Epithelial Na(+) Channel Expression Research Articles

Regulation of the epithelial Na+ channel by paraoxonase-2

Paraoxonase-2 (PON-2) is a membrane-bound lactonase with unique anti-oxidative and anti-atherosclerotic properties. PON-2 shares key structural elements with MEC-6, an endoplasmic reticulum-resident molecular chaperone in Caenorhabditis elegans MEC-6 modulates the expression of a mechanotransductive ion channel comprising MEC-4 and MEC-10 in touch-receptor neurons. Because pon-2 mRNA resides in multiple rat nephron segments, including the aldosterone-sensitive distal nephron where the epithelial Na+ channel (ENaC) is expressed, we hypothesized that PON-2 would similarly regulate ENaC expression. We observed PON-2 expression in aquaporin 2-positive principal cells of the distal nephron of adult human kidney. PON-2 also co-immunoprecipitated with ENaC when co-expressed in HEK293 cells. When PON-2 was co-expressed with ENaC in Xenopus oocytes, ENaC activity was reduced, reflecting a reduction in ENaC surface expression. MEC-6 also reduced ENaC activity when co-expressed in Xenopus oocytes. The PON-2 inhibitory effect was ENaC-specific, as PON-2 had no effect on functional expression of the renal outer medullary potassium channel. PON-2 did not alter the response of ENaC to extracellular Na+, mechanical shear stress, or α-chymotrypsin-mediated proteolysis, suggesting that PON-2 did not alter the regulation of ENaC by these factors. Together, our data suggest that PON-2 regulates ENaC activity by modulating its intracellular trafficking and surface expression.

Cyclic-AMP regulates postnatal development of neural and behavioral responses to NaCl in rats.

During postnatal development rats demonstrate an age-dependent increase in NaCl chorda tympani (CT) responses and the number of functional apical amiloride-sensitive epithelial Na+ channels (ENaCs) in salt sensing fungiform (FF) taste receptor cells (TRCs). Currently, the intracellular signals that regulate the postnatal development of salt taste have not been identified. We investigated the effect of cAMP, a downstream signal for arginine vasopressin (AVP) action, on the postnatal development of NaCl responses in 19–23 day old rats. ENaC-dependent NaCl CT responses were monitored after lingual application of 8-chlorophenylthio-cAMP (8-CPT-cAMP) under open-circuit conditions and under ±60 mV lingual voltage clamp. Behavioral responses were tested using 2 bottle/24h NaCl preference tests. The effect of [deamino-Cys1, D-Arg8]-vasopressin (dDAVP, a specific V2R agonist) was investigated on ENaC subunit trafficking in rat FF TRCs and on cAMP generation in cultured adult human FF taste cells (HBO cells). Our results show that in 19–23 day old rats, the ENaC-dependent maximum NaCl CT response was a saturating sigmoidal function of 8-CPT-cAMP concentration. 8-CPT-cAMP increased the voltage-sensitivity of the NaCl CT response and the apical Na+ response conductance. Intravenous injections of dDAVP increased ENaC expression and γ-ENaC trafficking from cytosolic compartment to the apical compartment in rat FF TRCs. In HBO cells dDAVP increased intracellular cAMP and cAMP increased trafficking of γ- and δ-ENaC from cytosolic compartment to the apical compartment 10 min post-cAMP treatment. Control 19–23 day old rats were indifferent to NaCl, but showed clear preference for appetitive NaCl concentrations after 8-CPT-cAMP treatment. Relative to adult rats, 14 day old rats demonstrated significantly less V2R antibody binding in circumvallate TRCs. We conclude that an age-dependent increase in V2R expression produces an AVP-induced incremental increase in cAMP that modulates the postnatal increase in TRC ENaC and the neural and behavioral responses to NaCl.

Derlin-1 promotes ubiquitylation and degradation of the epithelial Na+ channel, ENaC.

Ubiquitylation of the epithelial Na+ channel (ENaC) plays a critical role in cellular functions, including transmembrane transport of Na+, Na+ and water balance, and blood pressure stabilization. Published studies have suggested that ENaC subunits are targets of ER-related degradation (ERAD) in yeast systems. However, the molecular mechanism underlying proteasome-mediated degradation of ENaC subunits remains to be established. Derlin-1, an E3 ligase mediator, links recognized target proteins to ubiquitin-mediated proteasomal degradation in the cytosol. In the present study, we found that derlin-1 suppressed the expression of ENaC at the protein level and that the subunit α-ENaC (also known as SCNN1A) physically interacted with derlin-1 at the membrane-anchored domains or the loop regions, and that derlin-1 initiated α-ENaC retrotranslocation. In addition, HUWE1, an endoplasmic reticulum (ER)-resident E3 ubiquitin ligase, was recruited and promoted K11-linked polyubiquitylation of α-ENaC and, hence, formation of an α-ENaC ubiquitin-mediated degradation complex. These findings suggest that derlin-1 promotes ENaC ubiquitylation and enhances ENaC ubiquitin- mediated proteasome degradation. The derlin-1 pathway therefore may represent a significant early checkpoint in the recognition and degradation of ENaC in mammalian cells.

The Expression of Water and Ion Channels in Diffuse Alveolar Damage Is Not Dependent on DAD Etiology.

IntroductionAquaporins and ion channels are membrane proteins that facilitate the rapid movement of water and solutes across biological membranes. Experimental and in vitro studies reported that the function of these channels and pulmonary edema resolution are impaired in acute lung injury (ALI). Although current evidence indicates that alveolar fluid clearance is impaired in patients with ALI/diffuse alveolar damage (DAD), few human studies have addressed the alterations in pulmonary channels in this clinical condition. Additionally, it is not known whether the primary cause of DAD is a relevant variable for the channel dysfunction.MethodsAutopsied lungs of 43 patients with acute respiratory failure (ARF) due to DAD of three different etiologies, non-pulmonary sepsis, H1N1 viral infection and leptospirosis, were compared to 18 normal lungs. We quantified the expression of aquaporin (AQP) 1, AQP3, AQP5, epithelial Na+ channel (ENaC) and sodium potassium ATPase (Na-K-ATPase) in the alveolar septum using immunohistochemistry and image analysis.ResultsThe DAD group presented with increased expression of AQP3, AQP5 and Na-K-ATPase and decreased expression of ENaC compared to controls. However, there was no difference in protein expression within the DAD groups of different etiologies.ConclusionWater and ion channels are altered in patients with ARF due to DAD. The cause of DAD does not seem to influence the level of impairment of these channels.

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

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.

Disruption of KCNJ10 (Kir4.1) stimulates the expression of ENaC in the collecting duct.

Kcnj10 encodes the inwardly rectifying K(+) channel 4.1 (Kir4.1) and is expressed in the basolateral membrane of late thick ascending limb, distal convoluted tubule (DCT), connecting tubule (CNT), and cortical collecting duct (CCD). In the present study, we perform experiments in postneonatal day 9 Kcnj10(-/-) or wild-type mice to examine the role of Kir.4.1 in contributing to the basolateral K(+) conductance in the CNT and CCD, and to investigate whether the disruption of Kir4.1 upregulates the expression of the epithelial Na(+) channel (ENaC). Immunostaining shows that Kir4.1 is expressed in the basolateral membrane of CNT and CCD. Patch-clamp studies detect three types of K(+) channels (23, 40, and 60 pS) in the basolateral membrane of late CNT and initial CCD in wild-type (WT) mice. However, only 23- and 60-pS K(+) channels but not the 40-pS K(+) channel were detected in Kcnj10(-/-) mice, suggesting that Kir.4.1 is a key component of the 40-pS K(+) channel in the CNT/CCD. Moreover, the depletion of Kir.4.1 did not increase the probability of finding the 23- and 60-pS K(+) channel in the CNT/CCD. We next used the perforated whole cell recording to measure the K(+) reversal voltage in the CNT/CCD as an index of cell membrane potential. Under control conditions, the K(+) reversal potential was -69 mV in WT mice and -61 mV in Kcnj10(-/-) mice, suggesting that Kir4.1 partially participates in generating membrane potential in the CNT/CCD. Western blotting and immunostaining showed that the expression of ENaCβ and ENaCγ subunits from a renal medulla section of Kcnj10(-/-) mice was significantly increased compared with that of WT mice. Also, the disruption of Kir4.1 increased aquaporin 2 expression. We conclude that Kir4.1 is expressed in the CNT and CCD and partially participates in generating the cell membrane potential. Also, increased ENaC expression in medullary CD of Kcnj10(-/-) mice is a compensatory action in response to the impaired Na(+) transport in the DCT.

Is Salbutamol and Adrenalin Inhalation Effective in Management of Transient Tachypnia of Newborn?

Background: In the neonatal period, transient tachypnea of the newborn (TTN) is the most frequent cause of early respiratory distress because of delayed resorption of lung fluid, which fills the fetal airways. The inability of the lung to switch from fluid secretion to fluid absorption and immaturity in the expression of epithelial Nachannels (ENaC) may play an important role in the development of transient tachypnea of the newborn. Aims: Evaluation of inhalation therapy by BETA-2 agonist salbutamol and epinephrine for the management of transient tachypnea of newborn. Methods: The study was a prospective randomized control study. It included 60 neonates diagnosed clinically and radiologicaly as TTN. Inhaled salbutamol, epinephrine or saline 0.45% solution was administered. Results: Comparing studied groups after intervention, highly significant lower respiratory rate, TTN scores, respiratory support and duration of hospital stay was detected in group II (salbutamol group) while no significant difference between groups regarding heart rate after 4 hours from intervention. Original Research Article Nawar et al.; BJMMR, 14(3): 1-8, 2016; Article no.BJMMR.23922 2 Conclusion: Inhaled Salbutamol resulted in better outcome in decreasing the respiratory rate, TTN clinical score, lower FIO2, higher O2 saturation, decreasing the duration of respiratory support along with the total duration of hospitalization.

Epithelial Sodium Channels in Pulmonary Epithelial Progenitor and Stem Cells.

Regeneration of the epithelium of mammalian lungs is essential for restoring normal function following injury, and various cells and mechanisms contribute to this regeneration and repair. Club cells, bronchioalveolar stem cells (BASCs), and alveolar type II epithelial cells (ATII) are dominant stem/progenitor cells for maintaining epithelial turnover and repair. Epithelial Na+ channels (ENaC), a critical pathway for transapical salt and fluid transport, are expressed in lung epithelial progenitors, including club and ATII cells. Since ENaC activity and expression are development- and differentiation-dependent, apically located ENaC activity has therefore been used as a functional biomarker of lung injury repair. ENaC activity may be involved in the migration and differentiation of local and circulating stem/progenitor cells with diverse functions, eventually benefiting stem cells spreading to re-epithelialize injured lungs. This review summarizes the potential roles of ENaC expressed in native progenitor and stem cells in the development and regeneration of the respiratory epithelium.

Hypertension in NBCe2 Deficient Mice Caused by Enhanced ENaC Activity

Mutations of the electrogenic Na‐HCO3 cotransporter NBCe2 are associated with hypertension but the detailed pathogenesis is not well understood. Here we showed that up‐regulation of the epithelial Na channel (ENaC) in the distal nephron plays a critical role in hypertension caused by NBCe2 deficiency in mice. Wild type (WT) and NBCe2 knock out (KO) mice were fed an acid diet (1.5% NH4Cl) for 7‐10 days. Mean arterial pressure (MAP) was measured by tail cuff before and after the treatment with amiloride (Amil, 25 mg/L in drinking water). The MAP (mmHg) was higher in KO compared with WT before treatment (KO: 124.5 ± 2.2 vs WT: 110.6 ± 4.4). After treatment, the MAP in both groups dropped to a similar level (KO: 98.4 ± 3.7 vs WT: 99.1 ± 4.4). Western blot showed that the ENaC‐α expression in the kidney cortex was higher in KO compared with WT. Metabolic cage experiments showed that urinary Na excretion increased 3‐fold after Amil treatment (5 mg/kg, ip) in KO, while increasing only 20% in WT. The urinary K excretion also decreased significantly after Amil treatment in the KO but was similar before and after treatment in WT. Micropunture experiments showed that the [K] was significantly higher in the connecting tubule (CNT) of the KO compared with WT (KO: 21.9 ± 0.6 vs WT: 17.9 ± 1.5 mM), while the transepithelial potential (Vte, mV) was significantly lower (KO: ‐3.2 ± 0.2 vs WT: ‐1.6 ± 0.2). After Amil treatment, the Vte increased to ‐0.1 ± 0.2 in KO and 0.7 ± 0.4 in WT. These results show that both the ENaC expression and activity are increased in the KO, which contribute to the hypertension caused by NBCe2 deficiency.

Improvement of nephrotic syndrome by Banggihwanggitang.

Nephrotic syndrome is associated with altered renal handling of sodium along with changes of renal epithelial Na channel (ENaC), leading to the development of edema and ascites.The dried sclerotia of Banggihwanggitang (BHT) has been used for the treatment of chronic kidney disease. This study was conducted to evaluate the effects of BHT on puromycin aminonucleoside (PAN)‐induced renal functional derangement and the change of ENaC expression. The nephrotic syndrome animal models were constructed by PAN 75 mg/kg injection and then were treated with losartan (30 mg/kg/day) or BHT (200 mg/kg/day) for 7 days. Consequently, BHT group was significantly improved the proteinuria and ascites. In addition, the plasma level of triglyceride, total cholesterol, and low density lipoprotein (LDL)‐cholesterol were significantly decreased in BHT. Besides, BHT group attenuated the PAN‐induced increase in ENaC protein expression. BHT was significantly suppressed PAN‐induced organic osmolytes regulator such as serum‐ and glucocorticoid‐inducible protein kinase (Sgk1), and mineralocorticoid receptor (MR) mRNA expression.Taken together, BHT improve nephrotic syndrome including proteinuria and ascites, through inhibition of ENaC. Thus, Banggihwanggitang is involved in the body‐fluid regulation via inhibition of water and sodium channels against renal disorder such as edema or nephrosis.

Epigenetics of epithelial Na+channel-dependent sodium uptake and blood pressure regulation

The epithelial Na(+) channel (ENaC) consists of α, β, γ subunits. Its expression and function are regulated by aldosterone at multiple levels including transcription. ENaC plays a key role in Na(+) homeostasis and blood pressure. Mutations in ENaC subunit genes result in hypertension or hypotension, depending on the nature of the mutations. Transcription of αENaC is considered as the rate-limiting step in the formation of functional ENaC. As an aldosterone target gene, αENaC is activated upon aldosterone- mineralocorticoid receptor binding to the cis-elements in the αENaC promoter, which is packed into chromatin. However, how aldosterone alters chromatin structure to induce changes in transcription is poorly understood. Studies by others and us suggest that Dot1a-Af9 complex represses αENaC by directly binding and regulating targeted histone H3 K79 hypermethylation at the specific subregions of αENaC promoter. Aldosterone decreases Dot1a-Af9 formation by impairing expression of Dot1a and Af9 and by inducing Sgk1, which, in turn, phosphorylates Af9 at S435 to weaken Dot1a-Af9 interaction. MR attenuates Dot1a-Af9 effect by competing with Dot1a for binding Af9. Af17 relieves repression by interfering with Dot1a-Af9 interaction and promoting Dot1a nuclear export. Af17(-/-) mice exhibit defects in ENaC expression, renal Na(+) retention, and blood pressure control. This review gives a brief summary of these novel findings.

Proteases, ENaCs and cystic fibrosis.

Proteases, ENaCs and cystic fibrosis.

Impaired epithelial Na+ channel activity contributes to cystogenesis and development of autosomal recessive polycystic kidney disease in PCK rats.

BACKGROUNDAutosomal recessive polycystic kidney disease (ARPKD) is a genetic disorder characterized by the development of renal cysts of the tubular epithelial cell origin. Epithelial Na+ channel (ENaC) is responsible for sodium reabsorption in the aldosterone-sensitive distal nephron. Here we investigated the ENaC expression and activity in cystic tissue taken from rats with ARPKD.METHODSPCK rats were treated with the selective ENaC inhibitor benzamil given in the drinking water, and after 4 or 12 weeks the severity of morphological malformations in the kidneys was assessed. ENaC and AQP2 expression and ENaC activity were tested with immunohistochemistry and patch clamp electrophysiology, respectively.RESULTSTreatment with benzamil exacerbated development of cysts compared to the vehicle-treated animals. In contrast, the 12 weeks treatment with a loop diuretic furosemide had no effect on cystogenesis. Single channel patch clamp analysis revealed that ENaC activity in the freshly isolated cystic epithelium was significantly lower than in the non-cystic collecting ducts isolated from PCK or normal Sprague-Dawley rats. Immunohistochemical analysis confirmed that β-ENaC and AQP2 expressions in cysts are decreased compared to non-dilated tubules from PCK rat kidneys.CONCLUSIONWe demonstrated that cystic epithelium exhibits low ENaC activity and this phenomenon can contribute to cyst progression.

Collecting duct-specific knockout of renin attenuates angiotensin II-induced hypertension.

The physiological and pathophysiological significance of collecting duct (CD)-derived renin, particularly as it relates to blood pressure (BP) regulation, is unknown. To address this question, we generated CD-specific renin knockout (KO) mice and examined BP and renal salt and water excretion. Mice containing loxP-flanked exon 1 of the renin gene were crossed with mice transgenic for aquaporin-2-Cre recombinase to achieve CD-specific renin KO. Compared with controls, CD renin KO mice had 70% lower medullary renin mRNA and 90% lower renin mRNA in microdissected cortical CD. Urinary renin levels were significantly lower in KO mice (45% of control levels) while plasma renin concentration was significantly higher in KO mice (63% higher than controls) during normal-Na intake. While no observable differences were noted in BP between the two groups with varying Na intake, infusion of angiotensin II at 400 ng·kg(-1)·min(-1) resulted in an attenuated hypertensive response in the KO mice (mean arterial pressure 111 ± 4 mmHg in KO vs. 128 ± 3 mmHg in controls). Urinary renin excretion and epithelial Na(+) channel (ENaC) remained significantly lower in the KO mice following ANG II infusion compared with controls. Furthermore, membrane-associated ENaC protein levels were significantly lower in KO mice following ANG II infusion. These findings suggest that CD renin modulates BP in ANG II-infused hypertension and these effects are associated with changes in ENaC expression.

ENaC activity and expression is decreased in the lungs of protein kinase C-α knockout mice.

We used a PKC-α knockout model to investigate the regulation of alveolar epithelial Na(+) channels (ENaC) by PKC. Primary alveolar type II (ATII) cells were subjected to cell-attached patch clamp. In the absence of PKC-α, the open probability (Po) of ENaC was decreased by half compared with wild-type mice. The channel density (N) was also reduced in the knockout mice. Using in vivo biotinylation, membrane localization of all three ENaC subunits (α, β, and γ) was decreased in the PKC-α knockout lung, compared with the wild-type. Confocal microscopy of lung slices showed elevated levels of reactive oxygen species (ROS) in the lungs of the PKC-α knockout mice vs. the wild-type. High levels of ROS in the knockout lung can be explained by a decrease in both cytosolic and mitochondrial superoxide dismutase activity. Elevated levels of ROS in the knockout lung activates PKC-δ and leads to reduced dephosphorylation of ERK1/2 by MAP kinase phosphatase, which in turn causes increased internalization of ENaC via ubiquitination by the ubiquitin-ligase Nedd4-2. In addition, in the knockout lung, PKC-δ activates ERK, causing a decrease in ENaC density at the apical alveolar membrane. PKC-δ also phosphorylates MARCKS, leading to a decrease in ENaC Po. The effects of ROS and PKC-δ were confirmed with patch-clamp experiments on isolated ATII cells in which the ROS scavenger, Tempol, or a PKC-δ-specific inhibitor added to patches reversed the observed decrease in ENaC apical channel density and Po. These results explain the decrease in ENaC activity in PKC-α knockout lung.

Developmental changes of ENaC expression and function in the inner ear of pendrin knock-out mice as a perspective on the development of endolymphatic hydrops.

Pendrin mutations cause enlarged vestibular aqueducts and various degrees of sensorineural hearing loss. The selective abolition of pendrin causes dilation of the membranous labyrinth known as endolymphatic hydrops, loss of the endocochlear potential, and consequently loss of hearing function. Because Na+ transport is one of the most important driving forces for fluid transport, the epithelial Na+ channel (ENaC) is believed to play an important role in fluid volume regulation in the inner ear. Therefore, the dysfunction of Na+ transport through ENaC by the acidification of endolymph in Pendred syndrome is one of the potential causes of endolymphatic hydrops. We investigated the changes of ENaC expression and function during the development of the pendrin knock-out mouse. In the cochlea, the expression of β and γENaC was significantly increased at P56 in Pds−/− mice compared with Pds+/+ mice. In the vestibule, the expression of βENaC was significantly increased at P56, and γENaC expression significantly increased from P6 to P56 in Pds−/− mice. The ENaC-dependent trans-epithelial current was not significantly different between Pds+/+ and Pds−/− mice in Reissner’s membrane or the saccular extramacular roof epithelium at P0, but the current was significantly increased in Pds−/− mice at P56 compared with Pds+/+ mice. These findings indicate that the expression and function of ENaC were enhanced in Pds−/− mice after the development of endolymphatic hydrops as a compensatory mechanism. This result provides insight into the role of Na+ transport in the development and regulation of endolymphatic hydrops due to pendrin mutations.

Improvement of body‐fluid dysfunction by Poria cocos in nephrotic syndrome (701.3)

Improvement of body‐fluid dysfunction by Poria cocos in nephrotic syndromeSo Min Lee 1,2, Yun Jung Lee1,2, Jung Joo Yoon1,2, Byung Hyuk Han1,2, Dae Gill Kang1,2, and Ho Sub Lee1,21College of Oriental Medicine and Professional Graduate School of Oriental Medicine, Wonkwang University, Iksan, Jeonbuk 570‐749, Republic of Korea; 2 Hanbang Body‐fluid Research Center, Wonkwang University, Shinyong‐dong, Iksan, Jeonbuk, 570‐749, Republic of KoreaAbstract Nephrotic syndrome is associated with altered renal handling of water and sodium along with changes of aquaporins (AQP) and renal epithelial Na channel (ENaC).The dried sclerotia of Poria cocos Wolf (WPC) has been used for the treatment of chronic edema and nephrosis. This study was conducted to evaluate the effects of WPC on puromycin aminonucleoside (PAN)‐induced renal functional derangement and the change of renal AQP2 and ENaC expression. The nephrotic syndrome animal models were constructed by PAN 75 mg/kg injection and then were treated with losartan (30 mg/kg/day) or WPC (200 mg/kg/day) for 7 days. Consequently, WPC group was significantly improved the proteinuria and ascites. In addition, the plasma level of triglyceride, total cholesterol, and low density lipoprotein (LDL)‐cholesterol were significantly decreased in WPC. Besides, WPC group attenuated the PAN‐induced increase in protein, and mRNA levels of AQP2 and ENaC subunit α/β. WPC was significantly suppressed PAN‐induced organic osmolytes regulator such as serum‐ and glucocorticoid‐inducible protein kinase (Sgk1), and sodium‐myo‐inositol cotransporter (SMIT) mRNA expression.Taken together, WPC improve nephrotic syndrome including proteinuria and ascites, through inhibition of AQP2 and ENaC subunit. Thus, WPC is involved in the body‐fluid regulation via inhibition of water and sodium channels against renal disorder such as edema or nephrosis.

The epithelial Na+ channel

Approximately 20 years ago, a paradigm shift occurred questioning whether expression of the epithelial Na⁺ channel (ENaC) was mainly restricted to epithelial tissues. In this review, the recent findings of ENaC regulation, and its potential contributions to the function and dysfunction of the vasculature, is discussed. Over the last few years, the expression, localization, and functional properties of ENaC have been determined in the two main vascular cell types: endothelial cells, and vascular smooth muscle cells. A chronically increased ENaC membrane abundance can lead to endothelial stiffening and to a reduced release of nitric oxide, the hallmark of endothelial dysfunction. Endothelial ENaC was shown to determine vasoconstriction by negatively modulating nitric oxide release in mesenteric arteries, likely via the PI3K/Akt signaling pathway. ENaC has therefore been recognized as a potentially important regulator of vascular nanomechanics and as a transducer of mechanical forces. As ENaC expression is broader than anticipated, it has become clear that the protein may play a crucial role in the vasculature as it is located at the interface between blood and tissue, and is therefore implicated in the development of endothelial dysfunction and hypertension.

Effect of Poria cocos on Puromycin Aminonucleoside-Induced Nephrotic Syndrome in Rats.

Nephrotic syndrome is associated with altered renal handling of water and sodium and changes in the levels of aquaporins (AQPs) and epithelial Na channels (ENaCs). The dried sclerotia of Poria cocos Wolf (WPC) have been used for treating chronic edema and nephrosis. We evaluated the effects of WPC on puromycin aminonucleoside- (PAN-) induced renal functional derangement and altered renal AQP2 and ENaC expression. In the nephrotic syndrome rat model, animals were injected with 75 mg/kg PAN and then treated with Losartan (30 mg·kg−1 ·day−1) or WPC (200 mg·kg−1 ·day−1) for 7 days. In the WPC group, proteinuria and ascites improved significantly. Plasma levels of triglyceride, total cholesterol, and low-density lipoprotein- (LDL-) cholesterol reduced significantly in the WPC group. In addition, the WPC group exhibited attenuation of the PAN-induced increase in AQP2 and ENaC α/β subunit protein and mRNA levels. WPC suppressed significantly PAN-induced organic osmolyte regulators, reducing serum- and glucocorticoid-inducible protein kinase (Sgk1) and sodium-myo-inositol cotransporter (SMIT) mRNA expression. Our results show that WPC improves nephrotic syndrome, including proteinuria and ascites, through inhibition of AQP2 and ENaC expression. Therefore, WPC influences body-fluid regulation via inhibition of water and sodium channels, thereby, improving renal disorders such as edema or nephrosis.

Cycloheximide and lipopolysaccharide downregulate αENaC mRNA via different mechanisms in alveolar epithelial cells

Active Na(+) transport mediated by epithelial Na(+) channel (ENaC) is vital for fetal lung fluid reabsorption at birth and pulmonary edema resolution. Previously, we demonstrated that αENaC expression and activity are downregulated in alveolar epithelial cells by cycloheximide (Chx) and Pseudomonas aeruginosa. The regulatory mechanisms of αENaC mRNA expression by Chx and lipopolysaccharide (LPS) from P. aeruginosa were further studied in the present work. Both agents decreased αENaC mRNA expression to 50% of control values after 4 h. Chx repressed αENaC expression in a dose-dependent manner independently of protein synthesis. Although extracellular signal-regulated kinases 1 and 2 (ERK1/2) and p38 mitogen-activated protein kinase (MAPK) pathways were activated by the two treatments, their mechanisms of ENaC mRNA modulation were different. First, activation of the signaling pathways was sustained by Chx but only transiently by LPS. Second, ERK1/2 or p38 MAPK inhibition attenuated the effects of Chx on αENaC mRNA, whereas suppression of both signaling pathways was necessary to alleviate the outcome of LPS on αENaC mRNA. The molecular mechanisms involved in the decrease of αENaC expression were investigated in both conditions. LPS, but not Chx, significantly reduced αENaC promoter activity via the ERK1/2 and p38 MAPK pathways. These results suggest that LPS attenuates αENaC mRNA expression via diminution of transcription, whereas Chx could trigger some posttranscriptional mechanisms. Although LPS and Chx downregulate αENaC mRNA expression similarly and with similar signaling pathways, the mechanisms modulating ENaC expression are different depending on the nature of the cellular stress.