Electrolyte Homeostasis Research Articles

Mechanistic target of rapamycin: integrating growth factor and nutrient signaling in the collecting duct.

The renal collecting duct and other postmacula densa sites are the primary tubular regions for fine-tuning of electrolyte homeostasis in the body. A role for the mechanistic target of rapamycin (mTOR), a serine-threonine kinase, has recently been appreciated in this regulation. mTOR exists in two distinct multiprotein functional complexes, i.e., mTORC1 and mTORC2. Upregulation of mTORC1, by growth factors and amino acids, is associated with cell cycle regulation and hypertrophic changes. In contrast, mTORC2 has been demonstrated to have a role in regulating Na+ and K+ reabsorptive processes, including those downstream of insulin and serum- and glucocorticoid-regulated kinase (SGK). In addition, mTORC2 can upregulate mTORC1. A number of elegant in vitro and in vivo studies using cell systems and genetically modified mice have revealed mechanisms underlying activation of the epithelial Na+ channel (ENaC) and the renal outer medullary K+ channel (ROMK) by mTORC2. Overall, mTOR in its systematic integration of phosphorylative signaling facilitates the delicate balance of whole body electrolyte homeostasis in the face of changes in metabolic status. Thus, inappropriate regulation of renal mTOR has the potential to result in electrolyte disturbances, such as acidosis/alkalosis, hyponatremia, and hypertension. The goal of this minireview is to highlight the physiological role of mTOR in its complexes in regulating electrolyte homeostasis in the aldosterone-sensitive distal nephron.

The depressor axis of the renin-angiotensin system and brain disorders: a translational approach.

All the components of the classic renin-angiotensin system (RAS) have been identified in the brain. Today, the RAS is considered to be composed mainly of two axes: the pressor axis, represented by angiotensin (Ang) II/angiotensin-converting enzyme/AT1 receptors, and the depressor and protective one, represented by Ang-(1-7)/ angiotensin-converting enzyme 2/Mas receptors. Although the RAS exerts a pivotal role on electrolyte homeostasis and blood pressure regulation, their components are also implicated in higher brain functions, including cognition, memory, anxiety and depression, and several neurological disorders. Overactivity of the pressor axis of the RAS has been implicated in stroke and several brain disorders, such as cognitive impairment, dementia, and Alzheimer or Parkinson's disease. The present review is focused on the role of the protective axis of the RAS in brain disorders beyond its effects on blood pressure regulation. Furthermore, the use of drugs targeting centrally RAS and its beneficial effects on brain disorders are also discussed.

Effects of Acrolein on Adrenal Glands

Aldosterone and cortisol, two major hormones secreted from adrenal glands, are vital to life by maintaining electrolyte, body fluid and energy homeostasis in human. Both of hormones are regulated precisely. Acrolein, the well-known cytotoxic agent released during the incomplete combustion. It abounds in the smoke of cigarette and in enclosed fires. Previous studies have demonstrated that smoking has the adverse effects on cardiovascular, nerve and other system. In particular, the pathogenic functions of tobacco inhalation in cardiovascular disorders as well as alters plasma electrolyte levels have been well documented. It is thus important to evaluate cellular/molecular mechanisms of smoking that change these responses. In a recent paper published in Steroids , we demonstrated that the major chemical in cigarette, acrolein, has the abilities to enhance aldosterone secretion and decrease corticosterone secretion. These effects may associate with the progression of cardiovascular and other diseases.

Preamble to “Human Physiology: Water and Electrolyte Homeostasis” by Yu. Natochin

Preamble to “Human Physiology: Water and Electrolyte Homeostasis” by Yu. Natochin

Incorporation of the δ-subunit into the epithelial sodium channel (ENaC) generates protease-resistant ENaCs in Xenopus laevis

The epithelial sodium channel (ENaC) is a critical regulator of vertebrate electrolyte homeostasis. ENaC is the only constitutively open ion channel in the degenerin/ENaC protein family, and its expression, membrane abundance, and open probability therefore are tightly controlled. The canonical ENaC is composed of three subunits (α, β, and γ), but a fourth δ-subunit may replace α and form atypical δβγ-ENaCs. Using Xenopus laevis as a model, here we found that mRNAs of the α- and δ-subunits are differentially expressed in different tissues and that δ-ENaC predominantly is present in the urogenital tract. Using whole-cell and single-channel electrophysiology of oocytes expressing Xenopus αβγ- or δβγ-ENaC, we demonstrate that the presence of the δ-subunit enhances the amount of current generated by ENaC due to an increased open probability, but also changes current into a transient form. Activity of canonical ENaCs is critically dependent on proteolytic processing of the α- and γ-subunits, and immunoblotting with epitope-tagged ENaC subunits indicated that, unlike α-ENaC, the δ-subunit does not undergo proteolytic maturation by the endogenous protease furin. Furthermore, currents generated by δβγ-ENaC were insensitive to activation by extracellular chymotrypsin, and presence of the δ-subunit prevented cleavage of γ-ENaC at the cell surface. Our findings suggest that subunit composition constitutes an additional level of ENaC regulation, and we propose that the Xenopus δ-ENaC subunit represents a functional example that demonstrates the importance of proteolytic maturation during ENaC evolution.

Beneficial Effects of High Potassium: Contribution of Renal Basolateral K+ Channels.

The prevalence of hypertension in the United States is estimated to have reached 85.7 million adults1 without taking into account new American College of Cardiology/American Heart Association guideline for the prevention, detection, evaluation, and management of high blood pressure,2 which is certain to increase this estimate. Hypertension is a major risk factor for stroke, coronary heart disease, and renal failure. Additionally, high blood pressure (BP) increases cardiovascular mortality rates and progression of many other chronic diseases. Salt and water handling by the kidney directly affects BP, whereas renal ion channels and transporters maintain electrolyte homeostasis.3,4 Epidemiological and anthropological studies suggest that in isolated societies with diets composed primarily of fruits, vegetables, and nuts, hypertension affects only 1% of the population. In contrast, in countries consuming a modern Western diet high in processed foods and dietary sodium, the prevalence of hypertension is as high as 30%. This effect of the Western diet is not solely attributed to high sodium content but rather the dramatically decreased dietary potassium-to-sodium ratio. In industrialized countries, the daily intakes of potassium and sodium are ≈30 to 50 and 80 to 250 mmol per day, respectively. This is in sharp contrast with isolated or primitive societies, having the daily rates of 150 to 290 mmol for potassium and 20 to 40 mmol per day for sodium.5 Therefore, estimated potassium-to-sodium intake ratios range from 0.12 to 0.63 for industrialized societies and 3.8 to 14.5 for isolated societies. According to the National Health and Nutrition Examination Survey, only about one tenth of US adults have potassium-to-sodium intake ratios consistent with the World Health Organization guidelines for reduced risk of mortality.6 Potassium is the most abundant intracellular ion, and its role in the regulation of BP is well established. Dietary supplementation with potassium …

Study of association of mortality with electrolyte abnormalities in children admitted in pediatric intensive care unit

Background: Electrolyte abnormalities are common in critically ill children. In view of importance of electrolyte homeostasis and its significant impact on the final outcome of patient, the present study was undertaken. Objective of this study was to determine the prevalence of electrolyte abnormalities in children admitted in Pediatric Intensive Care Unit at the time of admission and its association with mortality and primary organ system involvement.Methods: The study enrolled all the patients from 29 days to 12 years admitted in PICU of a tertiary care hospital during April 2015 to September 2016 (total 18 months). The children were classified according to presence or absence of electrolyte abnormality. The children were further divided into subgroups based on electrolyte values and mortality and organ system involvement was analyzed in each of the sub groups.Results: The prevalence of electrolyte abnormality in terms of sodium or potassium abnormality in the present study was 44.31% (323 of 729). Hyponatremia (27.43%) was the most common electrolyte abnormality followed by hypokalemia (13.99%). The mortality in children with electrolyte abnormality was found to be 28.8% which was significantly higher than mortality in those without electrolyte abnormality. Maximum children with hyponatremia had central nervous system involvement (48.5%) and those with hypernatremia had gastrointestinal involvement (65.4%). Hypokalemia was most frequently observed with gastrointestinal involvement (54.9%) and hyperkalemia with renal involvement (34.8%).Conclusions: Present study has demonstrated significant association of electrolyte abnormalities at admission in PICU with mortality and primary organ system involvement. Close monitoring and correction of electrolyte abnormalities is essential to reduce morbidity and mortality.

Standard of care, controversies, and innovations in the medical treatment of severe traumatic brain injury.

Severe traumatic brain injury (STBI) is characterized by a primary injury which cannot be reversed and a secondary injury that can be prevented or reversed. Management of STBI patients in intensive care mainly aims at preventing the secondary injury. Treatment aims to: reducing ICP pressure (that can result in an ischemic insult); avoiding hypotension, hyperthermia, or hypoxemia; maintaining a normal electrolytes homeostasis; treating the autonomic dysfunction syndrome, coagulopathies, acute kidney injury and maintaining an adequate nutrition. Many treatment protocols are already well established, while many others are still debated. Moreover, new frontiers in STBI management are represented by the neurovascular regeneration and neurorestoration which are showing very promising results even if most of them still need a clinical validation. In this paper we review standard of care, controversies and innovations in the medical treatment of STBI.

The mineralocorticoid receptor as a modulator of innate immunity and atherosclerosis.

The mineralocorticoid receptor (MR) is a member of the nuclear receptor steroid-binding family. The classical MR ligand aldosterone controls electrolyte and fluid homeostasis after binding in renal epithelial cells. However, more recent evidence suggests that activation of extrarenal MRs by aldosterone negatively impacts cardiovascular health independent of its effects on blood pressure: high levels of aldosterone associate with an increased cardiovascular event rate, where MR antagonists exert beneficial effects on cardiovascular mortality. The most important cause for cardiovascular events is atherosclerosis that is currently considered a low-grade inflammatory disorder of the arterial wall. In this inflammatory process, the innate immune system plays a deciding role, with the monocyte-derived macrophage being the most abundant cell in the atherosclerotic plaque. Intriguingly, both monocytes and macrophages express the MR, and a growing body of evidence shows that these cells are skewed into a pro-inflammatory and pro-atherosclerotic phenotype via MR stimulation. In this review, we detail the current perspective on the role of the monocyte and macrophage MR in atherosclerosis development and provide a comprehensive framework of the effects of MR activation of the innate immune system that might drive the pro-atherosclerotic outcome.

Using a Physiological Model to Understand Water and Electrolyte Disturbances Following Transsphenoidal Pituitary Surgery

Disturbances in water and electrolyte homeostasis are common following transsphenoidal surgery. These disturbances are variable and unpredictable, increasing patient risk and complicating postsurgical treatment decisions. Clinically, it is generally accepted that damage to the pituitary stalk is the source of these disturbances, but the mechanisms behind the response variability and underlying pathophysiology remains unknown. The objective was to test the hypothesis that stepwise damage to the posterior pituitary produces a spectrum of water and electrolyte disturbance which can account for the full variety of disease presentation. We used HumMod, a large mathematical model of human physiology, to simulate pituitary stalk damage at differing fractions: 20, 40, 60, and 80%. The damaged neurons were modeled to undergo a 5‐day countdown to degeneration and release their stored ADH as they die, as is described in the literature. Pituitary damage changed the balance of water and electrolytes to various degrees. Lower damage (20%) resulted in transient polyuria, while intermediate damage (40%) was associated with a delayed polyuria and diabetes insipidus. Higher levels of damage (60 and 80%) resembled triphasic diabetes insipidus. We postulate that the results from this model can serve as a mechanistic basis for many varieties of postsurgical water and electrolyte disturbances, where increasing cellular damage to the pituitary potentiates the likelihood of a full triphasic response. However, our simulation suggests that water and electrolyte disturbances following pituitary surgery cannot be grouped into a spectrum of disease that depends only on the level of pituitary damage. Other mechanisms which are still unclear and not a part of this model are likely at play.Support or Funding InformationThis work is supported by the Hearin Foundation through the Medical Student Research Program as well as T32 HL105324.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Slc4a8 in the Kidney: Expression, Subcellular Localization and Role in Acid Base Homeostasis and Salt Reabsorption

BackgroundThe sodium‐dependent bicarbonate transporter Slc4a8 (a.k.a NDCBE) mediates the co‐transport of sodium and bicarbonate in exchange for chloride. It is abundantly detected in the brain, with low expression levels in the kidney. The cell distribution and subcellular localization of Slc4a8 in the kidney and its role in acid/base and electrolyte homeostasis has been the subject of conflicting reports. There are no conclusive localization or functional studies to pinpoint the location and demonstrate the function of Slc4a8 in the kidney.MethodsMolecular techniques, including in situ hybridization and RT‐PCR, were performed on kidney sections and tagged epitopes were used to examine the membrane targeting of Slc4a8 in polarized kidney cells. Crispr/Cas9 was used to generate and examine Slc4a8 KO mice.ResultIn situ hybridization and Zonal distribution studies showed no expression for Slc4a8 (NDCBE) in the cortex or in cortical collecting ducts (CCD). Slc4a8 was predominantly detected in the outer and inner medullary collecting ducts (OMCD and IMCD), and was targeted to the basolateral membrane of osmotically tolerant MDCK cells when confocal microscopy images were acquired. Slc4a8 KO mice did not show any abnormal salt or bicarbonate wasting under baseline conditions or in response to bicarbonate loading, salt restriction or furosemide‐induced diuresis.ConclusionsSlc4a8 (NDCBE) is absent in the CCD and is predominantly localized on the basolateral membrane of medullary collecting duct cells. Further, Slc4a8 deletion does not cause any acid base or electrolyte abnormality in pathophysiologic states such as furosemide‐induced salt wasting or salt restriction. We suggest that Slc4a8 is likely involved in intracellular pH and volume regulation in medullary collecting duct cells.Support or Funding InformationMerit Review 5 I01 BX001000‐06 award from the Department of Veterans Affairs, and funds from the Center on Genetics of Transport and Epithelial Biology at the University of Cincinnati.

Migraines and Ionic Variances

Chronic migraine is considered to be a disabling neurological disease, treated with dangerous and often brain damaging medicines. A good summary of recent findings labels migraine as “an inherited, episodic disorder involving sensory sensitivity”9. While crucial findings like this have been made, and even with the medical community's understanding and acceptance of the previous section's genetic explanation, research of dietary factors is still missing. Why? There are several reasons but one of the fundamental ones is the belief that migraine is a disease.Research showed in 1951 that migraineurs excrete 50% more sodium in their urine than non‐migraineurs do and have “busy brains”. These findings suggest an important clue for the mechanism of migraines even without the understanding of the genetic connection. This clue led me to the recognition of a promising hypothesis, namely, the possibility that in some manner sodium depletion is involved in migraine onset and persistence. As far as I could ascertain, the question of “Why sodium is depleted?” has not been asked—though some resent research found support for an inverse relationship between migraines and the amount of dietary sodium consumed: the more sodium was consumed, the fewer migraines appeared.Migraineurs are much more likely to have metabolic disorders than non‐migraineurs. This is likely connected to electrolyte disturbance caused by exogenous glucose. The extreme response is caused by genetic variations of glucose transporters, ionic channels that manage electrolyte homeostasis, and associated proteins. Migraineurs appear to genetically be predisposed to other forms of metabolic diseases as well.Some of the most critical genetic variances that affect migraineurs are as follows (GeneCards database): CACNA1A Calcium voltage gated channel ATP1A2 ATPase, Na/K transporting SCN1A Sodium voltage gated channel NOTCH3 intercellular signaling pathway that plays a key role in neural development PRRT2 Proline Rich Transmembrane Protein 2; α‐amino acid that is used in the biosynthesis of proteins; potential endogenous excitotoxin KCNK18 potassium channel EDNRA Endothelin Receptor Type A activation result in elevation of intracellular‐free calcium SLC2A1 Solute Carrier Family 2 Member 1 facilitative transporters (allow solutes to flow downhill with their electrochemical gradients); secondary active transporters (allow solutes to flow uphill against their electrochemical gradient by coupling to transport of a second solute that flows downhill with its gradient such that the overall free energy change is still favorable) MTHFR Methylation of folate SLCxxx just about all solute carriers Together, these hint at major electrolyte dysregulation potential if proper nutrition is not provided. Migraine appears to share much in common with channelopathy.More research is needed to specifically examine the electrolyte dysfunction caused by exogenous glucose consumption in migraineurs and the associated CSD and migraine pain.Support or Funding InformationThe author received no funding or support. There is no conflict of interest.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The Protective Effects of Atrial Natriuretic Peptide Infusion in Salt‐Sensitive Hypertension

Atrial Natriuretic Peptide (ANP) is a hormone secreted by cardiomyocytes in response to increased atrial volume; it stimulates water and sodium excretion by the kidneys and relieves pressure on the circulatory system. In our previous studies we compared wild type Dahl Salt Sensitive (SS) rats to SS rats lacking the Nppa gene which encodes ANP (SSNppa−/−). The data revealed that salt‐induced blood pressure development was significantly exacerbated in the SSNppa−/− rats. Furthermore, SSNppa−/− rats exhibited reduced urinary sodium excretion and diuresis, as well as aggravated kidney damage compared to wild type controls (FASEB J 2017 31:855.8.). These data led to a hypothesis that ANP infusion can have beneficial effects during the development of salt‐induced hypertension.Dahl SS rats were fed a high salt diet (HS, 4% NaCl) for 21 days, with a continuous i.v. infusion of ANP (100 ng/kg/day) administered from day 0 or day 14 onward to test preventive and therapeutic effects of ANP, respectively. Blood pressure was monitored throughout the study, and urine samples were collected in metabolic cages on days 0 and 21. At the end of the protocol animals were sacrificed and kidney tissues were harvested and stained with Masson's trichrome to evaluate renal damage. Fibrosis, glomerular injury, and protein casts formation were assessed; urine and plasma samples were also analyzed to estimate electrolyte homeostasis.ANP infusion had beneficial effects on BP and renal function compared to control animals. We observed a decrease in blood pressure in both groups infused with ANP during 0–21 days (21D) and 14–21 days (7D) of a HS challenge. Kidney to body weight ratio did not differ between the groups; however, we observed a trend for reduced renal hypertrophy in the 21D animals. Metabolic cage studies revealed no changes in diuresis; urinary electrolyte analysis showed a more effective sodium excretion in the 21D rats. Assessment of kidney damage demonstrated a decrease in protein casts in both 7D and 21D rats: 3.8% ± 0.47%, 2.7% ± 0.24%, and 2.0% ± 0.41% of the tissue surface in the control, 7D and 21D groups, respectively. Glomerular injury had a similar pattern: scores were 2.86 ± 0.09, 2.62 ± 0.07, and 2.41 ± 0.06. Renal fibrosis was alleviated in the 21D group (1.4% ± 0.18%, compared to 1.9% ± 0.49% and 3.1% ± 0.31% in the 7D and control groups).Therefore, ANP infusion had both preventive and therapeutic effects. Our data revealed that ANP administration during the HS diet challenge resulted in decreased blood pressure, increased electrolyte excretion, and reduced tissue damage.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Intracellular Chloride and Scaffold Protein Mo25 Cooperatively Regulate Transepithelial Ion Transport through WNK Signaling in the Malpighian Tubule

Background With No Lysine kinase (WNK) signaling regulates mammalian renal epithelial ion transport to maintain electrolyte and BP homeostasis. Our previous studies showed a conserved role for WNK in the regulation of transepithelial ion transport in the Drosophila Malpighian tubule.Methods Using in vitro assays and transgenic Drosophila lines, we examined two potential WNK regulators, chloride ion and the scaffold protein mouse protein 25 (Mo25), in the stimulation of transepithelial ion flux.ResultsIn vitro, autophosphorylation of purified Drosophila WNK decreased as chloride concentration increased. In conditions in which tubule intracellular chloride concentration decreased from 30 to 15 mM as measured using a transgenic sensor, Drosophila WNK activity acutely increased. Drosophila WNK activity in tubules also increased or decreased when bath potassium concentration decreased or increased, respectively. However, a mutation that reduces chloride sensitivity of Drosophila WNK failed to alter transepithelial ion transport in 30 mM chloride. We, therefore, examined a role for Mo25. In in vitro kinase assays, Drosophila Mo25 enhanced the activity of the Drosophila WNK downstream kinase Fray, the fly homolog of mammalian Ste20-related proline/alanine-rich kinase (SPAK), and oxidative stress-responsive 1 protein (OSR1). Knockdown of Drosophila Mo25 in the Malpighian tubule decreased transepithelial ion flux under stimulated but not basal conditions. Finally, whereas overexpression of wild-type Drosophila WNK, with or without Drosophila Mo25, did not affect transepithelial ion transport, Drosophila Mo25 overexpressed with chloride-insensitive Drosophila WNK increased ion flux.Conclusions Cooperative interactions between chloride and Mo25 regulate WNK signaling in a transporting renal epithelium.

Axial and cellular heterogeneity in electrolyte transport pathways along the thick ascending limb.

The thick ascending limb (TAL) extends from the border of the inner medulla to the renal cortex, thus ascending through regions with wide differences in tissue solute and electrolyte concentrations. Structural and functional differences between TAL cells in the medulla (mTAL) and the cortex (cTAL) would therefore be useful to adapt TAL transport function to a changing external fluid composition. While mechanisms common to all TAL cells play a central role in the reclamation of about 25% of the NaCl filtered by the kidney, morphological features, Na+ / K+ -ATPase activity, NKCC2 splicing and phosphorylation do vary between segments and cells. The TAL contributes to K+ homeostasis and TAL cells with high or low basolateral K+ conductances have been identified which may be involved in K+ reabsorption and secretion respectively. Although transport rates for HCO3- do not differ between mTAL and cTAL, divergent axial and cellular expression of H+ transport proteins in TAL have been documented. The reabsorption of the divalent cations Ca2+ and Mg2+ is highest in cTAL and paralleled by differences in divalent cation permeability and the expression of select claudins. Morphologically, two cell types with different cell surface phenotypes have been described that still need to be linked to specific functional characteristics. The unique external environment and its change along the longitudinal axis require an axial functional heterogeneity for the TAL to optimally participate in conserving electrolyte homeostasis. Despite substantial progress in understanding TAL function, there are still considerable knowledge gaps that are just beginning to become bridged.

SLC9 Gene Family: Function, Expression, and Regulation.

The Slc9 family of Na+ /H+ exchangers (NHEs) plays a critical role in electroneutral exchange of Na+ and H+ in the mammalian intestine as well as other absorptive and secretory epithelia of digestive organs. These transport proteins contribute to the transepithelial Na+ and water absorption, intracellular pH and cellular volume regulation as well as the electrolyte, acid-base, and fluid volume homeostasis at the systemic level. They also influence the function of other membrane transport mechanisms, affect cellular proliferation and apoptosis as well as cell migration, adherence to the extracellular matrix, and tissue repair. Additionally, they modulate the extracellular milieu to facilitate other nutrient absorption and to regulate the intestinal microbial microenvironment. Na+ /H+ exchange is inhibited in selected gastrointestinal diseases, either by intrinsic factors (e.g., bile acids, inflammatory mediators) or infectious agents and associated bacterial toxins. Disrupted NHE activity may contribute not only to local and systemic electrolyte imbalance but also to the disease severity via multiple mechanisms. In this review, we describe the cation proton antiporter superfamily of Na+ /H+ exchangers with a particular emphasis on the eight SLC9A isoforms found in the digestive tract, followed by a more integrative description in their roles in each of the digestive organs. We discuss regulatory mechanisms that determine the function of Na+ /H+ exchangers as pertinent to the digestive tract, their regulation in pathological states of the digestive organs, and reciprocally, the contribution of dysregulated Na+ /H+ exchange to the disease pathogenesis and progression. © 2018 American Physiological Society. Compr Physiol 8:555-583, 2018.

Dominant functional role of the novel phosphorylation site S811 in the human renal NaCl cotransporter.

The NaCl cotransporter (NCC) is essential for electrolyte homeostasis and control of blood pressure. The human SLC12A3 gene, which encodes NCC, gives rise to 3 isoforms, of which only the shortest isoform [NaCl cotransporter isoform 3 (NCC3)] has been studied extensively. All NCC isoforms share key phosphorylation sites at T55 and T60 that are essential mediators of NCC function. Recently, a novel phosphorylation site at S811 was identified in isoforms 1 and 2 [NaCl cotransporter splice variant (NCCSV)], which are only present in humans and higher primates. The aim of the current study, therefore, is to investigate the role of S811 phosphorylation in the regulation of NCC by a combination of biochemical and fluorescent microscopy analyses. We demonstrate that hypotonic low-chloride buffer increases S811 phosphorylation, whereas phosphorylation-deficient S811A mutant hinders phosphorylation at T55 and T60 in NCCSV and NCC3. NCCSV S811A impairs NCC3 activity in a dominant-negative fashion, although it does not affect plasma membrane abundance. This effect may be explained by the heterodimerization of NCCSV with NCC3. Taken together, our study highlights the dominant-negative effect of NCCSV on T55 and T60 phosphorylation and NCC activity. Here, we reveal a new function of NCCSV in humans that broadens the understanding on NCC regulation in blood pressure control.-Tutakhel, O. A. Z., Bianchi, F., Smits, D. A., Bindels, R. J. M., Hoenderop, J. G. J., van der Wijst, J. Dominant functional role of the novel phosphorylation site S811 in the human renal NaCl cotransporter.

A perfusion chamber for monitoring transepithelial NaCl transport in an in vitro model of the renal tubule.

Transepithelial electrical measurements in the renal tubule have provided a better understanding of how kidney regulates electrolyte and water homeostasis through the reabsorption of molecules and ions (e.g., H2 O and NaCl). While experiments and measurement techniques using native tissue are difficult to prepare and to reproduce, cell cultures conducted largely with the Ussing chamber lack the effect of fluid shear stress which is a key physiological stimulus in the renal tubule. To overcome these limitations, we present a modular perfusion chamber for long-term culture of renal epithelial cells under flow that allows the continuous and simultaneous monitoring of both transepithelial electrical parameters and transepithelial NaCl transport. The latter is obtained from electrical conductivity measurements since Na+ and Cl- are the ions that contribute most to the electrical conductivity of a standard physiological solution. The system was validated with epithelial monolayers of raTAL and NRK-52E cells that were characterized electrophysiologically for 5 days under different flow conditions (i.e., apical perfusion, basal, or both). In addition, apical to basal chemical gradients of NaCl (140/70 and 70/140 mM) were imposed in order to demonstrate the feasibility of this methodology for quantifying and monitoring in real time the transepithelial reabsorption of NaCl, which is a primary function of the renal tubule.

Moderate glucose supply reduces hemolysis during systemic inflammation.

BackgroundSystemic inflammation alters energy metabolism. A sufficient glucose level, however, is most important for erythrocytes, since erythrocytes rely on glucose as sole source of energy. Damage to erythrocytes leads to hemolysis. Both disorders of glucose metabolism and hemolysis are associated with an increased risk of death. The objective of the study was to investigate the impact of intravenous glucose on hemolysis during systemic inflammation.Materials and methodsSystemic inflammation was accomplished in male Wistar rats by continuous lipopolysaccharide (LPS) infusion (1 mg LPS/kg and h, 300 min). Sham control group rats received Ringer’s solution. Glucose was supplied moderately (70 mg glucose/kg and h) or excessively (210 mg glucose/kg and h) during systemic inflammation. Vital parameters (eg, systemic blood pressure) as well as blood and plasma parameters (eg, concentrations of glucose, lactate and cell-free hemoglobin, and activity of lactate dehydrogenase) were measured hourly. Clot formation was analyzed by thromboelastometry.ResultsContinuous infusion of LPS led to a so-called post-aggression syndrome with disturbed electrolyte homeostasis (hypocalcemia, hyperkalemia, and hypernatremia), changes in hemodynamics (tachycardia and hypertension), and a catabolic metabolism (early hyperglycemia, late hypoglycemia, and lactate formation). It induced severe tissue injury (significant increases in plasma concentrations of transaminases and lactate dehydrogenase), alterations in blood coagulation (disturbed clot formation), and massive hemolysis. Both moderate and excessive glucose supply reduced LPS-induced increase in systemic blood pressure. Excessive but not moderate glucose supply increased blood glucose level and enhanced tissue injury. Glucose supply did not reduce LPS-induced alterations in coagulation, but significantly reduced hemolysis induced by LPS.ConclusionIntravenous glucose infusion can diminish LPS-related changes in hemodynamics, glucose metabolism, and, more interestingly, LPS-induced hemolysis. Since cell-free hemoglobin is known to be a predictor for patient’s survival, a reduction of hemolysis by 35% only by the addition of a small amount of glucose is another step to minimize mortality during systemic inflammation.

Cheminformatics Analysis of Dynamic WNK-Inhibitor Interactions.

The With-No-Lysine (WNK) serine/threonine kinase family constitutes a unique and distinctive branch of the kinome. The four proteins of this family (WNK1/2/3/4) are involved in blood pressure regulation, body fluid, and electrolyte homeostasis. Herein, we modeled and analyzed the binding modes of all publicly-available small orthosteric and allosteric binders (including WNK463 and WNK467) experimentally tested towards any of the WNK family member. To do so, we relied on state-of-the-art cheminformatics approaches including structure-based molecular docking and molecular dynamics simulations. In particular, we computed and analyzed the (i) molecular selectivity of known inhibitors when docked in the binding site of each WNK family member, (ii) the dynamic WNK-inhibitor interactions at both orthosteric and allosteric sites to derive new structure-activity relationships, and (iii) the key specific interactions present in each binding site. This study reports on the first, cheminformatics-powered analysis of the entire chemical space of known WNK inhibitors. We discuss the conservation of critical WNK-inhibitor interactions and the existence of isoform-specific interactions that could enable the rational design of more potent and selective WNK binders.