Perspectives on edema in childhood nephrotic syndrome

Chia Wei Teoh, Lisa A. Robinson, and Damien NooneDivision of Nephrology, The Hospital for Sick Children, Toronto, Ontario, Canada

Submitted 29 May 2015; accepted in final form 11 August 2015

Teoh CW, Robinson LA, Noone D. Perspectives on edema in childhoodnephrotic syndrome. Am J Physiol Renal Physiol 309: F575–F582, 2015.First published August 19, 2015; doi:10.1152/ajprenal.00229.2015.—Therehave been two major theories surrounding the development of edemain nephrotic syndrome (NS), namely, the under- and overfill hypoth-eses. Edema is one of the cardinal features of NS and remains one ofthe principal reasons for admission of children to the hospital. Re-cently, the discovery that proteases in the glomerular filtrate ofpatients with NS are activating the epithelial sodium channel (ENaC),resulting in intrarenal salt retention and thereby contributing to edema,might suggest that targeting ENaC with amiloride might be a suitablestrategy to manage the edema of NS. Other potential agents, partic-ularly urearetics and aquaretics, might also prove useful in NS. Recentevidence also suggests that there may be other areas involved in saltstorage, especially the skin, and it will be intriguing to study theimplications of this in NS.

aquaretic; edema; ENaC; nephrotic syndrome; urearetic

NEPHROTIC SYNDROME (NS) is one of the most common child-hood kidney diseases with a cumulative prevalence of �16 in100,000 children (26). The diagnosis of NS is clinical, char-acterized by heavy proteinuria, hypoalbuminemia, and edema.NS can be classified into two main groups: the geneticallydetermined forms, involving podocyte and slit diaphragm pro-teins, which usually manifest in the first year of life (39), andthe idiopathic form. The main causes of idiopathic childhoodNS are minimal-change disease (MCD) (77.1%) and focalsegmental glomerulosclerosis (FSGS) (7.9%) (16).

The pathophysiological mechanisms of the syndrome re-main poorly defined despite decades of study. It is a multifac-torial disease, where the immune system targets structuralcomponents of the glomerular filtration barrier in a geneticallysusceptible patient. Many causative factors, including circulat-ing factors, genetic polymorphisms implicated in lymphocytematuration and differentiation, and DNA epigenetic modifica-tions, have been proposed and tested; none fully explains thepathogenesis of the disease (43).

The familial type of NS is primarily considered a “podocy-topathy” as it involves inherited abnormalities in slit dia-phragm or podocyte proteins, and the translational opportuni-ties and challenges associated with this form of NS haverecently been reviewed in this journal (37). However, the termpodocytopathy fails to take into account the fact that mutationsin proteins that make up the glomerular basement membrane,such as the collagen chains, �3, �4, and �5 (62) or thelaminins, may also lead to a proteinuric state, with laminin �2(LAMB2) being rarely associated with infantile or the congen-ital form of NS (17). The search also continues for the elusiveT lymphocyte-derived soluble factor proposed by Shalhoub in

1974 (78). The reader is also directed to recent comprehensivereviews on the subject of permeability factors in NS, detailingthe soluble urokinase plasminogen activator receptor story andproposing putative factors and translational strategies (18, 59).More recently, it has been suggested that even in MCD, both apermeability factor of immune origin and podocyte-derivedfactors may contribute to the development of NS (reviewed inRef. 43).

Edema is one of the classic clinical features of childhood NSand one of the principal reasons for admission to the hospital.The underlying mechanisms of edema formation remain asubject of ongoing investigation. This review aims to focus onthe current understanding of edema formation in NS and todiscuss recent developments in this area, potential targets fortherapy, and novel areas that need further scientific explora-tion.

Pathogenesis of Edema in NS: Under- vs. OverfillHypothesis

Two major opposing theories on the pathophysiologicalmechanisms underlying the development of edema in NS havebeen proposed: 1) the underfill hypothesis and 2) the overfillhypothesis (7, 13).

The underfill hypothesis. The underfill hypothesis was gen-erated almost a century ago (27). It postulates that high-gradeproteinuria results in hypoalbuminemia, leading to a reductionin plasma oncotic pressure with consequent leakage of plasmawater into the interstitium, generating edema. The resultantdiminished intravascular volume manifests with tachycardia,hypotension, and oliguria as well as peripheral vasoconstric-tion and water and sodium retention. This is affected byactivation of the renin-angiotensin-aldosterone system (RAAS),coupled with an increase in plasma norepinephrine and argi-nine vasopressin (AVP) concentrations (65, 76, 95). Together,this neurohormonal activation results in a highly concentratedurine with very low sodium content (44). In a study of 16pediatric and adult patients with NS with normal renal func-tion, Usberti et al. (92) found decreased plasma sodium con-centration, increased plasma AVP concentration, and an ele-vation in plasma renin activity (PRA) and urinary norepineph-rine levels, coupled with impaired excretion of an acute waterload, compared with controls. They demonstrated a highlysignificant inverse correlation between plasma AVP concen-tration and blood volume in these nephrotic patients (92).Therefore, in this scenario, the renal sodium retention wasthought to occur as a secondary physiological response tounderfilling (a reduced effective intravascular volume) (8). Thetherapeutic implications of this hypothesis are clear: expansionof intravascular volume and restoration of plasma oncoticpressure by administration of a colloid such as albumin wouldprove beneficial. Indeed, albumin infusion-induced volumeexpansion in children with NS has been shown to reduce PRA,

Address for reprint requests and other correspondence: D. Noone, Div. ofNephrology, The Hospital for Sick Children, 555 Univ. Ave., Toronto, ONM5G 1X8, Canada (e-mail: damien.noone@sickkids.ca).

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aldosterone, and AVP and to improve glomerular filtration rate(GFR), urine output, and sodium excretion (70, 90).

However, a decrease in measured plasma volume in ne-phrotic patients has not been a universal finding, and in somenephrotic states plasma volume may actually be increased (24,34, 66). Furthermore, elevated PRA or aldosterone, expected inthe setting of decreased blood volume, has only been con-firmed in about half of studied nephrotic subjects (35, 66).However, these variations in findings may be accounted for bythe fact that these parameters are dependent on the hemody-namic status of the patients at the time of the study.

Some clinical situations also do not fit well with the underfillhypothesis. The treatment of edema in NS with albumin alonereduces PRA, but it is often insufficient to induce diuresiswithout the addition of a diuretic (11, 94). In contrast, somepatients respond to diuretics alone without the need for albu-min (44). Moreover, the reduction of the renin-aldosterone axisby mineralocorticoid receptor antagonists or angiotensin-con-verting enzyme inhibitors does not result in increased sodiumexcretion in most patients with NS (10, 94). In fact, neurohor-monal markers such as renin and aldosterone levels do notshow a consistent pattern, suggesting volume depletion insome, and normal or excess volume in other patients (9, 11, 52,61, 93, 94, 96). In some patients, the administration of albuminis associated with volume overload and causes hypertensionand pulmonary edema (71). To the observant physician (andparent), the earliest sign of the patient entering remission is alarge diuresis, subsequently confirmed by the improvement inproteinuria. This occurs well before plasma protein levels (andthus plasma oncotic pressure) have normalized (64). Finally,there are patients with other causes of analbuminemia who donot typically suffer from edema (12, 54). Taken together, thesefindings suggest that the generation of edema in NS is not justsecondary to a reduction in plasma oncotic pressure as a resultof hypoalbuminemia caused by high-grade proteinuria.

Ichikawa et al. (42) provided strong experimental evidenceagainst the underfill hypothesis in a rat model of nephrosis, inwhich they performed nephron puncture after infusing puro-mycin aminoglycoside (PAN) into one kidney to independentlyassess the effect of proteinuria in both the nephrotic and thenormal kidney. They noted that only the nephrotic kidneyavidly conserved sodium, despite maintaining euvolemia bycontrolling plasma protein levels within the normal range byinfusion of homologous rat plasma. This suggested an intrare-nal mechanism of salt retention as opposed to a circulatingneurohumoral factor. In a separate study, Chandra et al. (15)found that the control kidney actually increased sodium excre-tion, as if sensing intravascular volume excess. They showedthat sodium delivery to the collecting duct was comparable inboth kidneys, whereas the final sodium excretion was higher inthe normal kidney, therefore suggesting that the collecting ductwas the main site of avid sodium reabsorption in the nephrotickidney.

The overfill hypothesis. Due to the inconsistent findingsdescribed above, the overfill hypothesis was proposed as analternative explanation for the development of edema in NS:proteinuria causes primary sodium retention with consequentvolume expansion and leakage of excess fluid into the inter-stitium (25, 61). In support of this idea, Dorhout Mees et al.(25) observed that blood pressure and plasma volume fell inpatients with NS after prednisone-induced remission (25). In

88 patients with NS, Gur et al. (36, 77) reported higher plasmaand blood volume corrected for estimated lean body masscompared with controls.

The Epithelial Sodium Channel and Sodium Retention in NS

What mediates the primary sodium retention? Ichikawa et al.(42) identified the collecting duct as the main segment involvedin sodium retention in NS (42). Initial focus was on thebasolateral Na-K-ATPase, as there is evidence from studies inrodent models of nephrosis to suggest an increase in levels andactivity of Na-K-ATPase in the collecting ducts of PAN-induced nephrotic rats (20, 97). Recent evidence suggests theprincipal means of sodium reabsorption in NS occurs in thedistal segment of the nephron via the epithelial sodium channel(ENaC) (Fig. 1) (50). Using both the PAN and HgCl2 rodentmodels of NS, Kim et al. (46, 47) demonstrated an increase inthe expression and targeting of ENaC to the apical cell surface,which is aldosterone dependent (19, 57). Lourdel et al. (57)showed a linear correlation between plasma aldosterone andENaC abundance. Although the prevention of hyperaldosteron-emia by adrenalectomy prevented the increase in apical ENaCtargeting, it did not prevent sodium retention and the develop-ment of NS in the PAN-treated rats (19, 57). Although theadrenalectomized rats did not show an increase in ENaCexpression, they had a similarly low urinary sodium excretionas adrenal-intact nephrotic rats with increased ENaC expres-sion (57). Treatment with amiloride returned sodium excretionto normal despite a lack of increase in ENaC expression (57).This aldosterone-independent sodium retention in NS by theENaC was further illustrated by Deschenes et al. (22), whoshowed that sodium retention can be prevented in the PAN-induced rat by amiloride and not aldosterone inhibition.

ENaC activity depends on channel density at the apicalmembrane, which is aldosterone and vasopressin dependent,and on channel activity, which is regulated by proteolyticprocessing and by anionic phospholipids on the inner cellmembrane (Fig. 1) (38, 58). The ENaC is composed of the �-,�-, and �-subunits (74), of which the �- and �-chains have aregulatory role. An ENaC channel with uncleaved �- and�-chains has low open channel probability and conducts littlesodium (40). Sequential proteolytic cleavage of the �- and�-subunits results in increasing open channel probability (40).Cleavage of an inhibitory tract from the �-subunit by serineproteases such as plasmin results in near full activation of theENaC (14). More recently, plasmin has been directly shown toactivate ENaC and promote sodium and water retention innephrotic rodents and humans (Fig. 2) (83). The plasminprecursor, plasminogen, is filtered by the nephrotic kidney andis activated to plasmin by the tubular urokinase-type plasmin-ogen activator present in the rat and human kidney. Use of thepotassium-sparing diuretic, amiloride, resulted in increasedurine sodium excretion and reduced ascites volume in the PANnephrosis rat. These effects were attributed both to inhibitionof ENaC and inhibition of urokinase-type plasminogen activa-tor by amiloride, thus reducing the amount of active plasminpresent in the urine (Fig. 2).

The dominant role of the ENaC in sodium retention and thedevelopment of nephrotic edema are underscored by the rever-sal of sodium retention by amiloride in animal models. Itprovides a rationale for specific blockade of ENaC with

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amiloride for the treatment of sodium retention and edemaformation in NS. While the administration of amiloride beforethe onset of sodium retention fully prevented sodium retentionand edema formation in PAN-induced nephrotic rats (22),clinicians usually encounter patients who are well beyond thisearly stage, many of whom have anasarca. In this situation,treatment aims both to limit further sodium retention and topromote diuresis and natriuresis. Under normal conditions,sodium reabsorption along the collecting duct is quantitativelylow, thus reducing the ability of amiloride to promote massivesodium excretion. Loop diuretics decrease sodium reabsorptionalong the thick ascending limb of the loop of Henle, increasingdistal sodium delivery, which, in turn, overloads the sodiumreabsorptive capacity of the distal nephron, resulting in natri-uresis. However, nephrotic patients display increased sodiumreabsorptive capacity along the cortical collecting ducts,thereby blunting the natriuretic effect. Combined use of loopdiuretics and amiloride, which inhibits distal sodium reabsorp-tion, could overcome the resistance to therapy with a loopdiuretic alone. Accordingly, Deschenes et al. (21) showed thatcombined therapy with amiloride and furosemide resulted ingreater natriuresis and weight loss than monotherapy witheither agent alone in pediatric patients (21). There has been nostudy interrogating the effectiveness of amiloride monotherapyin childhood NS. It has to be emphasized that most of thefindings in support of the overfill hypothesis are derived fromanimal studies with few studies in humans.

In NS, ENaC activation occurs following proteolytic pro-cessing of the protein. Therefore, inhibition of serine proteaseactivity could represent another potential therapeutic target(82). Accordingly, camostat mesilate (CM), a synthetic inhib-

itor of serine proteases such as plasmin and prostasin, wasshown to reduce aldosterone-induced renal ENaC proteolysisin rats (91). In a small case series, treatment with CM causeda significant reduction in proteinuria in patients with NS ofvarious etiologies. Although these reports suggest that sys-temic administration of serine protease inhibitors to humanpatients is well tolerated, improves proteinuria, and attenuatesedema formation through inhibition of ENaC-mediated sodiumretention, the potential for systemic effects related to un-specific protease inhibition suggests a note of caution (82).Using inhibitors that are freely filtered across the glomerularbarrier or secreted into the tubules could potentially reducethe side effects associated with systemic inhibition of pro-tease activity (82).

The Role of Corin

Corin is an endogenous type II transmembrane serine pro-tease that cleaves pro atrial natriuretic peptide (ANP) and probrain natriuretic peptide (BNP) to their biologically activeforms and is detected in multiple organs, including the heart,brain, and kidney as well as in the serum and urine (23, 31, 68,102, 103). Its importance in blood pressure was demonstratedby the demonstration of salt-sensitive hypertension in corin-deficient mice (102). ANP has three important roles in sodiumand water excretion in the kidney, namely, 1) inducing afferentarteriolar vasodilation and efferent vasoconstriction, therebyincreasing GFR; 2) causing natriuresis through inhibition of theN�/H� exchanger in the proximal tubule, the distal Na�-Cl�

cotransporter, and ENaC; and 3) causing diuresis throughinhibition of aquaporin-2 insertion in the apical membrane of

Fig. 1. Model of epithelial sodium channel(ENaC) regulation by aldosterone and vaso-pressin. Upon binding of aldosterone to itsreceptor (MR), the ligand-receptor translo-cates into the nucleus, leading to the induc-tion of a specific set of aldosterone-inducedtranscripts (AIT) such as ENaC, the Na/K-ATPase, or SGK1. Vasopressin binds to itsreceptor (V2R), leading to the activation of adownstream protein kinase A (PKA). BothSGK1 and PKA inactivate Nedd4 uponphosphorylation, leading to retention of ac-tive ENaC at the cell surface. Adapted fromRef. 34 with permission.

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the collecting duct, normally mediated via arginine vasopressin(reviewed in Ref. 2). Polzin et al. (69) also suggested the rolefor corin in the salt retention in NS. They demonstrated areduction in kidney corin expression in the PAN nephrosis ratcoupled with a concomitant increase in pro-ANP and decreasedANP levels. They then proceeded to demonstrate increasedamounts of renal �-ENaC and its activators (phosphodiesterase5 and protein kinase GII) in corin knockout mice comparedwith wild-type mice (69). Further exploration of the linkbetween corin and ENaC may further enhance our understand-ing of the salt retention in NS (48).

The Role of Aquaretics

In NS, vasopressin levels are reported to be elevated, leadingto the production of concentrated urine and impaired excretionof a water load (44, 65, 95). In keeping with this observation,nephrotic patients are often hyponatremic, consistent with a

relative water excess (92). Type 2 vasopressin receptors(AVPR2) mediate urinary concentration in the collecting duct,and AVPR2 inhibitors have proven effective in the treatment ofedema (1). Konomoto et al. (51) highlighted a potential role ofvasopressin in edema formation in a child with relapsingsteroid-sensitive NS who developed cranial diabetes insipidus.Relapses of NS before the development of cranial diabetesinsipidus and after treatment with DDAVP were associatedwith edema. However, when a relapse occurred in the presenceof untreated cranial diabetes insipidus, the patient was edemafree despite high levels of proteinuria and low plasma albumin.In another report, a child with steroid-resistant NS and edema,pleural and pericardial effusions resistant to treatment withalbumin/furosemide responded significantly to tolvaptan (aselective, competitive vasopressin receptor 2 antagonist), de-spite persistent nephrotic range proteinuria and unchangedplasma albumin levels (79).

Fig. 2. Plasmin stimulates ENaC-mediatedsodium reabsorption in proteinuria. Duringproteinuria, plasminogen that is present inthe tubular lumen is activated to plasmin byurokinase-type plasminogen activator (uPA),which is bound to its receptor (uPAR). Plas-min can subsequently activate ENaC eitherthrough interaction with prostasin or throughdirect cleavage of the �-ENaC subunits.Adapted from Ref. 84 with permission.

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While these case reports offer a novel approach to treatingedema in nephrotic patients, injudicious use of aquaretics maybe fraught with potential risks of serious hypovolemia, hyper-natremic dehydration, shock, and thrombosis (6), especially inthe consideration of the thrombophilic tendencies in NS (80).Aquaretics are, thus, best avoided in NS.

“Urearetics”: Blocking Urea Transporter Proteins

Since urea transport plays an important role in urinaryconcentration, urea-blocking agents may represent anothertherapeutic tool to treat NS-related edema (67, 72). Ureatransporter (UT) proteins facilitate the passive transport ofurea, driven by a concentration gradient, across cell mem-

branes, and are essential in the urinary concentrating mecha-nism (28). Different isoforms of the transporters UT-A andUT-B are expressed in the descending limb and the collectingduct of the medullary nephrons (28, 45, 81, 85, 98). UTreceptor accumulation in the plasma membrane is regulatedmainly by vasopressin (via V2 receptors) (4, 5, 41, 49, 106)and hypertonicity (3, 99, 100). UT expressed in the innermedullary collecting duct is responsible for urea reabsorption,which maintains a hypertonic medullary interstitium. Thehighly urea-permeable fenestrated ascending vasa recta facili-tate transfer of urea from the inner to the outer medulla. Theinterstitial urea from the ascending vasa recta is partiallyreabsorbed by the nonfenestrated descending vasa recta and the

Fig. 3. Urea transporter blockade by “urearetics”, salt-sparing diuretics. Aquaporin and urea transporters in the descending thin limb of the loop of Henle andinner medullary collecting duct facilitate the passive transport of urea, driven by a concentration gradient, across cell membranes, and are essential in the urinaryconcentrating mechanism. Urea transporter inhibition and its subsequent effect on the countercurrent mechanism may be promising therapeutic targets as it hasthe potential to be effective in refractory edema, producing a relatively salt-sparing diuresis compared with conventional diuretics. Adapted from Ref. 33.

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thin descending limb of the loop of Henle by a countercurrentexchange mechanism in which urea recycling conserves renalurea (67, 72, 73). UT-deficient mice demonstrate a reducedcapacity to concentrate urea in the urine, manifesting as areduction in maximum urinary osmolality and up to a twofoldincrease in daily urine output (32, 104) (Fig. 3).

Various small-molecule inhibitors of UT isoforms have beendeveloped and tested successfully as diuretics in differentrodent models (29, 30, 55, 105). Selective UT-A isoforminhibition has an added advantage for potential diuretic therapyin that it is primarily expressed in the kidneys, reducing thepotential of extrarenal effects compared with UT-B, which isrelatively ubiquitous (28). These studies collectively highlighturea transporter inhibition as potential targets for therapeuticdrug development. Their effect on the countercurrent mecha-nism may be promising as it has the potential to be effective inrefractory edema, producing a relatively salt-sparing diuresiscompared with conventional diuretics. Its use (with or withoutconventional diuretics) may be effective in hyponatremia dueto the syndrome of inappropriate antidiuretic hormone secre-tion and in fluid-overload states like congestive cardiac failure,chronic liver failure, chronic kidney disease, and NS. However,further testing of these inhibitors in animal models, and ulti-mately in clinical trials, is needed to define their safety andefficacy before mainstream therapeutic use.

Alternative View on Extrarenal Sodium Balance

Conventional understanding of sodium balance maintainsthat extracellular body fluids readily equilibrate across com-partments, that electrolyte concentrations in various compart-ments are constant, and that the kidney is the main player incontrolling the body’s sodium content. This paradigm has beenquestioned by recent studies that showed “kidney-like” lym-phatic and blood vessel countercurrent systems in the skin,intestines, bone, and elsewhere in the body (53, 56, 86–88).The notion that the immune system plays a role as homeostaticregulator of interstitial electrolyte homeostasis and that saltinduces proinflammatory immune cell polarization has ex-tended the role of the immune system to physiological adap-tation, interstitial fluid matrix regulation, blood pressure con-trol, and cardiovascular disease (60, 101). The protein-rich andnegatively charged glycosaminoglycan-rich endothelial sur-face layer may act as a buffer to excess intravascular sodium,by binding and osmotically inactivating it (89), and a barrier towater entering the interstitium, which make this an intriguingarea of future study for sodium hemostasis in both health anddisease (reviewed in Ref. 63). These findings may have impli-cations for our understanding of the sodium and fluid retentionin NS and drive the development of entirely novel therapeuticagents.

Conclusion

Understanding the mechanism of edema formation in NS hasfascinated clinicians and scientists for decades, and the studyof renal physiology in the nephrotic state has shed light onnormal physiology. The observation that proteases in the glo-merular filtrate activate the sodium-retaining ENaC, thus pro-moting intrarenal salt retention, has yet to be translated into anew therapeutic approach, despite the availability of amiloride.Is this because of concerns about hyperkalemia and the poten-

tial ineffectiveness of ENaC inhibition alone, or is there moreto the story? Can aquaretics and urearetics be used to manageedema in NS? These key questions need to be answered.

DISCLOSURES

No conflicts of interest, financial or otherwise, are declared by the authors.

AUTHOR CONTRIBUTIONS

Author contributions: C.W.T., L.A.R., and D.G.N. provided conception anddesign of research; C.W.T., L.A.R., and D.G.N. prepared figures; C.W.T.,L.A.R., and D.G.N. drafted manuscript; C.W.T., L.A.R., and D.G.N. editedand revised manuscript; C.W.T., L.A.R., and D.G.N. approved final version ofmanuscript.

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