A Disintegrin And Metalloproteinase

A disintegrin and metalloproteinase 17 (ADAM17) is a ubiquitously expressed membrane-bound sheddase that cleaves a diverse variety of membrane-bound molecules, including cytokines, growth factors, and their receptors to activate or inactivate various cellular signaling pathways. Although it was reported that ADAM17 may mediate renal diseases, the role of ADAM17 in the regulation of normal kidney function has never been identified. The objective of this study is to investigate whether renal ADAM17 plays a role in maintaining normal kidney function and structure. Tamoxifen-inducible kidney-specific cre (Ksp) and ADAM17-floxed mice were cross-bred for generating Ksp/ADAM17-floxed mice. Injection of tamoxifen initiated deletion of the ADAM17 gene in renal tubule cells. We found that conditional kidney-specific knockout of ADAM1 7 gene (Ksp-ADAM17 -/-) decreased urinary creatinine and sodium excretion were decreased in Ksp-ADAM17 -/- mice, indicating that ADAM17 gene deficiency impairs kidney function. H&E staining showed glomerulus collapse and tubule dilation in Ksp-ADAM17 -/- mice. The epithelial cells fall off into the lumen in the renal tubule. Mesangial expansion and fibrosis were found in glomeruli in Ksp-ADAM17 -/- mice. Moreover, apoptosis was increased in tubule cells in both cortex and medulla areas in Ksp-ADAM17 -/- mice. In conclusion, ADAM17 is critical to the maintenance of normal renal function and structure.

scholarly journals Salt loading decreases urinary excretion and increases intracellular accumulation of uromodulin in stroke-prone spontaneously hypertensive rats.

2021 ◽  
Author(s):  
Sheon Mary ◽  
Philipp Boder ◽  
Giacomo Rossitto ◽  
Lesley Graham ◽  
Kayley Scott ◽  
...  
Keyword(s):  
Blood Pressure ◽  
Spontaneously Hypertensive Rats ◽  
Ex Vivo ◽  
Chronic Hypertension ◽  
Thick Ascending Limb ◽  
Mrna Levels ◽  
Hypertensive Rats ◽  
Salt Loading ◽  
Spontaneously Hypertensive

Uromodulin (UMOD) is the most abundant renal protein secreted into urine by the thick ascending limb (TAL) epithelial cells of the loop of Henle. Genetic studies have demonstrated an association between UMOD risk variants and hypertension. We aimed to dissect the role of dietary salt in renal UMOD excretion in normotension and chronic hypertension. Normotensive Wistar-Kyoto rats (WKY) and stroke-prone spontaneously hypertensive rats (SHRSP) (n=8/sex/strain) were maintained on 1% NaCl for three weeks. A subset of salt-loaded SHRSP was treated with nifedipine. Salt-loading in SHRSP increased blood pressure (ΔSBP 35 ± 5 mmHg, p<0.0001) and kidney injury markers such as KIM-1 (fold change, FC 3.4; p=0.003), NGAL (FC, 2.0; p=0.012) and proteinuria. After salt-loading there was a reduction in urinary UMOD excretion in WKY and SHRSP by 26% and 55% respectively, compared to baseline. Nifedipine treatment reduced blood pressure in SHRSP, however, did not prevent salt-induced reduction in urinary UMOD excretion. In all experiments, changes in urinary UMOD excretion were dissociated from kidney UMOD protein and mRNA levels. Colocalization and ex-vivo studies showed that salt-loading increased intracellular UMOD retention in both WKY and SHRSP. Our study provides novel insights into the interplay between salt, UMOD, and blood pressure. The role of UMOD as a cardiovascular risk marker deserves mechanistic reappraisal and further investigations based on our findings.

Abstract 014: Collecting Duct Specific Deletion of the (Pro)renin Receptor Modulates ENaC Expression

Hypertension ◽  
2016 ◽  
Vol 68 (suppl_1) ◽  
Author(s):  
Nirupama Ramkumar ◽  
Deborah Stuart ◽  
Kai Song ◽  
Nikita Abraham ◽  
Shuping Wang ◽  
...  
Keyword(s):  
Blood Pressure ◽  
Collecting Duct ◽  
Plasma Renin ◽  
Cre Recombinase ◽  
Protein Isoforms ◽  
Ang Ii ◽  
Renal Tubular ◽  
Renal Expression ◽  
Specific Deletion

The renal tubular (pro)renin receptor (PRR) has been shown to modulate water balance, blood pressure and Na + homeostasis. We recently reported that inducible nephron wide deletion of the PRR results in Na + wasting, reduced epithelial Na + channel (ENaC) expression in the kidney and attenuated hypertensive response to angiotensin-II (Ang-II) infusion. In this study, we examined the effects of PRR deletion in collecting duct (CD) specific mouse models targeting either the principal cells (PC) or intercalated cells (IC). PC-specific PRR knockout (KO) mice were obtained by crossing floxed PRR mice with mice harboring AQP-2 Cre recombinase. Compared to floxed mice, PC specific KO PRR mice had no differences in PRR immunostaining but had 50% reduction in PRR mRNA in micro-dissected cortical CDs. No differences in blood pressure were observed between the two groups at baseline or following Ang-II infusion at 600 ng/kg/min. Similarly, plasma renin concentration and renal expression of ENaC protein isoforms were comparable between the two groups. To achieve IC-specific PRR deletion, floxed PRR mice were bred with mice expressing B-1 Cre recombinase. Compared to floxed controls, IC-specific PRR KO mice were smaller (KO body weight: 5.9 ± 1.3 g vs controls: 11.1± 1.2 g) and did not survive beyond 30 days after birth. IC-specific PRR KO mice also demonstrated marked reduction in renal medullary PRR immunostaining along with decreased renal expression of ENaC-α protein (50% reduction compared to controls), similar to the findings in nephron wide deletion of PRR. Taken together, these findings suggest that IC specific deletion of PRR but not PC-specific deletion modulates renal ENaC expression. Further studies evaluating ENaC activity in isolated cortical CDs from PC and IC specific PRR KO mice will help delineate the functional role of CD PRR in Na + homeostasis.

scholarly journals Renal localization and function of the tight junction protein, claudin-19

2007 ◽  
Vol 293 (1) ◽  
pp. F166-F177 ◽  
Author(s):  
Susanne Angelow ◽  
Randa El-Husseini ◽  
Sanae A. Kanzawa ◽  
Alan S. L. Yu
Keyword(s):  
Tight Junction ◽  
Renal Tubule ◽  
Divalent Cations ◽  
Physiological Role ◽  
Hereditary Disease ◽  
Paracellular Transport ◽  
Thick Ascending Limb ◽  
Peripheral Neurons ◽  
Junction Protein

Claudins form a family of transmembrane tight junction proteins that play a key role in control and selectivity of paracellular transport. Mutations in claudin-19, which is expressed in kidney, retina, and myelinated peripheral neurons, were identified in familial hypomagnesemia with hypercalciuria and nephrocalcinosis, a hereditary disease causing renal Mg2+ and Ca2+ wasting. Here, we studied the distribution and possible functional role of claudin-19 in the renal tubule. By immunofluorescence staining of mouse kidney, claudin-19 was found to be expressed at the tight junction of the thick ascending limb of Henle, the major site of paracellular Mg2+ reabsorption, where it colocalized with claudin-16, as well as in the thin ascending limb. The role of claudin-19 in paracellular transport was tested by stable transfection into Madin Darby canine kidney II TetOff cells to generate inducible cell lines. Claudin-19 increased the transepithelial electrical resistance and decreased permeability to monovalent and divalent cations, while anion and urea permeability were not affected. Our data suggest that claudin-19 acts as a selective cation barrier at the tight junction. This would be consistent with its physiological role to electrically seal myelinated peripheral neurons. The normal role of claudin-19 in renal tubule function remains to be determined.

Abstract 101: Alms1 (Alstrom Syndrome 1), a Novel Gene Involved in Blood Pressure Regulation, Renal Na Handling and Thick Ascending Limb (TAL) Function

Hypertension ◽  
2016 ◽  
Vol 68 (suppl_1) ◽  
Author(s):  
Ankita Bachhawat Jaykumar ◽  
Paulo Caceres ◽  
Gustavo Ares ◽  
William H Beierwaltes ◽  
Pablo A Ortiz
Keyword(s):  
Blood Pressure ◽  
Urine Volume ◽  
Dietary Factors ◽  
Thick Ascending Limb ◽  
Nucleotide Polymorphisms ◽  
Interacting Protein ◽  
Alström Syndrome ◽  
Alstrom Syndrome

Single nucleotide polymorphisms in the Alstrom syndrome 1 (ALMS1) gene are associated to hypertension, renal dysfunction, and obesity in the general population. The role of ALMS1 in regulating blood pressure or renal Na handling is unknown. In a proteomics screen, we identified ALMS1 as an interacting protein with the Na/K/2Cl cotransporter NKCC2. NKCC2 mediates NaCl reabsorption by the TAL. Thus, we hypothesized that ALMS1 regulates NKCC2 endocytosis and activity as well as blood pressure. First, we confirmed the expression of ALMS1 in isolated perfused TALs. To study ALMS1 function, we generated ALMS1 knockout (KO) rats in collaboration with the rat genome editing consortium at MCW and confirmed deletion of the ALMS1 gene. We found that 3 month old ALMS1 KO rats are hypertensive compared to wild type littermates (ALMS1 MAP: 141 ± 5 mmHg vs WT: 99 ± 6 mmHg, p< 0.005) fed a normal Na diet. We measured surface and intracellular NKCC2 and found a higher percentage of NKCC2 at the surface in TALs from ALMS1 KO (ALMS1: 13.8 ± 1.2% vs WT: 9.1 ± 1.0%, p<0.05, n=6). The increase in surface NKCC2 is due to lower endocytosis because the rate of NKCC2 internalization was lower in TALs from ALMS1 KO (ALMS1: 13.1 ± 1.2% vs WT: 28.2 ± 2.8% over 20 min, p<0.01, n=6). To study NKCC2-mediated Na transport in vivo , we measured bumetanide-induced natriuresis and diuresis. ALMS1 KO rats exhibited higher bumetanide-induced natriuresis (ALMS1: 1292 ± 65 vs WT: 564 ± 31 μmoles, p<0.01, n=5) and diuresis (ALMS1: 3.1 ± 0.32 vs WT: 1.6 ± 0.13 ml, p<0.05), indicative of higher TAL Na reabsorption. To study if this decreases the ability to excrete a volume and Na load in ALMS1 KO, we instrumented anesthetized rats and measured sodium excretion (U Na V) and urine volume (UV) over 150 min after an acute saline load. We found that the cumulative U Na V was lower in ALMS1 KO rats (ALMS1: 72 ± 38 vs WT: 219 ± 55 μmoles, p<0.05, n=8) as was the UV (ALMS1: 0.70 ± 0.24 vs WT: 1.78 ± 0.39 ml, p<0.05). We conclude that deletion of ALMS1 decreases NKCC2 endocytosis and increases TAL Na reabsorption. Thus, the hypertension observed in ALMS1 KO rats may be in part due to higher renal Na reabsorption. It is not known whether the expression of ALMS1 protein is decreased in hypertensive patients or lowered by dietary factors that increase BP in humans.

scholarly journals A two-hit mechanism for sepsis-induced impairment of renal tubule function

2013 ◽  
Vol 304 (7) ◽  
pp. F863-F874 ◽  
Author(s):  
Bruns A. Watts ◽  
Thampi George ◽  
Edward R. Sherwood ◽  
David W. Good
Keyword(s):  
Renal Tubule ◽  
Basolateral Membrane ◽  
Cecal Ligation ◽  
Erk Signaling ◽  
Thick Ascending Limb ◽  
Loss Of Function ◽  
Inhibitory Mechanisms ◽  
Erk Phosphorylation ◽  
Underlying Mechanisms

Renal insufficiency is a common and severe complication of sepsis, and the development of kidney dysfunction increases morbidity and mortality in septic patients. Sepsis is associated with a variety of defects in renal tubule function, but the underlying mechanisms are incompletely understood. We used a cecal ligation and puncture (CLP) model to examine mechanisms by which sepsis influences the transport function of the medullary thick ascending limb (MTAL). MTALs from sham and CLP mice were studied in vitro 18 h after surgery. The results show that sepsis impairs the ability of the MTAL to absorb HCO3−through two distinct mechanisms. First, sepsis induces an adaptive decrease in the intrinsic capacity of the tubules to absorb HCO3−. This effect is associated with an increase in ERK phosphorylation in MTAL cells and is prevented by pretreatment of CLP mice with a MEK/ERK inhibitor. The CLP-induced reduction in intrinsic HCO3−absorption rate appears to involve loss of function of basolateral Na+/H+exchange. Second, sepsis enhances the ability of LPS to inhibit HCO3−absorption, mediated through upregulation of Toll-like receptor 4 (TLR4)-ERK signaling in the basolateral membrane. The two inhibitory mechanisms are additive and thus can function in a two-hit capacity to impair renal tubule function in sepsis. Both effects depend on ERK and are eliminated by interventions that prevent ERK activation. Thus the TLR4 and ERK signaling pathways represent potential therapeutic targets to treat or prevent sepsis-induced renal tubule dysfunction.

Pre-emptive Analgesia for Postoperative Pain Relief in Children – Role of Paracetamol

2009 ◽  
pp. 9-16
Author(s):  
Rubina Yasmin ◽  
AKM Akhtaruzzaman ◽  
Paresh Chandra Sarker ◽  
Neaz Ahmed ◽  
Ranadhir Kumar Kundu ◽  
...  
Keyword(s):  
Blood Pressure ◽  
Heart Rate ◽  
Postoperative Pain ◽  
Diclofenac Sodium ◽  
Total Duration ◽  
Pain Scale ◽  
Prospective Clinical Study ◽  
High Dose ◽  
Induction Of Anaesthesia

This prospective clinical study was carried out in the Dept. of Anaesthesia, Analgesia and Intensive Care Medicine, BSMMU, Dhaka, during the period of May 2003 to July 2003. The study was done to emphasize the importance of giving analgesics preemptively instead of waiting for the child to complain of pain and to produce smooth recovery after surgery by decreasing immediate postoperative pain in children by a simple, safe acceptable drug. The children scheduled for tonsillectomy under general anaesthesia were recruited in this study. The analgesic efficiency of rectal paracetamol in two doses, 25 mg/kg bodywt.(Gr-P25) and 50 mg/kg. bodywt. (Gr-P50) were compared with Diclofenac Sodium suppository 1mg/ kg body weight (Gr-D) given half an hour before induction of anaesthesia. Pain scoring was done by TPPPS (Toddler Pre-schooler postoperative pain scale). Heart rate and blood pressure were stable in Gr-P50 and Gr-D. Time of first demand of analgesic was delayed in Gr-P50 and Gr-D. Total paracetamol consumption in 24 hours was less in Gr-P50(181±14.25) and Gr-D (212±25) than Gr-P25(318± 26.39). Total duration of analgesia in Gr- P50 (657±9.94) mins. and in Gr- D(502±10.63) mins. and in Gr-P25(288±23.17) mins. Pre-emptive high dose rectal paracetamol appears to be more effective than diclofenac sodium suppository for postoperative analgesia in children undergoing tonsillectomy. Journal of BSA, Vol. 18, No. 1 & 2, 2005 p.9-16

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