bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sodium butyrate suppresses Angiotensin II‐induced hypertension by inhibition of renal (pro)renin receptor and intrarenal RAS
Authors: Wang Lei , Zhu Qing , Lu Aihua , Zhang Linlin , Xu Chuangmin , Liu Xiyang , Yang Tianxin 
Published: November 2023

BackgroundButyrate, one of Short‐chain fatty acids, is end product produced from the fermentation of complex carbohydrates by the gut microbiota. Recently, sodium butyrate (NaBu) is found to play protective roles in a number of chronic diseases. However, it is still unclear whether NaBu processes a therapeutic potential in hypertension. The present study was aimed to investigate the role of NaBu in Ang II‐induced hypertension and further explore the underlying mechanism.MethodsAng II (200 ng/min) was infused in uninephrectomized Sprague‐Dawley rats plus intramedullary infusion of NaBu (1μg/kg·min) or not for 14 days. Blood pressure were monitored by the telemetry system. The renal tissues and serum samples as well as the last 24‐hour urine samples were collected to examine renal injury and regulation of (pro)renin receptor (PRR) and renin.ResultsIntramedullary infusion of NaBu in Sprague‐Dawley rats lowered the Ang II‐induced mean arterial pressure from 128.8 ± 6.2 mmHg to 108.0 ± 3.9 mmHg (P<0.01). This was paralleled with improvement of Ang II‐induced renal injury, including urinary albumin (μg/24h: Ctrl 270.7±49.2 vs AngII 1012.0±219.2 vs AngII‐NaBu 449.4±73.3, P<0.05), glomerulosclerosis (GDI: Ctrl 1.48±0.08 vs AngII 2.70±0.06 vs AngII‐NaBu 1.68±0.08, P<0.01), and renal fibrosis (α‐SMA protein: Ctrl 1.00±0.06 vs AngII 1.38±0.05 vs AngII‐NaBu 0.64±0.05, P<0.01), as well as the expression of inflammatory mediators TNF‐α and IL‐6. The renal protein expression of (pro)renin receptor, urinary excretion of soluble (pro)renin receptor and renin were all increased by AngII infusion but decreased by NaBu treatment. In cultured IMCD cells, NaBu treatment attenuated AngII‐induced expression of PRR and renin.ConclusionThese results demonstrated NaBu exerts antihypertensive action likely via suppressing PRR‐mediated intrarenal RAS.Support or Funding InformationThis work was supported by National Natural Science Foundation of China Grant No. 91439205, No. 31330037 and No. 81600322; Fundamental Research Funds of Sun Yat‐sen University 16ykpy47; Veterans Affairs Merit Review, and National Institutes of Health Grant DK094956. T. Yang is Research Career Scientist in the Department of Veterans Affairs.
Renal‐Specific Dendritic Cell Depletion Reduces Urinary Interleukin 6 Levels Following Angiotensin II‐Mediated Hypertension and Dietary High Salt
Authors: Wynne Brandi Michele , Hecht Gillian G , Kronk Trinity Augusta , van Elst Henrieke , Tansey Malu , Eaton Douglas C , Hoover Robert S. 
Published: November 2023

During hypertension, dendritic cells (DCs) have been shown to activate other immune cells and secrete cytokines, such as interleukin 6 (IL‐6). Recently, DCs were found to form an intricate network within the renal cortex and especially along the distal nephron. Studies reveal that renal‐specific DCs (rDCs) play an important role in the progression of hypertension. Previously, our data suggest that IL‐6 can increase sodium chloride cotransporter (NCC) expression and activity. We hypothesize that rDCs modulate intra‐renal levels of IL‐6 during hypertension or high salt (HS), which may affect distal nephron sodium reabsorption.To test this hypothesis, mice lacking a functional CX3CR1 chemokine receptor (CX3CR1‐EGFP+/+ or rDC mice) were used. CX3CR1 is required for DC localization to the kidney; thus these mice have a renal‐specific DC‐depletion. Mice were either given: vehicle or angiotensin II (AngII) (60ng/min, 28 days + 4% high salt) via osmotic minipump or fed a high salt (4%, 7 days) or normal diet. Systolic BP measurements were obtained with tail‐cuff plethysmography. Metabolic cage studies were performed and urine collected. Urine and serum were used to perform a cytokine array via Meso‐Scale Discovery system. Whole kidney cortex lysates were used for mRNA isolation and RT‐PCR.Mice with renal‐specific loss of DCs had a decreased systolic blood pressure at baseline. Following AngII hypertension, similar BP increases were observed (Δ38mmHg rDC vs. Δ36mmHg Wt). Baseline serum IL‐6 levels were not different; however, AngII‐infusion resulted in ≈3 fold decrease in IL‐6 mRNA in rDC mice when compared to Wt mice. At baseline, urinary IL‐6 levels were not different between rDC and Wt (4.41 ± 0.85 pmol rDC vs. 4.14 ± 1.20 pmol Wt, n=7–10) mice. However, HS feeding increased urinary IL‐6 levels in Wt mice, which was prevented in rDC mice (2.03 ± 0.20 pmol rDC vs. 7.34 ± 2.51 pmol Wt, n=6, p<0.05).These data suggest that baseline BP is modulated by resident innate immune cells specific to the kidney and rDC loss reduces systolic BP; this reduction is maintained during AngII‐hypertension. Notably, our data demonstrate that while systemic IL‐6 levels are unchanged with rDC‐depletion, renal mRNA and urinary IL‐6 levels are decreased during hypertension or HS feeding. Thus together, these data propose a model for rDC‐mediated increases in IL‐6 levels during hypertension or increased salt consumption, altering the pro‐inflammatory milieu and increasing distal sodium reabsorption.Support or Funding InformationNIDDK085097 and Department of Veteran Affairs MERIT Award I01BX002322‐01 to RSH; 2T32DK7656‐21 Emory Renal Division to BMW; R37 DK037963 to DCE;
Intracellular Cl− and the Scaffold Protein Mo25/Cab39 Regulate WNK‐SPAK/OSR1 Signaling in Transepithelial Ion Transport
Authors: Rodan Aylin , Wu Yipin , Sun Qifei , Humphreys John , Schellinger Jeffrey , Goldsmith Elizabeth 
Published: November 2023

ObjectiveThe WNK (with‐no‐lysine kinase)‐SPAK (Ste20/SPS1‐related proline/alanine‐rich kinase)/OSR1 (oxidative stress responsive‐1) kinase cascade regulates SLC12 family cation chloride cotransporters. Here, we investigate mechanisms of pathway activation in the regulation of transepithelial ion transport in the Drosophila melanogaster renal tubule.MethodsTransepithelial K+ flux in the Drosophila tubule main segment was measured using the Ramsay assay and ion‐specific electrodes in control and mutant tubules. Intracellular Cl− concentration was measured using the transgenic sensor ChlopHensor. Phosphorylation of transgenically expressed kinase‐dead rat SPAK was used as a readout of tubule WNK activity. Drosophila WNK autophosphorylation was measured using mass spectrometry. The effect of Mo25 on Fray activity was measured using in vitro kinase assays.ResultsWe have previously demonstrated that transepithelial K+ flux is increased in Drosophila renal tubules bathed in hypotonic medium in a WNK‐SPAK/OSR1 (fray in flies)‐NKCC dependent manner. We hypothesized that in hypotonic conditions, intracellular Cl− is decreased, leading to WNK activation. Intracellular Cl− was 30±4 mM in isotonic conditions; after switching to hypotonic medium, Cl− decreased to 20±3 mM at 10 minutes, and 16±2 and 15±2 mM at 30 and 60 min. Cl− inhibits Drosophila WNK kinase domain autophosphorylation in vitro: 13% of Drosophila WNK was autophosphorylated at Ser434 after 4 hours in 150 mM NaCl, compared to 52% in 50 mM NaCl. Paralleling the fall in Cl− in the tubule in hypotonic conditions, tubule WNK activity increased at 30 and 60 minutes after the tubules were transferred to hypotonic medium. However, overexpression of a predicted Cl−‐insensitive WNK, WNKL421F, during adulthood was not sufficient to increase transepithelial K+ flux in isotonic conditions. We hypothesized that the scaffold protein, Mo25/Cab39, may additionally be required. Using purified Drosophila proteins, we showed that Mo25 stimulates Fray phosphorylation of the fly NKCC in vitro. Mo25 knockdown in the tubule did not alter transepithelial K+ flux in isotonic conditions, but abolished hypotonic stimulation of K+ flux, indicating that Mo25 is required for maximal activation of the pathway. Furthermore, while overexpression of Mo25 in adult tubules did not increase K+ flux in isotonic conditions, simultaneous overexpression of Mo25 and WNKL421F increased K+ flux.ConclusionIntracellular Cl− and Mo25 act together to stimulate WNK‐SPAK/OSR1‐regulated transepithelial ion transport.Support or Funding InformationNational Institutes of Health
Modularity in the ENaC/Degenerin Family Facilitates Transfer of Functional Properties Between Family Members
Authors: DeMarco Michael A , Wang Xue‐Ping , Balchak Deidra M , Powell Koehler , Mukherjee Anindit , Blobner Brandon M , Kashlan Ossama B 
Published: November 2023

Members of the epithelial Na+ channel (ENaC)/Degenerin family are cation selective channels that respond to the extracellular environment. ENaCs and acid sensing ion channels (ASICs) are found in all tetrapods and are involved in diverse processes ranging from renal Na+ reabsorption and regulation of airway surface liquid height (ENaCs) to pain sensation and fear conditioning (ASICs). ASIC and ENaC subunits share significant sequence identity both in the core of the extracellular fold, characterized by β‐sheets, and in the transmembrane helices. The three peripheral helical domains of the extracellular structure, most notably the so‐called finger domain, are poorly conserved. We hypothesized that diversity of function in this protein family rose from an inherently modular architecture. To test our hypothesis, we swapped the finger domain of the mouse ENaC α subunit into mouse ASIC1, and examined its functional and biochemical properties. When we expressed chimeric channels in Xenopus oocytes and assessed function by two‐electrode voltage clamp, we found that the channel had functional attributes of both channels. Like ENaC, channels were constitutively active, and like ASIC1, channels were activated by acidification. Channels exhibited a basal current at pH 8. Upon rapid acidification to pH 4.5, currents substantially increased, and did not desensitize as is characteristic for ASIC1 currents. Consistent with retention of an ASIC1 pore, these channels were permeable to most cations tested, including Na+, K+, and Ca2+, and were blocked by benzamil, an amiloride analog. ENaC α subunits are cleaved at R205 and R231 by furin in the finger domain during biosynthesis, effectively activating quiescent channels. To examine whether the chimeric channel was similarly processed, we added an HA epitope tag to the C‐terminus of the channel and performed an immunoblot. We observed two bands at sizes expected for full length and furin‐cleaved channels. Furthermore, mutating the putative furin‐cleavage sites within the swapped‐in ENaC α subunit finger domain eliminated the lower molecular weight band, but had no apparent effect on channel activity. We also tested whether swapping the ENaC α subunit finger domain into ASIC1 transferred other functional properties, but found none. Our data suggest that the ASIC1 finger domain is critical to channel desensitization, and like the ENaC finger domain, functions as an autoinhibitory domain. Our data suggests a modular architecture for ENaC/Degenerin proteins, and that specific channel functions can be attributed to specific domains.Support or Funding InformationThis work is supported by NIH grant DK098204.
Chronic Loop Diuretic Treatment Mediates Segment‐Specific Hypertrophy in the Nephron
Authors: Krauson Aram , Walczak Elisabeth , Iyer Sonali , Velarde Nona , Nizar Jonathan , Bhalla Vivek 
Published: November 2023

Furosemide, a commonly prescribed loop diuretic to treat hypertension and edema, increases salt excretion by inhibiting reabsorption via the sodium‐potassium 2 chloride cotransporter (NKCC2) in the thick ascending limb. Chronic use of loop diuretics can lead to resistance and the need for higher doses and/or additional classes of diuretics with further long term adverse effects. We previously have shown mice chronically fed furosemide develop hyperplasia in the nephron after 7 days and hypertrophy after 21 days. Furthermore, our data suggested the IGF‐1/IGF‐1R mediated pathway to be involved in tubular hypertrophy. From chronic furosemide treatment, cell size increased in the proximal tubule, distal tubule, and connecting tubule/cortical collecting duct segments. In contrast, long term furosemide treatment had no effect on cell size in the furosemide‐inhibitable thick ascending limb. Using novel, inducible kidney tubule‐specific IGF‐1R knockout mice (iT‐IGF‐1R KO), we demonstrate diminished kidney growth in furosemide‐treated knockout mice only in the proximal tubule. In contrast, knocking out IGF‐1R did not affect cell size increase in the distal tubule or the connecting tubule/cortical collecting duct. These results suggest long term furosemide use induces IGF‐1R‐dependent hypertrophy of the proximal tubule but IGF‐1R‐independent hypertrophy of the distal tubule. Interestingly, using immunofluorescence, IGF‐1R is not detected in the proximal tubule but in more distal segments implicating paracrine activation in hypertrophy. Overall, we demonstrate furosemide‐treated mice undergo segment‐specific hypertrophic responses and mechanisms. To uncover the mechanisms of these segment‐specific responses we are using single cell RNA seq analysis and data will be forthcoming.Support or Funding InformationNIH/NIDDK: 5T32DK007357‐32 (T32), DK09156501A1 (R01)
Calcium‐Mediated Activation of the BK K+ Secretory Channel in Collecting Duct Cells is Dynamically Regulated by the Ca2+‐Activated SK3 and IK1 K+ Channels
Authors: Li Yue , Hu Hongxiang , Tian Jin Bin , Zhu Michael , O'Neil Roger 
Published: November 2023

The Ca2+‐activated BK K+ channel is the dominant pathway for flow‐ and Ca2+‐induced K+ secretion/K+ wasting in collecting duct system (CDS) of the kidney. The goal of the current study was to determine the role of two newly identified Ca2+‐activated K+ channels in CDS, SK3 and IK1 (expressed at both luminal and anti‐luminal cell borders), on BK channel activation (expressed at the luminal border). Like BK, these KCa channels are activated by TRPV4‐induced activation of Ca2+ influx (Berrout et al., PLoS ONE 9:e0156368; Li et al., PLoS ONE 11:e0155006, 2016), but have a much higher Ca2+‐binding affinity (300–600 nM) compared to BK (1–11 uM), properties consistent with a “Ca2+ sensing” function of SK3/IK1. In mCCDcl1 cells, a K+ secreting mouse collecting duct cell line, activation of each of these KCa channels following TRPV4 activation was observed to hyperpolarizes Vm, where selective inhibition of SK3, IK1 and BK, by addition of selective blockers apamin (300 nM), TRAM‐34 (300 nM), and IbTX (100 nM), respectively, each depolarized Vm and markedly reduce TRPV4‐mediated Ca2+i levels. Further, activation of BK was observed to be directly proportional to the TRPV4‐mediated increase in Ca2+i levels. Little or no activation of BK were observed when stimulated Ca2+i levels were low and strong activation of BK when Ca2+i levels were high. Inhibition of SK3 and IK1 each reduced the stimulated Ca2+i levels as expected. Addition of both apamin and TRAM‐34 during TRPV4 activation markedly depolarized Vm, reduced the increase in TRPV4‐mediated Ca2+i levels from 766 +/− 25 nM to 348 +/− 40 nM (n = 30), and reduced the initial rate of rise of TRPV4‐mediated Ca2+i from 215 +/− 14 to 30 +/− 6 nM/min (n = 30). Further, TRPV4‐mediated activation of BK was markedly reduced or abolished in the presence of apamin and TRAM‐34. It is concluded that SK3 and IK1 are functionally expressed in collecting duct cells and that both are activated by low levels of TRPV4‐mediated Ca2+ influx that leads to an initial hyperpolarization of Vm and further stimulation of Ca2+ influx. This SK3/IK1‐mediated enhance Ca2+ influx, in turn, is critical for activation of the BK channel where we show that strong dynamic coupling between TRPV4 and SK3/IK1 plays a central role in modulating BK channel activity in the CDS.Support or Funding InformationThis work was supported by the American Heart Association, Grant # 14GRNT20380916 and the National Institutes of Health, Grant #: 1R01DK098401
Role of Sodium Hydrogen Exchange Regulatory Factor‐1 (NHERF1) in Aging: Tale of Two Rat Models
Authors: Ahmad Asrar , Pushpakumar Sathnur , Ketchem Corey J , Sen Utpal , Lederer Eleanor DeLand , Khundmiri Syed Jalal 
Published: November 2023

Aging animals develop salt‐sensitive hypertension due, in part, to desensitization of dopamine receptors and sensitization of AT1R in renal proximal tubules. We demonstrated old FBN rats develop salt‐dependent hypertension. Preliminary data showed loss of NHERF1 expression in 22mo old F344 rats. We hypothesized that loss of NHERF1 may contribute to salt reabsorption in aging. To address this hypothesis, Fischer Brown Norway (FBN) rats (1, 4, 12, and 24mo old) were fed 1% or 8% NaCl diet for one week and, urinary volume, sodium, potassium, and chloride and expression of Na‐K ATPase a1 subunit (NKA), NHERF1, NKCC2, NCC, and D1R were measured. Feeding 8% NaCl increased 24 h urine volume compared to rats fed 1% NaCl diet in 1mo (13.6±0.98 vs 29.2±3.8 mL) and 4mo (15.3±1.5 vs 39.2±2.4) but not in 24 mo rats (20.7±4.7 vs 15.9±2.4). UNaV and UClV increased in young rats fed with 8% NaCl but not in old rats. UKV did not change in young rats but significantly decreased in older rats. To our surprise, in contrast to F344 rats, NHERF1 expression increased with age in FBN rats (0.21±.03 (4mo) vs 0.71±0.07 (24mo) AU ratio NHERF1:actin). Increased NHERF1 expression with age in FBN rats was associated with decreased D1R expression (0.8±0.16 vs 0.4±0.04) while in F344 rats D1R expression increased with age (0.6±0.1 vs 0.9±0.1). To determine the role of NHERF1 in the two models of rats, we determined TNFa in kidney homogenates from FBN and F344 rats. TNFa increased with age in both F344 (4.04±0.15 (4 mo) vs 6.9±0.42 (24 mo)) and FBN rats (2.1±0.3 vs 4.05±0.31 ng/mg protein). TNFa levels were significantly higher in young F344 rats as compared to young FBN rats. To determine if lack of NHERF1 is responsible for salt wasting, we measured urine volume and Na in 18 mo old WT and NHERF1 KO mice. The urine volume in WT mice (2.7±0.8 (1% NaCl) vs 5.6±1.2 mL/24 h (8% NaCl)) were significantly lower than KO mice (3.6±0.57(1% NaCl) vs 7.5±0.88 mL/24 h (8% NaCl)). However, there was no significant difference in UNaV between WT and KO mice fed an 8% NaCl diet. D1R expression increased in NHERF1 KO mice as compared to WT mice. TNFa increased significantly in NHERF1 KO mice kidneys as compared to WT mice. We conclude that increased NHERF1 expression as seen in FBN rats result in salt retention while lack of NHERF1 as seen F344 rats and NHERF1 KO mice increases renal inflammation. Further studies are required to delineate the two roles of NHERF1.Support or Funding InformationNIA grants R21AG047474 and R25AG047843 and AHA grant 16GRNT31030019
Assessing the Role of OCRL1 in Proximal Tubule Function in a Lowe Syndrome Cell Model
Authors: Eshbach Megan L. , Shipman Katherine E. , Raghavan Venkatesan , Rbaibi Youssef , Long Kimberly R. , Labilloy Anatália , Baty Catherine J. , Weisz Ora A. 
Published: November 2023

Lowe syndrome is an X‐linked recessive disorder caused by mutations in OCRL, which encodes the phosphatidylinositol 5′‐phosphatase OCRL1. Hallmark symptoms of Lowe syndrome include proximal tubule (PT) dysfunction, typically characterized by low molecular weight proteinuria (LMWP), renal tubular acidosis, aminoaciduria, and hypercalciuria. OCRL1 has been hypothesized to play a role in myriad cellular processes including membrane trafficking, actin dynamics, cell polarity, ciliogenesis, and cell division. However, consensus is lacking for how loss of OCRL1 leads to impaired PT function. To this end, we created a PT cell culture model of the disease. Using the CRISPR/Cas9 genome editing system, we knocked out OCRL1 expression in a human PT cell line. In parallel, we transiently knocked down OCRL1 using siRNA to confirm that chronic depletion did not result in cell adaptation. Neither acute nor chronic depletion of OCRL1 had any effect on endocytic uptake of the megalin/cubilin ligand albumin. However, OCRL1 depleted cells consistently proliferated more slowly and exhibited a higher percentage of multinucleated cells than matched controls. We hypothesize that defects in cell division during kidney development and/or repair after kidney injury may compromise PT length and/or diameter in patients and have developed a mathematical model to assess the effect of such changes on PT albumin uptake. Our data suggest a new explanation for how loss of OCRL1 function results in LMWP as well as the other commonly observed renal manifestations of Lowe syndrome.Support or Funding InformationNational Institutes of Health R01‐DK101484, R01‐DK100357, P30‐DK079307, T32‐DK061296; Lowe Syndrome Association
Corticosterone activates the thiazide‐sensitive NaCl transporter through the glucocorticoid receptor
Authors: Bailey Matthew A , Ivy Jessica R 
Published: November 2023

The Na‐Cl co‐transporter (NCC) is the major route for apical Na entry into cells of the distal convoluted tubule (DCT). Thiazide diuretics inhibit NCC and are a mainstay hypertension treatment. We have previously shown that glucocorticoid excess activates NCC and induces hypertension. The DCT expresses both glucocorticoid receptors (GR) and mineralocorticoid receptors (MR) but does not express the glucocorticoid‐metabolizing enzyme 11bHSD2 at high levels. This suggests that glucocorticoids may directly activate the transporter. Here we investigated whether glucocorticoids activate NCC in vivo and whether GR or MR is involved in this process.Male C57BL6 mice were anaesthetized under isofluorane and implanted with silastic pellets containing spironolactone (50 mg; chronic MR blockade) or RU486 (60 mg; chronic GR blockade). Control mice received no implant. After 5 days, corticosterone (6 mg/kg, s.c.) or vehicle (veh, 2% DMSO, s.c.) was injected at 0800, coincident with the nadir of the endogenous corticosterone circadian rhythm. Mice were culled at 1200 and the kidneys snap frozen. NCC threonine phosphorylation (on T53; pT53‐NCC) was measured by western analysis as an index of transporter activation.Data are mean±SD. In control mice, acute corticosterone doubled pT53‐NCC abundance (p=0.004, Student's t test, n=6 in each group). In mice receiving chronic MR blockade (n=5), total NCC protein was significantly (P<0.001) reduced compared to controls (n=6); corticosterone still significantly increased pT53‐NCC abundance (1.0±0.46 AU (veh; n=5) vs 3.7±1.4 AU (cort; n=7), p=0.0018, Student's t test). Chronic GR blockade did not affect total NCC abundance (n=6 in each group) but prevented pT53‐NCC phosphorylation induced by acute cort injection (n=6 in each group).Our main conclusion are: 1) MR activation is required for maintenance of tonic levels of NCC protein; 2) glucocorticoids acutely increase NCC phosphorylation through GR activation. Direct regulation of NCC activity by glucocorticoids has implications for blood pressure control, particularly in individuals with inappropriately elevated glucocorticoids, such as in diabetes, obesity or during chronic stress.Support or Funding InformationWe thank The British Heart Foundation Centre of Research Excellence and Kidney Research UK for funding