Proximal Reabsorption Research Articles

Peculiarities of ionoregulatory renal function of rats in the dynamics of experimental diabetes mellitus development

Background. The purpose of the research was to study the peculiarities of ionoregulatory renal function in the dynamics of alloxan-induced experimental diabetes mellitus. Materials and methods. The experiments were carried out on 63 white non-linear mature male rats, 53 with experimental diabetes mellitus of varying duration induced by intraperitoneal administration of alloxan in a dose of 160 mg/kg of body weight, and 10 intact rats (control group). Ten, 20, 25, 30, 40 and 45 days after administration of the diabetogenic substance, the animals were withdrawn from the experiment. Ionoregulatory function of the kidneys was studied by means of the clearance method under condition of water induced 2-hour diuresis to determine the clearance of endogenous creatinine, glomerular filtration rate, sodium and potassium content in the urine and blood plasma. It was followed by calculation of the electrolyte excretion, intensity of their filtration, absolute and relative reabsorption, their proximal and distal tubular transport (including standardized by glomerular filtrate volume). Results. The ionoregulatory function of the kidneys in rats with alloxan-induced experimental diabetes is characterized by the intensification of natriuresis and kaliuresis at all stages of the experiment. An increase in the urinary sodium loss in the early stages of alloxan-induced experimental diabetes is primarily stipulated by glomerular hyperfiltration, followed by an enhancement of filtration sodium load to the nephron. The loss of proportionality between the filtered amount of sodium and its proximal reabsorption causes a decrease in the total reabsorption potential of the tubular segment of the nephron in the dynamics of alloxan-induced experimental diabetes. It is reflected primarily on the proximal tubules, and subsequently induces a functional weakening of the tubule-tubular connection and relative dysfunction of the distal segment of the nephron with subsequent inhibition of aldosterone-dependent regulatory mechanisms. Conclusions. The kaliuric reaction of the diabetic kidney may serve as one of the signs of decompensation of the renal blood flow autoregulation by tubuloglomerular feedback, which is an initiating factor for the dysfunction of the tubular apparatus of diabetic kidney.

Glucose/Fructose Delivery to the Distal Nephron Activates the Sodium-Chloride Cotransporter via the Calcium-Sensing Receptor.

The calcium-sensing receptor (CaSR) in the distal convoluted tubule (DCT) activates the NaCl cotransporter (NCC). Glucose acts as a positive allosteric modulator of the CaSR. Under physiologic conditions, no glucose is delivered to the DCT, and fructose delivery depends on consumption. We hypothesized that glucose/fructose delivery to the DCT modulates the CaSR in a positive allosteric way, activating the WNK4-SPAK-NCC pathway and thus increasing salt retention. We evaluated the effect of glucose/fructose arrival to the distal nephron on the CaSR-WNK4-SPAK-NCC pathway using HEK-293 cells, C57BL/6 and WNK4-knockout mice, ex vivo perfused kidneys, and healthy humans. HEK-293 cells exposed to glucose/fructose increased SPAK phosphorylation in a WNK4- and CaSR-dependent manner. C57BL/6 mice exposed to fructose or a single dose of dapagliflozin to induce transient glycosuria showed increased activity of the WNK4-SPAK-NCC pathway. The calcilytic NPS2143 ameliorated this effect, which was not observed in WNK4-KO mice. C57BL/6 mice treated with fructose or dapagliflozin showed markedly increased natriuresis after thiazide challenge. Ex vivo rat kidney perfused with glucose above the physiologic threshold levels for proximal reabsorption showed increased NCC and SPAK phosphorylation. NPS2143 prevented this effect. In healthy volunteers, cinacalcet administration, fructose intake, or a single dose of dapagliflozin increased SPAK and NCC phosphorylation in urinary extracellular vesicles. Glycosuria or fructosuria was associated with increased NCC, SPAK, and WNK4 phosphorylation in a CaSR-dependent manner.

20-OR: The Renal and Vascular Effects of Combined SGLT2 and Angiotensin Converting Enzyme Inhibition

The goal of this mechanistic trial was to determine the kidney and cardiovascular effects of combined treatment with an SGLT2i (empagliflozin 25mg QD) and an ACEi (ramipril 10mg QD) for 4 weeks in patients at risk of renal hyperfiltration. In this randomized, double-blind, placebo-controlled, cross-over trial, measurements were obtained following each of the 4 treatment phases: 1) no treatment, 2) 4-week ramipril treatment alone, 3) 4-week empagliflozin-ramipril combination treatment, and 4) 4-week placebo-ramipril combination treatment. The primary endpoint was GFR after empagliflozin-ramipril treatment compared to placebo-ramipril. At the end of each study phase the following measurements were performed under clamped euglycemia (4-6 mmol/L) : inulin (GFR) and paraaminohippurate (effective renal plasma flow, ERPF) clearances, tubular sodium handling, ambulatory blood pressure, arterial stiffness, heart rate variability, non-invasive cardiac output monitoring, plasma and urine biochemistry. Empagliflozin-ramipril treatment resulted in an 8ml/min/1.73m2 lower GFR (p=0.0061) , lower absolute proximal fluid reabsorption rate (p=0.0092) , lower absolute proximal sodium reabsorption rate (p=0.0056) , and lower urinary 8-isoprostane (p=0.0084) relative to placebo-ramipril treatment. Empagliflozin-ramipril treatment resulted in additive blood pressure lowering effects (SBP p=0.0112; DBP p=0.0032) and a lower total peripheral resistance (p=0.0368) . There were no other significant changes observed with the addition of empagliflozin. SGLT2i-ACEi combination treatment resulted in an expected GFR “dip”, suppression of oxidative stress markers, and additive declines in blood pressure and total peripheral resistance in this mechanistic study. These changes are consistent with a protective physiological profile characterized by the lowering of intraglomerular pressure and related cardiorenal risk when adding an SGLT2i to conservative therapy. Disclosure Y. Lytvyn: None. K. Kimura: Employee; 3Boehringer Ingelheim Canada Ltd./Ltée. N. Peter: Employee; Boehringer Ingelheim Pharma GmbH & Co.KG. V.S. Lai: Advisory Panel; Baxter. J. Tse: None. L. Cham: None. B.A. Perkins: Advisory Panel; Abbott Diabetes, Insulet Corporation, Sanofi. Board Member; Novo Nordisk. Research Support; BMO Bank of Montreal, Novo Nordisk. Other Relationship; Abbott Diabetes, Insulet Corporation, Medtronic, Novo Nordisk. N. Soleymanlou: Employee; Boehringer Ingelheim Pharmaceuticals Inc., Boehringer Ingelheim Pharmaceuticals Inc. D. Cherney: Research Support; Boehringer Ingelheim-Lilly, Merck, Janssen, Sanofi, AstraZeneca and Novo-Nordisk. Other Relationship; AbbVie Inc., AstraZeneca, Bayer AG, Boehringer Ingelheim International GmbH, Janssen Research & Development, LLC, Lilly, Maze, BMS, CSL-Behring, Merck, Otsuka, Novartis and Novo-Nordisk, Mitsubishi Tanabe Pharma Corporation, Sanofi. Funding Boehringer Ingelheim and CIHR, Diabetes Canada and the Heart and Stroke Richard Lewar Centre of Excellence and the Heart and Stroke Foundation of Canada

123-LB: Extended Fasting Reveals Effect of Canagliflozin to Mobilize Ketones in Canines

SGLT2 inhibitors (SGLT2i) improve glycemic control by inhibiting renal proximal tubular glucose reabsorption. SGLT2i may also elevate ketones. Fasting is expected to increase endogenous ketones, and SGLT2i may enhance ketosis. We asked if SGLT2i alters circulating ketones (β-hydroxybutyrate; BOHB) and disrupts glycemia during prolonged fasting. Dogs (n=7) were studied with a crossover design. Dogs received either canagliflozin (CANA; 300 mg) or placebo (PL) once/day for 7 d, after which a 48 h fast was performed while treatment was continued. To begin the fast, after morning feeding (9 am to noon) , remaining food was removed (“t=0”) . Blood was sampled at regular intervals over a 48 h period of total fasting (urine collected over final 3 h) . After the fast, dogs were fed over 3 h. Treatment was then discontinued for 2 wk washout, and dogs were then assigned to the alternate treatment (CANA or PL) for 7 d, after which fasting experiment was repeated. Glycemia was sustained in both treatment groups during initial 22 h of fasting, presumably due to reduction of insulin (↓50.7±13.1% and ↓49.5±21.5% for CANA and PL, respectively; p<0.035 for both) . But marked effects of CANA alone were revealed after 22 h of fasting, when glucose declined markedly in CANA (52.9±3.4 mg/dl at t=48 h) , but not PL (86.6±2.5 mg/dl at 48 h; p=0.0002 vs. CANA) , due to drug-induced glycosuria (13948±543 vs. 19.6±1.5 mg/dl; p<0.0001) . After 22 h of fasting, CANA induced ∼5-fold increase in FFA (p<0.03) and marked increase in plasma BOHB that persisted for duration of fast (CANA: 0.560±0.065 mM, PL: 0.048±0.006 mM; p<0.0001) . After refeeding, glucose and BOHB normalized, although BOHB remained higher in CANA (p<0.0007) . In conclusion, effect of SGLT2i on ketones is manifest after 24 h of total food restriction. CANA treatment induces glycosuria, after which reduction in carbohydrate stores switches animals to fat-based metabolism, characterized by increased FFA and ketosis. CANA appears to induce ketosis when carbohydrate stores are depleted. Disclosure M. Ader: None. R. Sedighi: None. M. Kabir: None. R. N. Bergman: Consultant; Fractyl Health, Inc., Novo Nordisk, Research Support; AstraZeneca, Janssen Research & Development, LLC. Funding Janssen Pharmaceutica, Inc.; NIH (DK29867, DK27619)

Influence of water of negative oxidation-reduction potential with hydrogen saturation on kidney function in rats

Objective. To study the effect of water load of negative redox potential with hydrogen saturation on renal function in comparison with induced diuresis with water of negative redox potential without hydrogen saturation.Material and methods. In experiments on 60 male white-nonlinear rats weighing 0.16-0.18 kg, the effect of loading with ordinary tap water (redox potential 90.4 ± 1.45 mV), water of negative redox potential without saturation with hydrogen (redox potential - 304.5 ± 4.79 mV) and water of negative redox potential with hydrogen saturation 1.0 - 1.2 ppm and redox potential -297.3 ± 5.27 mV. Used: experimental, physiological, biochemical, chemical, physicochemical, statistical research methods.Results. Under water load of negative redox potential with hydrogen saturation in comparison with induced diuresis of negative redox potential with the use of microhydrin, there was no inhibition of glomerular filtration, filtration fraction and proximal reabsorption of sodium ions, change restorative potential of urine. These changes are explained by the lack of activation of the dysregulatory mechanism of basal vascular tone of the renal cortex under conditions of water load of negative redox potential with hydrogen saturation.Conclusion. Water loading of negative redox potential with hydrogen saturation in comparison with induced diuresis of negative redox potential without hydrogen saturation causes recovery of glomerular filtration rate, filtration fraction and proximal reabsorption of sodium ions, due to the lack of activation of the mechanism of basal vascular tone of the renal cortical substance, due to the high permeability of molecular hydrogen and uniform delivery of electrons to all organs and tissues.

Pattern Recognition versus Pathogenesis: Electrolytes in a Patient with Adrenal Insufficiency.

Pattern Recognition versus Pathogenesis: Electrolytes in a Patient with Adrenal Insufficiency.

Abstract 10460: Urinary Fatty Acid Binding Protein 1 is a Biomarker for Impaired Proximal Tubular Protein Reabsorption and is Synergistically Enhanced by Concurrent Liver Injury

Introduction: Urinary fatty acid binding protein 1 (FABP1, also known as liver-type FABP) has been implicated as a biomarker of acute kidney injury (AKI) in humans. However, the precise biological mechanisms underlying its elevation remain elusive. Here, we show that urinary FABP1 primarily reflects impaired protein reabsorption in proximal tubule epithelial cells (PTECs). Methods and Results: Bilateral nephrectomy resulted in a marked increase in serum FABP1 levels, suggesting that the kidney is an essential organ for removing serum FABP1. Injected recombinant FABP1 was filtered through the glomeruli and robustly reabsorbed via the apical membrane of PTECs. Urinary FABP1 was significantly elevated in mice devoid of megalin, a giant endocytic receptor for protein reabsorption. Elevation of urinary FABP1 was also observed in patients with Dent disease, a rare genetic disease characterized by defective megalin function in PTECs. Urinary FABP1 levels were exponentially increased following acetaminophen overdose, with both nephrotoxicity and hepatotoxicity observed. FABP1-deficient mice with liver-specific overexpression of FABP1 showed a massive increase in urinary FABP1 levels upon acetaminophen injection, indicating that urinary FABP1 is liver-derived. Lastly, we employed transgenic mice expressing diphtheria toxin receptor (DT-R) either in a hepatocyte- or PTEC-specific manner, or both. Upon diphtheria toxin (DT) administration, massive excretion of urinary FABP1 was induced in mice with both kidney and liver injury, while mice with either injury type showed marginal excretion. Conclusions: Collectively, our data demonstrated that intact PTECs have a considerable capacity to reabsorb liver-derived FABP1 through a megalin-mediated mechanism. Thus, urinary FABP1, which is synergistically enhanced by concurrent liver injury, is a biomarker for impaired protein reabsorption in AKI. These findings address the use of urinary FABP1 as a biomarker of histologically injured PTECs that secrete FABP1 into primary urine, and suggest the use of this biomarker to simultaneously monitor impaired tubular reabsorption and liver function.

Current Challenges and Future Perspectives of Renal Tubular Dysfunction in Diabetic Kidney Disease.

Over decades, substantial progress has been achieved in understanding the pathogenesis of proteinuria in diabetic kidney disease (DKD), biomarkers for DKD screening, diagnosis, and prognosis, as well as novel hypoglycemia agents in clinical trials, thereby rendering more attention focused on the role of renal tubules in DKD. Previous studies have demonstrated that morphological and functional changes in renal tubules are highly involved in the occurrence and development of DKD. Novel tubular biomarkers have shown some clinical importance. However, there are many challenges to transition into personalized diagnosis and guidance for individual therapy in clinical practice. Large-scale clinical trials suggested the clinical relevance of increased proximal reabsorption and hyperfiltration by sodium-glucose cotransporter-2 (SGLT2) to improve renal outcomes in patients with diabetes, further promoting the emergence of renal tubulocentric research. Therefore, this review summarized the recent progress in the pathophysiology associated with involved mechanisms of renal tubules, potential tubular biomarkers with clinical application, and renal tubular factors in DKD management. The mechanism of kidney protection and impressive results from clinical trials of SGLT2 inhibitors were summarized and discussed, offering a comprehensive update on therapeutic strategies targeting renal tubules.

Pathogenetical aspects of tubulointerstitial syndrome development in alloxan-induced experimental diabetes mellitus

Annotation. The aim of our study was to explore the pathogenetical aspects of tubulointerstitial syndrome development in alloxan-induced experimental diabetes mellitus. The experiments were carried out on 20 white non-linear mature male rats, 10 with experimental diabetes mellitus (EDМ) induced by intraperitoneal administration of alloxan at a dose of 160 mg/kg of body weight, 10 intact rats served as the control group. 25 days after administration of the diabetogenic substance, the animals were withdrawn from the experiment. The concentration of sodium and potassium ions in urine and blood plasma samples was determined, followed by calculation (considering water-induced 2-hour diuresis and endogenous creatinine clearance) of glomerular filtration rate, electrolyte excretion, their filtration rate, absolute and relative reabsorption, clearance, their proximal and distal renal transport. Removed after decapitation rats’ kidneys were dissected to 3 parts – renal cortex, medulla and papilla, sodium and potassium content was determined in water-extract of the corresponding part of the renal parenchyma, and papillary-cortical, papillary-medullar and medullary-cortical concentration ion gradients were calculated. Significant suppression of papillary-medullar and papillary-cortical concentration sodium gradients, as well as a slight limitation of its medullary-cortical gradient were established. The concentration potassium gradients were significantly reduced. Statistical processing of the obtained data was carried out with the determination of the average value, standard deviations. To assess the probability of the difference between the study groups used non-parametric Mann-Whitney ranking criterion according to the algorithms implemented in the computer program “Statistica for Windows”, “Version 8.0”. There was a decrease of the sodium-potassium ratio in urine, enhanced urinary excretion of potassium and an increase of its content in urine, as well as intensification of absolute transtubular sodium transport due to equivalent augmentation of the filtration charge of this cation, increase of proximal sodium reabsorption and, to a lesser degree, – of distal one. The distal and proximal sodium reabsorption, reduced to a unit of active nephrons, was found to be decreased, and the relative reabsorption of the cation significantly exceeded the control values, contributing to the limitation of natriuresis. The results of the study suggest that in 26-day alloxan-induced experimental diabetes hemodynamic-hyperperfusion overload on the tubular apparatus of the kidney causes the development of relative insufficiency of the proximal and distal tubules, disorders of hormone-dependent reabsorption of cations, limitation of regulatory influence of aldosterone and ADH with further tubulointerstitial disturbances that unable adequate osmotic concentration of urine.

Disorders of Acid-Base and Potassium Balance

Water accounts for approximately half of an adult human's body weight. Two thirds of body water is intracellular, and the remaining one third is contained in the extracellular fluid compartment, which includes intravascular (plasma) and interstitial fluid. Small amounts of water are also contained in bone, dense connective tissue, digestive secretions, and cerebrospinal fluid. To maintain stability of the internal milieu, body fluids are processed by the kidney, guided by intricate physiologic control systems that regulate fluid volume and composition. This chapter reviews the principles of body fluid hemostasis and discusses disorders of water excess (hyponatremia), water deficiency (hypernatremia), water conservation (diabetes insipidus), saltwater excess (edematous states), and saltwater deficiency (volume depletion). Figures illustrate the relation between the plasma hydrogen ion concentration and the blood pH, proximal tubular bicarbonate reabsorption, renal secretion of hydrogen ions, renal glutamine metabolism and ammonia diffusion, metabolic alkalosis, and the electrocardiographic changes in and approach to causes of hypokalemia and hyperkalemia. Tables describe causes of metabolic acidosis with a high and a normal ion gap; causes of type 1 renal tubular acidosis; causes of type 4 renal tubular acidosis and aldosterone resistance; causes of metabolic alkalosis, respiratory acidosis, and respiratory alkalosis; evaluation of the renal defect in hyperkalemia; and treatment of hyperkalemia. This chapter contains 10 figures, 12 tables, and 53 references.

Dysregulation mechanism of the influence of water of negative redox potential on kidney function

Elucidate the dysregulatory mechanism of the effect of water load of negative redox potential in comparison with induced diuresis with ordinary tap water on the functional state of the kidneys.Material and methods. The experiments were carried out on 40 male white nonlinear rats weighing 0.16-0.18 kg with the analysis of the effect of water loading with a negative redox potential on renal function, which was obtained by treating tap water with microhydrin. The redox potential of water and urine was determined with an ORP meter. From the indicators of renal function studied: protein excretion, glomerular filtration, filtration fraction of sodium ions, its absolute and proximal reabsorption. Statistical processing of the data obtained, including multivariate regression analysis, was carried out using the Statgrafics, Statistica, and Excel 2003 programs.Results. Comparative multivariate analysis of the effect of water loading of negative redox potential in comparison with induced diuresis with ordinary tap water revealed an ambiguous nature of reliable correlations (p <0.05) between the studied indicators of renal function and the redox potential of urine, which is due to the simultaneous presence of both protective mechanisms of negative redox potential influence (improvement of energy supply, antioxidant effect), and damaging (transient renal ischemia due to activation of the basal tone mechanism due to an increase in the contractile function of the heart as a result of improving its energy supply), the final result will depend on the dominance of classical or dysregulation mechanisms in this particular situation. Conclusion. The dysregulation mechanism of the effect of water of negative redox potential on renal function is due to the initial mobilization of defense reactions with improved contractile function of the heart, which is subsequently transformed into an injury response due to increased basal renal vascular tone with the development of transient ischemia and renal dysfunction, decrease оf glomerular filtration, filtration fraction of sodium ions, its absolute and proximal reabsorption and development of proteinuria.

The tubular hypothesis of nephron filtration and diabetic kidney disease.

Kidney size and glomerular filtration rate (GFR) often increase with the onset of diabetes, and elevated GFR is a risk factor for the development of diabetic kidney disease. Hyperfiltration mainly occurs in response to signals passed from the tubule to the glomerulus: high levels of glucose in the glomerular filtrate drive increased reabsorption of glucose and sodium by the sodium-glucose cotransporters SGLT2 and SGLT1 in the proximal tubule. Passive reabsorption of chloride and water also increases. The overall capacity for proximal reabsorption is augmented by growth of the proximal tubule, which (alongside sodium-glucose cotransport) further limits urinary glucose loss. Hyperreabsorption of sodium and chloride induces tubuloglomerular feedback from the macula densa to increase GFR. In addition, sodium-glucose cotransport by SGLT1 on macula densa cells triggers the production of nitric oxide, which also contributes to glomerular hyperfiltration. Although hyperfiltration restores sodium and chloride excretion it imposes added physical stress on the filtration barrier and increases the oxygen demand to drive reabsorption. Tubular growth is associated with the development of a senescence-like molecular signature that sets the stage for inflammation and fibrosis. SGLT2 inhibitors attenuate the proximal reabsorption of sodium and glucose, normalize tubuloglomerular feedback signals and mitigate hyperfiltration. This tubule-centred model of diabetic kidney physiology predicts the salutary effect of SGLT2 inhibitors on hard renal outcomes, as shown in large-scale clinical trials.

Principles of Evolution of the Excretory Organs and the System of Homeostasis

The function of the excretory organs in metazoan invertebrates and vertebrates is aimed at maintaining homeostasis. Two patterns of the morpho-functional organization of these organs can be distinguished: I—a combination of ultrafiltration and subsequent reabsorption with partial secretion of substances, or II—only a secretion of molecules of a certain type. Here we substantiate the principle according to which in the type I organs the key role of the proximal tubule is to reabsorb fluid, which is ideal in terms of its solute composition and concentration, and to secrete a number of organic acids and bases, while the distal tubule is responsible for adjusting the composition of inorganic substances. Everything what does not meet these criteria is to be excreted. The type II organs (salt glands, aglomerular kidneys) are inherent to those animals whose kidneys do not ensure osmoregulation. During evolution, the mammalian kidney developed the mechanism which regulates the redistribution of fluid within the nephron: altered reabsorption of ions and water in the proximal segment can cause an influx of some additional fluid to the distal segment where the regulatory systems adjust the amount of reabsorbed substances. The central tendency in the evolution of the kidney in vertebrates, including humans, is to increase the rates of glomerular filtration and proximal reabsorption. A similarity between molecular mechanisms of transmembrane and transepithelial transport of substances has been revealed in the evolution of excretory organs, with distinctions concerning mainly the structure of the regulatory molecules and the intensity of urine formation.

Abstract 080: Dietary Fructose Enhances Protein Kinase C Activation by Angiotensin II in Proximal Tubules via Changes in Intracellular Calcium

Dietary fructose causes salt-sensitive hypertension. This is in part due to increasing the sensitivity of proximal nephron Na reabsorption to angiotensin II (Ang II) such that lower concentrations stimulate transport to a greater extent. Ang II stimulates Na transport in this segment by increasing protein kinase C (PKC) α, a calcium- and lipid-dependent kinase. We hypothesized that dietary fructose increases the ability of Ang II to elevate intracellular calcium (Cai) and, thereby, activate PKC α in proximal tubules. This, in turn, stimulates Na/H exchange activity, the primary transporter involved in Na reabsorption. To test our hypothesis rats were maintained on a diet of normal chow plus tap water or normal chow plus 20% fructose. The effect of Ang II on Cai was measured using Fura2 in perfused S2 segments. Na/H exchange (NHE) was measured in perfused tubules using the pH-sensitive dye BCECF. PKC α activity was measured by separating particulate and soluble fractions, performing Western blots and recording the particulate to soluble ratio. Basal Cai was 143±29 nM in proximal tubules from control rats while it was 160±30 nM in those given fructose, not significantly different. Ang II (1 nM) increased Cai by 43±10 nM in control tubules and by 148±53 nM in tubules from rats fed fructose (p < 0.03). A higher concentration of Ang II (100 nM) had a similar effect in tubules from both groups (Fructose: 237±100 nM vs. Control: 190±34 nM). Ang II increased the particulate to soluble ratio of PKC α, a measure of activation, by 0.134±0.026 in tubules from rats fed fructose (p <0.001) but not significantly in control tubules (0.060±0.061). Finally we measured NHE activity. Ang II (1 pM) increased NHE activity by 0.7±0.1 fluorescent units/s in tubules from rats given fructose but had no effect on NHE activity in control tubules (p<0.01). With Go6976, a PKC α/β1 inhibitor, Ang II was unable to stimulate NHE activity in tubules from rats fed fructose. We conclude that dietary fructose increases the ability of Ang II to elevate Cai, and consequently PKCα. This, in turn, stimulates NHE activity which likely contributes to fructose-induced salt-sensitivity of blood pressure.

Abstract P2038: A Nobel I551F Mutant of Sodium Handling Transporter NBCe1 Has Cytosolic Retention and Diminishes Transport Activity

The electrogenic Na + /HCO 3 - cotransporter NBCe1 is expressed in many organs and controls sodium handling. NBCe1 homozygous mutation causes not only severe hypotension but also proximal renal tubular acidosis in human and rodents. Here we shows the functional significance of the mutations reported from NCBI database on NBCe1 function. We identified 14 mutations in NBCe1 variant A(kidney type), resulting in the single amino acid substitutions, A425T, F461L, A465S, L494F, N503H, A518T, A518G, I523V, V533D, Y535H, I551F, Y554H, M753V, and L785I. Immunocytological analysis with confocal microscopy revealed only I551F mutant was expressed predominantly in the cytoplasm of HEK 293 cells and MDCK cells. Functional analysis using Xenopus oocytes revealed that I551F mutant had a significantly reduced activity corresponding to 52% of that of wild-type (p<0.01). Whole and biotinylated western blotting in HEK293 cells confirmed that the surface expression of I551F mutant was significantly reduced. Deglycosylation study revealed that I551F has impaired N-glycosylation. To examine the role of I551 in more detail, we also examined the properties of artificial mutants I551A, I551P, I551K, I551R, I551D and I551E in HEK293 and MDCK cells. All of artificial mutants were properly expressed in the plasma membrane. By contrast, only I551F mutant was predominantly expressed in cytoplasmic regions. These results indicate that I551F mutation inactivates the NBCe1 function with cytoplasmic retention. Because NBCe1 plays a major role in renal proximal sodium and bicarbonate reabsorption, I551F mutation may be associated with the disturbance in systemic acid-base balance or the changes in blood pressure.

Role of carbonic anhydrase in acute recovery following renal ischemia reperfusion injury.

Ischemia reperfusion (IR) injury can cause acute kidney injury. It has previously been reported that kidney oxygen consumption (QO2) in relation to glomerular filtration rate (GFR), and thus tubular sodium load, is markedly increased following IR injury, indicating reduced electrolyte transport efficiency. Since proximal tubular sodium reabsorption (TNa) is a major contributor to overall kidney QO2, we investigated whether inhibition of proximal tubular sodium transport through carbonic anhydrase (CA) inhibition would improve renal oxygenation following ischemia reperfusion. Anesthetized adult male Sprague Dawley rats were administered the CA inhibitor acetazolamide (50 mg/kg bolus iv), or volume-matched vehicle, and kidney function, hemodynamics and QO2 were estimated before and after 45 minutes of unilateral complete warm renal ischemia. CA inhibition per se reduced GFR (-20%) and TNa (-22%), while it increased urine flow and urinary sodium excretion (36-fold). Renal blood flow was reduced (-31%) due to increased renal vascular resistance (+37%) without affecting QO2. IR per se resulted in similar decrease in GFR and TNa, independently of CA activity. However, the QO2/TNa ratio following ischemia-reperfusion was profoundly increased in the group receiving CA inhibition, indicating a significant contribution of basal oxygen metabolism to the total kidney QO2 following inhibition of proximal tubular function after IR injury. Ischemia increased urinary excretion of kidney injury molecule-1, an effect that was unaffected by CA. In conclusion, this study demonstrates that CA inhibition further impairs renal oxygenation and does not protect tubular function in the acute phase following IR injury. Furthermore, these results indicate a major role of the proximal tubule in the acute recovery from an ischemic insult.

Role of sodium/glucose cotransporter inhibition on a rat model of angiotensin II\u2013dependent kidney damage

BackgroundRenal proximal tubular sodium and glucose reabsorption are regulated by the sodium-glucose cotransporter (SGLT2). Changes in this transporter can play a role in hyperglycaemia and reactive oxygen species (ROS) production. We demonstrated increased glucose absorption in proximal tubule membrane vesicles and increased expression of SGLT2 in hypertensive rats. Here we investigated Angiotensin II (Ang II) -dependent SGLT2 expression induction and the role of SGLT2 induction in the development of Ang II-dependent kidney damage. The aim of this study was to determine whether SGLT2 induction by Ang II is associated with Ang II-dependent kidney damage. We propose the following objectives a) to demonstrate that Ang II induces SGLT2 expression and b) to demonstrate that prevention of SGLT2 expression and activity prevent Ang II-induced kidney damage.MethodsWe used chronic Ang II infusion as a model of kidney damage in male Wistar rats and evaluated systolic blood pressure by telemetric methods. SGLT2 mRNA and protein expression were evaluated by PCR and immunoblotting. SGLT2 activity was evaluated in brush border membrane vesicles by measuring glucose uptake. ROS production was measured by confocal microscopy. The glomerular filtration rate (GFR) was evaluated by the inulin excretion method, and urinary protein excretion was evaluated by the Bradford method. Biological parameter evaluations were performed, after two weeks of infusion of Ang II. We compared the effects of Angiotensin II (AT1) receptor blockade by Losartan and SGLT2 inhibition by Empagliflozin both as monotherapy treatments and in combination on the development of kidney damage.ResultsChronic Ang II infusion led to a blood pressure elevation and increased SGLT2 mRNA expression and activity as well as kidney damage, as reflected by increased ROS production, decreased GFR and increased urinary protein excretion. AT1 receptor blockade prevented all these changes. By contrast, SGLT2 inhibition did not affect blood pressure and had a small effect on kidney damage. However, the combination of both drugs resulted in the potentiation of the effects observed by AT1 receptor blockade alone.ConclusionsWe suggest that Ang II-dependent increased SGLT2 induction is one mechanism by which Ang II induces kidney damage.

THE ROLE OF CYTOKINES IN THE KIDNEY DYSFUNCTION UNDER CONDITIONS OF IRRITABLE BOWELS SYNDROME DEVELOPMENT

The research is dedicated to the elucidation of the cytokines’ role in the renal dysfunction under conditions of irritable bowels syndrome.Objective - to carry out pathological analysis of cytokines’ role of the blood plasma in the kidney dysfunction at irritable bowels syndrome with constipation and diarrhea.Material and methods. 60 patients (18 – men, 42 – women aged 28 to 62 years) with irritable bowels syndrome were examined. Among them there were 28 patients with irritable bowels syndrome with constipation, 32 patients had irritable bowels syndrome with diarrhea. Control group consisted of 25 practically healthy patients. The level of cytokines was determined in the blood plasma: necrosis factor of tumor-alpha, interleukin- 1-beta, interleukin-4 and interleukin-6. Statistical data processing was conducted by means of computer programs “Statgrafics” and “Exel 7.0”.Results. Contents increase of necrosis factor of tumor-alpha, interleukin-1-beta, interleukine-4 and interleukine-6 is accompanied by inhibition of the total, enzymatic, non-enzymatic fibrinolytic activity of the urine and proximal reabsorption of sodium ions in patients with irritable bowel syndrome.Conclusions. More significant imbalance manifestations of cytokines and functional state of kidneys revealed in patients with the syndrome of irritable bowels with diarrhea in comparison with the previously mentioned syndrome with constipation are explained by the development of dehydration, hemoconcentration, hypoxia, lipid peroxidation activation and more essential damageable manifestations of inflammation process due to diarrhea.

The role of abdominal microflora of the colon in the impaired renal function in the development of irritable bowel syndrome

Objective: to conduct population-level pathophysiological analysis of aerobic, anaerobic cavitary microflora of the large intestine with irritable bowel syndrome with constipation and diarrhea with the analysis of the functional state of the kidneys.Material and methods. The study included 60 patients with irritable bowel syndrome. Men — 18 patients, women — 42 patients from 28 to 62 years of age, out of which: with irritable bowel syndrome with constipation comprised 28 patients, and with irritable bowel syndrome with diarrhea — 32. The control group consisted of 25 healthy patients. We determined the population levels of aerobic (S. aureus, E. fecalis, E. coli, P. vulgaris, K. pneumoniae), anaerobic (B. Bifidum, B. lactis), evaluated the functional state of the kidneys, the general condition of patients on Likert scale. Statistical data processing was performed using the computer programs “Statgrafics” and “Excel 7.0”. Results. In patients with irritable bowel syndrome, the increase in the content of cavitary microflora of the large intestine E. Coli, S. aureus, P. vulgaris, K. pneumoniae, E. faecalis under decreased B. Bifidum, B. Lactis is accompanied by violation of kidney function with the development of tubular proteinuria and inhibition of proximal reabsorption of sodium ions.Conclusion. It has been revealed more significant manifestations of dysbacteriosis, deterioration of the general condition of patients according to Likert scale and the functional state of kidneys in patients with irritable bowel syndrome with diarrhea in comparison with the above syndrome with constipation due to the development of dehydration under the conditions of diarrhea.

Acid-Base Disorders

Under physiologic conditions, the acid-base balance of the body is maintained via changes in ventilation that eliminate carbon dioxide, buffering of acid loads, and renal excretion of hydrogen ions. Failure to maintain the pH of the blood between 7.35 and 7.45 can result in life-threatening conditions. This review details the pathophysiology, stabilization and assessment, diagnosis and treatment, and disposition and outcomes of acid-base disorders. Figures show the relationship between hydrogen ions and blood pH, proximal tubular bicarbonate reabsorption, the secretion of hydrogen ions, renal ammonia production, ammonium diffusion, metabolic alkalosis, electrocardiographic changes in hypokalemia and hyperkalemia, pseudoinfarction caused by hyperkalemia, and an algorithmic approach to suspected acid-base disorders. Tables list causes of high–anion gap metabolic acidosis, metabolic acidosis with a normal anion gap, type 1 renal tubular acidosis, type 4 renal tubular acidosis and aldosterone resistance, metabolic alkalosis, respiratory acidosis, and respiratory alkalosis; treatment of hyperkalemia; and a stepwise approach for the evaluation of suspected acid-base disorders. This review contains 9 highly rendered figures, 9 tables, 64 references, and a list of pertinent Web sites.