On the role of the epithelium in a model of sodium exchange in renal tubules.

In this study we present a mathematical model describing the transport of sodium in a fluid circulating in a counter-current tubular architecture, which constitutes a simplified model of Henle's loop in a kidney nephron. The model explicitly takes into account the epithelial layer at the interface between the tubular lumen and the surrounding interstitium. In a specific range of parameters, we show that explicitly accounting for transport across the apical and basolateral membranes of epithelial cells, instead of assuming a single barrier, affects the axial concentration gradient, an essential determinant of the urinary concentrating capacity. We present the solution related to the stationary system, and we perform numerical simulations to understand the physiological behaviour of the system. We prove that when time grows large, our dynamic model converges towards the stationary system at an exponential rate. In order to prove rigorously this global asymptotic stability result, we study eigen-problems of an auxiliary linear operator and its dual.

Salt stress in the renal tubules is linked to TAL-specific expression of uromodulin and an upregulation of heat shock genes.

Previously, our comprehensive cardiovascular characterization study validated Uromodulin as a blood pressure gene. Uromodulin is a glycoprotein exclusively synthesized at the thick ascending limb of the loop of Henle and is encoded by the Umod gene. Umod-/- mice have significantly lower blood pressure than Umod+/+ mice, are resistant to salt-induced changes in blood pressure, and show a leftward shift in pressure-natriuresis curves reflecting changes of sodium reabsorption. Salt stress triggers transcription factors and genes that alter renal sodium reabsorption. To date there are no studies on renal transcriptome responses to salt stress. Here we aimed use RNA-Seq to delineate salt stress pathways in tubules isolated from Umod+/+ mice (a model of sodium retention) and Umod-/- mice (a model of sodium depletion) ± 300 mosmol sodium chloride ( n = 3 per group). In response to salt stress, the tubules of Umod+/+ mice displayed an upregulation of heat shock transcripts. The greatest changes occurred in the expression of: Hspa1a (Log2 fold change 4.35, P = 2.48 e-12) and Hspa1b (Log2 fold change 4.05, P = 2.48 e-12). This response was absent in tubules of Umod-/- mice. Interestingly, seven of the genes discordantly expressed in the Umod-/- tubules were electrolyte transporters. Our results are the first to show that salt stress in renal tubules alters the transcriptome, increasing the expression of heat shock genes. This direction of effect in Umod+/+ tubules suggest the difference is due to the presence of Umod facilitating greater sodium entry into the tubule cell reflecting a specific response to salt stress.

Paracellular transport and energy utilization in the renal tubule.

PURPOSE OF REVIEW: Paracellular transport across the tight junction is a general mechanism for transepithelial transport of solutes in epithelia, including the renal tubule. However, why paracellular transport evolved, given the existence of a highly versatile system for transcellular transport, is unknown. RECENT FINDINGS: Recent studies have identified the paracellular channel, claudin-2, that is responsible for paracellular reabsorption of sodium in the proximal renal tubule. Knockout of claudin-2 in mice impairs proximal sodium and fluid reabsorption but is compensated by upregulation of sodium reabsorption in the loop of Henle. This occurs at the expense of increased renal oxygen consumption, hypoxia of the outer medulla and increased susceptibility to ischemic kidney injury. SUMMARY: Paracellular transport can be viewed as a mechanism to exploit the potential energy in existing electrochemical gradients to drive passive transepithelial transport without consuming additional energy. In this way, it enhances the efficiency of energy utilization by transporting epithelia.

Role of renal transporters and novel regulatory interactions in the TAL that control blood pressure.

Hypertension (HTN), a major public health issue is currently the leading factor in the global burden of disease, where associated complications account for 9.4 million deaths worldwide every year. Excessive dietary salt intake is among the environmental factors that contribute to HTN, known as salt sensitivity. The heterogeneity of salt sensitivity and the multiple mechanisms that link high salt intake to increases in blood pressure are of upmost importance for therapeutic application. A continual increase in the kidney's reabsorption of sodium (Na+) relies on sequential actions at various segments along the nephron. When the distal segments of the nephron fail to regulate Na+, the effects on Na+ homeostasis are unfavorable. We propose that the specific nephron region where increased active uptake occurs as a result of variations in Na+ reabsorption is at the thick ascending limb of the loop of Henle (TAL). The purpose of this review is to urge the consideration of the TAL as contributing to the pathophysiology of salt-sensitive HTN. Further research in this area will enable development of a therapeutic application for targeted treatment.

Genetic causes of hypomagnesemia, a clinical overview.

Magnesium is essential to the proper functioning of numerous cellular processes. Magnesium ion (Mg2+) deficits, as reflected in hypomagnesemia, can cause neuromuscular irritability, seizures and cardiac arrhythmias. With normal Mg2+ intake, homeostasis is maintained primarily through the regulated reabsorption of Mg2+ by the thick ascending limb of Henle's loop and distal convoluted tubule of the kidney. Inadequate reabsorption results in renal Mg2+ wasting, as evidenced by an inappropriately high fractional Mg2+ excretion. Familial renal Mg2+ wasting is suggestive of a genetic cause, and subsequent studies in these hypomagnesemic families have revealed over a dozen genes directly or indirectly involved in Mg2+ transport. Those can be classified into four groups: hypercalciuric hypomagnesemias (encompassing mutations in CLDN16, CLDN19, CASR, CLCNKB), Gitelman-like hypomagnesemias (CLCNKB, SLC12A3, BSND, KCNJ10, FYXD2, HNF1B, PCBD1), mitochondrial hypomagnesemias (SARS2, MT-TI, Kearns-Sayre syndrome) and other hypomagnesemias (TRPM6, CNMM2, EGF, EGFR, KCNA1, FAM111A). Although identification of these genes has not yet changed treatment, which remains Mg2+ supplementation, it has contributed enormously to our understanding of Mg2+ transport and renal function. In this review, we discuss general mechanisms and symptoms of genetic causes of hypomagnesemia as well as the specific molecular mechanisms and clinical phenotypes associated with each syndrome.

Mosaic expression of claudins in thick ascending limbs of Henle results in spatial separation of paracellular Na+ and Mg2+ transport.

The thick ascending limb (TAL) of Henle's loop drives paracellular Na+, Ca2+, and Mg2+ reabsorption via the tight junction (TJ). The TJ is composed of claudins that consist of four transmembrane segments, two extracellular segments (ECS1 and -2), and one intracellular loop. Claudins interact within the same (cis) and opposing (trans) plasma membranes. The claudins Cldn10b, -16, and -19 facilitate cation reabsorption in the TAL, and their absence leads to a severe disturbance of renal ion homeostasis. We combined electrophysiological measurements on microperfused mouse TAL segments with subsequent analysis of claudin expression by immunostaining and confocal microscopy. Claudin interaction properties were examined using heterologous expression in the TJ-free cell line HEK 293, live-cell imaging, and Förster/FRET. To reveal determinants of interaction properties, a set of TAL claudin protein chimeras was created and analyzed. Our main findings are that (i) TAL TJs show a mosaic expression pattern of either cldn10b or cldn3/cldn16/cldn19 in a complex; (ii) TJs dominated by cldn10b prefer Na+ over Mg2+, whereas TJs dominated by cldn16 favor Mg2+ over Na+; (iii) cldn10b does not interact with other TAL claudins, whereas cldn3 and cldn16 can interact with cldn19 to form joint strands; and (iv) further claudin segments in addition to ECS2 are crucial for trans interaction. We suggest the existence of at least two spatially distinct types of paracellular channels in TAL: a cldn10b-based channel for monovalent cations such as Na+ and a spatially distinct site for reabsorption of divalent cations such as Ca2+ and Mg2.

The C-Terminal Fragment of Agrin (CAF), a Novel Marker of Renal Function, Is Filtered by the Kidney and Reabsorbed by the Proximal Tubule.

Agrin, a multidomain proteoglycan and neurotrypsin, a neuronal serine protease, are important for forming (neuromuscular) synapses. Proteolytical activity of neurotrypsin produces a C-terminal fragment of agrin, termed CAF, of approximately 22 kDA molecular size which also circulates in blood. The presence of CAF in urine suggests either glomerular filtration or secretion into urine. Blood levels of CAF have been identified as a potential novel marker of kidney function. Here we describe that several nephron segments in the mouse kidney express agrin and neutrotrypsin in addition to the localization of both protein in the glomerulum. Agrin mRNA and protein was detected in almost all nephron segments and mRNA abundance was highest in the inner medullary collecting duct. Neurotrypsin mRNA was mostly detected in the thick ascending limb of the loop of Henle, the distal convoluted tubule, and the inner medullary collecting duct. Moreover, we show that the proximal tubule absorbs injected recombinant CAF by a process shared with receptor-mediated and fluid phase endocytosis. Co-injection of CAF with recombinant human transferrin, a substrate of the receptor-mediated endocytic pathway as well as with FITC-labelled dextran (10 kDa), a marker of fluid phase endocytosis, showed partial colocalization of CAF with both markers. Further colocalization of CAF with the lysosomal marker cathepsin B suggested degradation of CAF by the lysosome in the proximal tubule. Thus, the murine kidney expresses agrin and neurotrypsin in nephron segments beyond the glomerulum. CAF is filtered by the glomerulum and is reabsorbed by endocytosis by the proximal tubule. Thus, impaired kidney function could impair glomerular clearance of CAF and thereby increase circulating CAF levels.

Mathematical Model of Ammonia Handling in the Rat Renal Medulla.

The kidney is one of the main organs that produces ammonia and release it into the circulation. Under normal conditions, between 30 and 50% of the ammonia produced in the kidney is excreted in the urine, the rest being absorbed into the systemic circulation via the renal vein. In acidosis and in some pathological conditions, the proportion of urinary excretion can increase to 70% of the ammonia produced in the kidney. Mechanisms regulating the balance between urinary excretion and renal vein release are not fully understood. We developed a mathematical model that reflects current thinking about renal ammonia handling in order to investigate the role of each tubular segment and identify some of the components which might control this balance. The model treats the movements of water, sodium chloride, urea, NH3 and [Formula: see text], and non-reabsorbable solute in an idealized renal medulla of the rat at steady state. A parameter study was performed to identify the transport parameters and microenvironmental conditions that most affect the rate of urinary ammonia excretion. Our results suggest that urinary ammonia excretion is mainly determined by those parameters that affect ammonia recycling in the loops of Henle. In particular, our results suggest a critical role for interstitial pH in the outer medulla and for luminal pH along the inner medullary collecting ducts.

A mathematical model of rat proximal tubule and loop of Henle.

Proximal tubule and loop of Henle function are coupled, with proximal transport determining loop fluid composition, and loop transport modulating glomerular filtration via tubuloglomerular feedback (TGF). To examine this interaction, we begin with published models of the superficial rat proximal convoluted tubule (PCT; including flow-dependent transport in a compliant tubule), and the rat thick ascending Henle limb (AHL). Transport parameters for this PCT are scaled down to represent the proximal straight tubule (PST), which is connected to the thick AHL via a short descending limb. Transport parameters for superficial PCT and PST are scaled up for a juxtamedullary nephron, and connected to AHL via outer and inner medullary descending limbs, and inner medullary thin AHL. Medullary interstitial solute concentrations are specified. End-AHL hydrostatic pressure is determined by distal nephron flow resistance, and the TGF signal is represented as a linear function of end-AHL cytosolic Cl concentration. These two distal conditions required iterative solution of the model. Model calculations capture inner medullary countercurrent flux of urea, and also suggest the presence of an outer medullary countercurrent flux of ammonia, with reabsorption in AHL and secretion in PST. For a realistically strong TGF signal, there is the expected homeostatic impact on distal flows, and in addition, a homeostatic effect on proximal tubule pressure. The model glycosuria threshold is compatible with rat data, and predicted glucose excretion with selective 1Na(+):1glucose cotransporter (SGLT2) inhibition comports with observations in the mouse. Model calculations suggest that enhanced proximal tubule Na(+) reabsorption during hyperglycemia is sufficient to activate TGF and contribute to diabetic hyperfiltration.

Hormonal regulation of salt and water excretion: a mathematical model of whole kidney function and pressure natriuresis.

We present a lumped-nephron model that explicitly represents the main features of the underlying physiology, incorporating the major hormonal regulatory effects on both tubular and vascular function, and that accurately simulates hormonal regulation of renal salt and water excretion. This is the first model to explicitly couple glomerulovascular and medullary dynamics, and it is much more detailed in structure than existing whole organ models and renal portions of multiorgan models. In contrast to previous medullary models, which have only considered the antidiuretic state, our model is able to regulate water and sodium excretion over a variety of experimental conditions in good agreement with data from experimental studies of the rat. Since the properties of the vasculature and epithelia are explicitly represented, they can be altered to simulate pathophysiological conditions and pharmacological interventions. The model serves as an appropriate starting point for simulations of physiological, pathophysiological, and pharmacological renal conditions and for exploring the relationship between the extrarenal environment and renal excretory function in physiological and pathophysiological contexts.

Urine-concentrating mechanism in the inner medulla: function of the thin limbs of the loops of Henle.

The ability of mammals to produce urine hyperosmotic to plasma requires the generation of a gradient of increasing osmolality along the medulla from the corticomedullary junction to the papilla tip. Countercurrent multiplication apparently establishes this gradient in the outer medulla, where there is substantial transepithelial reabsorption of NaCl from the water-impermeable thick ascending limbs of the loops of Henle. However, this process does not establish the much steeper osmotic gradient in the inner medulla, where there are no thick ascending limbs of the loops of Henle and the water-impermeable ascending thin limbs lack active transepithelial transport of NaCl or any other solute. The mechanism generating the osmotic gradient in the inner medulla remains an unsolved mystery, although it is generally considered to involve countercurrent flows in the tubules and vessels. A possible role for the three-dimensional interactions between these inner medullary tubules and vessels in the concentrating process is suggested by creation of physiologic models that depict the three-dimensional relationships of tubules and vessels and their solute and water permeabilities in rat kidneys and by creation of mathematical models based on biologic phenomena. The current mathematical model, which incorporates experimentally determined or estimated solute and water flows through clearly defined tubular and interstitial compartments, predicts a urine osmolality in good agreement with that observed in moderately antidiuretic rats. The current model provides substantially better predictions than previous models; however, the current model still fails to predict urine osmolalities of maximally concentrating rats.

Tubular fluid flow and distal NaCl delivery mediated by tubuloglomerular feedback in the rat kidney.

The glomerular filtration rate in the kidney is controlled, in part, by the tubuloglomerular feedback (TGF) system, which is a negative feedback loop that mediates oscillations in tubular fluid flow and in fluid NaCl concentration of the loop of Henle. In this study, we developed a mathematical model of the TGF system that represents NaCl transport along a short loop of Henle with compliant walls. The proximal tubule and the outer-stripe segment of the descending limb are assumed to be highly water permeable; the thick ascending limb (TAL) is assumed to be water impermeable and have active NaCl transport. A bifurcation analysis of the TGF model equations was performed by computing parameter boundaries, as functions of TGF gain and delay, that separate differing model behaviors. The analysis revealed a complex parameter region that allows a variety of qualitatively different model equations: a regime having one stable, time-independent steady-state solution and regimes having stable oscillatory solutions of different frequencies. A comparison with a previous model, which represents only the TAL explicitly and other segments using phenomenological relations, indicates that explicit representation of the proximal tubule and descending limb of the loop of Henle lowers the stability of the TGF system. Model simulations also suggest that the onset of limit-cycle oscillations results in increases in the time-averaged distal NaCl delivery, whereas distal fluid delivery is not much affected.

The loop of Henle as the milestone of mammalian kindey concentrating ability: a historical review.

The first description of the renal tubules is attributed to Lorenzo Bellini in 1662 and four years later Marcello Malpighi described the glomerulus. In 1842 Sir William Bowman described the capsule that surrounds the Malpighian body and its connection with the renal tubule and introduced the "excretory" hypothesis of urine formation. In the same year, Carl Ludwig introduced the "filtration-reabsorption" hypothesis of urine formation. Bowman's hypothesis was accepted by the so-called "vitalists" and Ludwig's hypothesis by the so-called "mechanists". In the middle of this confliction, Jacob Henle described in 1862 the homonymous "U" shaped loop but his discovery has neglected. In 1942 Werner Kuhn, a physical chemist, proposed that the loop of Henle may be the natural analog of the hairpin countercurrent multiplication system which concentrates urine in mammalian kidneys. In 1951 Kuhn, Hargitay and Wirz showed experimentally that the loop of Henle was the most important part of the countercurrent multiplication system of urine-concentrating mechanism in mammalian kidneys. The new theory was accepted by English-speaking scientists later, in 1958, when Carl Gottschalk and Margaret Mylle published their experimental work and proved that Kuhn's theory was correct. Gottschalk summarized the evidence of the accumulated knowledge in 1962, three centuries after the first description of renal tubules and one century after description of Henle's loop.

Effect of tubular inhomogeneities on feedback-mediated dynamics of a model of a thick ascending limb.

One of the key mechanisms that mediate renal autoregulation is the tubuloglomerular feedback (TGF) system, which is a negative feedback loop in the kidney that balances glomerular filtration with tubular reabsorptive capacity. Tubular fluid flow, NaCl concentration and other related variables are known to exhibit TGF-mediated oscillations. In this study, we used a mathematical model of the thick ascending limb (TAL) of a short loop of Henle of the rat kidney to study the effects of (i) spatially inhomogeneous TAL NaCl active transport rate, (ii) spatially inhomogeneous tubular radius and (iii) compliance of the tubular walls on TGF-mediated dynamics. A bifurcation analysis of the TGF model equations was performed by deriving a characteristic equation and finding its roots. Results of the bifurcation analysis were validated via numerical simulations of the full model equations. Model results suggest that a higher TAL NaCl active transport rate or a smaller TAL radius near the loop bend gives rise to stable oscillatory solutions at sufficiently high TGF gain values, even with zero TGF delay. In addition, when the TAL walls are assumed to be compliant, the TGF system exhibits a heightened tendency to oscillate, a result that is consistent with predictions of a previous modelling study.

An improved method of renal tissue engineering, by combining renal dissociation and reaggregation with a low-volume culture technique, results in development of engineered kidneys complete with loops of Henle.

BACKGROUND: Tissue engineering of functional kidney tissue is an important goal for clinical restoration of renal function in patients damaged by infectious, toxicological, or genetic disease. One promising approach is the use of the self-organizing abilities of embryonic kidney cells to arrange themselves, from a simply reaggregated cell suspension, into engineered organs similar to fetal kidneys. The previous state-of-the-art method for this results in the formation of a branched collecting duct tree, immature nephrons (S-shaped bodies) beside and connected to it, and supportive stroma. It does not, though, result in the significant formation of morphologically detectable loops of Henle - anatomical features of the nephron that are critical to physiological function. METHODS: We have combined the best existing technique for renal tissue engineering from cell suspensions with a low-volume culture technique that allows intact kidney rudiments to make loops of Henle to test whether engineered kidneys can produce these loops. RESULTS: The result is the formation of loops of Henle in engineered cultured 'fetal kidneys', very similar in both morphology and in number to those formed by intact organ rudiments. CONCLUSION: This brings the engineering technique one important step closer to production of a fully realistic organ.

[Magnesium homeostasis and its disturbances].

Magnesium homeostasis is maintained through normal functions of the kidney, intestine, and bone. In the kidney, approximately 80% magnesium is filtered by the glomeruli. In general, 95% filtered magnesium is collectively reabsorbed in the proximal tubule (15%-20%) , thick ascending limb of Henle (TAL, 65%-75%) , and the distal convoluted tubule (DCT, 5%-10%) . In the TAL, magnesium reabsorption regulated by the paracellular pathway via claudin-16 is driven by electrochemical voltage. Chloride channel Kb and renal outer medullary potassium channels control this lumen-positive voltage. In the DCT, the transcellular pathway via transient receptor potential melastatin 6 (TRPM6) plays a fundamental role in the final 5%-10% magnesium reabsorption. The functions of TRPM6 depend on Na-Cl co-transporters and Na( + )-K( + )-ATPase. Defects in these regulatory proteins may cause inherited or drug-induced disorders of magnesium metabolism. Recently, some proteins have been confirmed to be responsible for magnesium homeostasis ; however, further research is required to elucidate the mechanisms underlying the maintenance of magnesium homeostasis.

Mitochondrial proteomic analysis reveals deficiencies in oxygen utilization in medullary thick ascending limb of Henle in the Dahl salt-sensitive rat.

The renal medullary thick ascending limb (mTAL) of the Dahl salt-sensitive (SS) rat is the site of enhanced NaCl reabsorption and excess superoxide production. In the present studies we isolated mitochondria from mTAL of SS and salt-resistant control strain SS.13(BN) rats on 0.4 and 8% salt diet for 7 days and performed a proteomic analysis. Purity of mTAL and mitochondria isolations exceeded 93.6 and 55%, respectively. Using LC/MS spectral analysis techniques we identified 96 mitochondrial proteins in four biological mTAL mitochondria samples, run in duplicate, as defined by proteins with a false discovery rate <5% and scan count ≥2. Seven of these 96 proteins, including IDH2, ACADM, SCOT, Hsp60, ATPA, EFTu, and VDAC2 were differentially expressed between the two rat strains. Oxygen consumption and high-resolution respirometry analyses showed that mTAL cells and the mitochondria in the outer medulla of SS rats fed high-salt diet exhibited lower rates of oxygen utilization compared with those from SS.13(BN) rats. These studies advance the conventional proteomic paradigm of focusing exclusively upon whole tissue homogenates to a focus upon a single cell type and specific subcellular organelle. The results reveal the importance of a largely unexplored role for deficiencies of mTAL mitochondrial metabolism and oxygen utilization in salt-induced hypertension and renal medullary oxidative stress.

Signal transduction in a compliant short loop of Henle.

To study the transformation of fluctuations in filtration rate into tubular fluid chloride concentration oscillations alongside the macula densa, we have developed a mathematical model for tubuloglomerular feedback (TGF) signal transduction along the pars recta, the descending limb, and the thick ascending limb (TAL) of a short-looped nephron. The model tubules are assumed to have compliant walls and, thus, a tubular radius that depends on the transmural pressure difference. Previously, it has been predicted that TGF transduction by the TAL is a generator of nonlinearities: if a sinusoidal oscillation is added to a constant TAL flow rate, then the time required for a fluid element to traverse the TAL is oscillatory in time but nonsinusoidal. The results from the new model simulations presented here predict that TGF transduction by the loop of Henle is also, in the same sense, a generator of nonlinearities. Thus, this model predicts that oscillations in tubular fluid alongside the macula densa will be nonsinusoidal and will exhibit harmonics of sinusoidal perturbations of pars recta flow. Model results also indicate that the loop acts as a low-pass filter in the transduction of the TGF signal.