Calcium Oxalate Nephrolithiasis Research Articles

A Simple, Low-Cost, and Useful Preconcentration Method for Quantification of Soluble, Insoluble, and Total Oxalate in Selected Vegetables Through Spectrophotometry

The amount of dietary oxalate from food may be an important risk factor in the development of idiopathic calcium oxalate nephrolithiasis. The main aim of this research was to develop an accurate and reliable method by cloud point extraction (CPE) combined with spectrophotometry to measure the oxalate contents of selected vegetables. The method is based on ion association of stable anionic complex, which is produced by the reaction of oxalate with Mo(VI), with Toluidine blue (TBH2+), and then by extraction into micelles of octylphenol ethoxylate (Triton X-45) in the presence of NH4F as a salting-out agent at pH 6.0. Using the optimized conditions, the calibration graph was highly linear in the range of 1.2–240 μg L−1. The detection limit of the method (LOD (3σblank/m) was 0.36 μg L−1, and the relative standard deviation (RSD %) as a measure of precision was in the range of 1.1–5.3 % (n = 5 for 5, 25, and 50 μg L−1). The method was successfully applied to the speciative determination of soluble and total oxalate in vegetables after ultrasonic-assisted extraction and dilution at suitable ratios. The amount of insoluble oxalate was calculated from the analytical difference between total oxalate and soluble oxalate contents of samples with and without acidic extraction under ultrasonic power (300 W, 50 Hz) for 15 min at 60 °C. The accuracy of the method was intrinsically controlled and verified by comparing the results obtained with those of an independent kinetic method as well as recoveries of spiked samples.

Soft Tissue Metastasis of Parathyroid Carcinoma: Description of a Difficult Case

Parathyroid carcinoma is an extremely rare cause of hyperparathyroidism. Histologic diagnosis may be challenging. DAB, male aged 66 was submitted to subtotal gastrectomy because of a localized gastric adenocarcinoma (intestinal type). A mild primary hyperparathyroidism was simultaneously found. Calcium oxalate nephrolithiasis was diagnosed 16 years before. Sonography revealed a nodular lesion in the right lower thyroid pole, while NMR showed a nodular lesion at the left lower thyroid pole. The sestamibi scan found a late image hyperfixation at the right lower pole. Bilateral inferior parathy roidectomy was performed and revealed on both sides adenoma and moderate nuclear polymorphism and no mitotic figures; on the left side a central cystic area was found. Clinical and biochemical cure apparently followed. Six months later a presumed unrelated nodular lesion in the right thigh muscular mass was removed and found to have numerous mitotic figures and immunoreactivity for PTH. A sestamibi scan showed marked hyperfixation in the right leg and PET-FDG-F 18 hyperfixated the right quadricipital area. There is evidence for a long standing disease since nephrolithiasis was found some years before. Long standing hyperparathyroidism and increased gastric acid secretion can lead to chronic gastritis, intestinal metaplasia and gastric adenocarcinoma. There is evidence for evolution along bilateral hyperplasia, adenoma and carcinoma since bilateral disease was present and left cystic nonfunctioning lesion was found. Most surprisingly a single metastatic lesion was later found in a very unusual location and this lesion was highly undifferentiated in sharp contrast with the primitive lesion.

Study on inhibitory effect of EGCG on Calcium oxalate nephrolithiasis in rats and its related mechanism

In the study, the inhibitory effect of epigallocatechin gallate (EGCG) on Calcium oxalate nephrolithiasis and its possible mechanism were investigated. The rat Calcium oxalate nephrolithiasis model was induced through the combined oral administration of ethylene glycol and ammonium chloride, which was intervened with EGCG. Rat blood samples were collected to detect blood creatinine (Cr), blood urea nitrogen (BUN) and blood calcium. Rat urine samples were collected to observe and compare 24-hour urine volume, oxalic acid (Ox) and calcium in urine. Renal samples were collected to prepare tissue slices and observe the pathological changes in Calcium oxalate nephrolithiasis. The expression of osteopontin (OPN) in renal tissues was evaluated by Real-time PCR and Western blot. According to the results, compared with normal rats, rats in the nephrolithiasis model showed significant increases in Cr, BUN, urine Calcium, urine Ox and renal OPN expression (P < 0.05), but obvious decrease in 24-hour urine volume (P < 0.05); Compared with rats with nephrolithiasis, those processed with EGCG revealed remarkable declines in Cr, BUN, urine Calcium and urine Ox (P < 0.05), with significant rise in 24-hour urine volume (P < 0.05) in a concentration-dependent manner. Additionally, compared with the control group, nephrolithiasis rats showed significant pathological changes in Calcium oxalate calculus. After ECCG treatment, the renal pathological changes and OPN expression attenuated significantly in a concentration-dependent manner. The results showed that EGCG inhibits the formation of calcium oxalate nephrolithiasis in rats and shows a notable protective effect on renal functions.

Efficacy of Mixtures of Magnesium, Citrate and Phytate as Calcium Oxalate Crystallization Inhibitors in Urine

The main aim of the current study was to evaluate the effectiveness of mixtures of magnesium, citrate and phytate as calcium oxalate crystallization inhibitors. A turbidimetric assay in synthetic urine was performed to obtain induction times for calcium oxalate crystallization in the absence and presence of different mixtures of inhibitors. The morphology of calcium oxalate crystals in the absence or presence of inhibitors and mixtures of the inhibitors was evaluated in 2 crystallization experiments at low and high calcium oxalate supersaturation. The crystals formed were examined using scanning electron microscopy. Examination of crystallization induction times revealed clear inhibitory effects of magnesium, citrate and phytate on calcium oxalate crystallization, supporting usefulness in the treatment and prevention of calcium oxalate nephrolithiasis. Significant synergistic effects between magnesium and phytate were observed. Scanning electron microscopy images revealed that phytate is a powerful crystal growth inhibitor of calcium oxalate, totally preventing the formation of trihydrate and monohydrate. In addition to crystallization inhibition capacity, citrate and magnesium avoided calcium oxalate crystallization by decreasing its supersaturation. The synergistic effect between magnesium and phytate on calcium oxalate crystallization suggests that a combination of these 2 compounds may be highly useful as antilithiasis therapy.

Protective effects of the aqueous extract of Crocus sativus against ethylene glycol induced nephrolithiasis in rats.

This study evaluated the possible protective effect of Crocus sativus L. (saffron) in the treatment of renal calculi. Aqueous extract of saffron (25, 50 and 100 mg/kg, daily) was administered intraperitoneally in two regimens of protective or curative, using male Wistar rats. Urolithiasis was induced by ethylene glycol (% 0.75) in drinking water. Urine was collected for biochemical analysis and the kidneys were prepared for total lipid peroxide and histological evaluation. Ethylene glycol feeding resulted in an increased urine output, renal excretion of oxalate and decreased excretion of citrate and magnesium. Saffron did not show diuretic effect; however, it significantly reduced the elevated urinary oxalate in prophylactic (50 and 100 mg/kg) and curative (100 mg/kg) studies. Only the high dose of prophylactic regimen restored citrate concentration of urine. Increased number of calcium oxalate crystals deposits in the kidney tissue of calculogenic rats was significantly reverted by the prophylactic and high dose of curative saffron treatment. Malondialdehyde (MDA, a lipid peroxidation product) in the kidneys was increased following the lithogenic treatment; however, prophylactic (50, 100 mg/kg) and curative (100 mg/kg) regimens with saffron reduced the elevated levels of MDA. Results in the current study indicate that saffron can protect against ethylene glycol induced calcium oxalate (CaOx) nephrolithiasis. The mechanisms underlying this effect are mediated possibly through effect on the urinary concentration of stone-forming constituents and an antioxidant effect.

Soft Tissue Metastasis of Parathyroid Carcinoma: Description of a Difficult Case

Parathyroid carcinoma is an extremely rare cause of hyperparathyroidism. Histologic diagnosis may be challenging. DAB, male aged 66 was submitted to subtotal gastrectomy because of a localized gastric adenocarcinoma (intestinal type). A mild primary hyperparathyroidism was simultaneously found. Calcium oxalate nephrolithiasis was diagnosed 16 years before. Sonography revealed a nodular lesion in the right lower thyroid pole, while NMR showed a nodular lesion at the left lower thyroid pole. The sestamibi scan found a late image hyperfixation at the right lower pole. Bilateral inferior parathyroidectomy was performed and revealed on both sides adenoma and moderate nuclear polymorphism and no mitotic figures; on the left side a central cystic area was found. Clinical and biochemical cure apparently followed. Six months later a presumed unrelated nodular lesion in the right thigh muscular mass was removed and found to have numerous mitotic figures and immunoreactivity for PTH. A sestamibi scan showed marked hyperfixation in the right leg and PET-FDG-F 18 hyperfixated the right quadricipital area. There is evidence for a long standing disease since nephrolithiasis was found some years before. Long standing hyperparathyroidism and increased gastric acid secretion can lead to chronic gastritis, intestinal metaplasia and gastric adenocarcinoma. There is evidence for evolution along bilateral hyperplasia, adenoma and carcinoma since bilateral disease was present and left cystic nonfunctioning lesion was found. Most surprisingly a single metastatic lesion was later found in a very unusual location and this lesion was highly undifferentiated in sharp contrast with the primitive lesion. J Endocrinol Metab. 2015;5(1-2):184-188 doi: http://dx.doi.org/10.14740/jem240w

Reactive oxygen species may unite many mechanisms by which calcium oxalate stones form.

The pathogenesis of calcium oxalate nephrolithiasis remains a mystery, but the suggestion that it is simply due to perturbations in urinary super saturations remains an inadequate explanation (1). It is likely due to much more complex and nuanced mechanisms that incorporate inorganic and organic components. How these components propagate into Randall plaques or calculi or even where these stone-forming events occur (vasa recta, collecting ducts, or the basement membrane of the loops of Henle) is debatable. Metabolic derangements leading to uncontrolled reactive oxygen species (ROS) generation or a reduced antioxidant capacity to alleviate oxidative stresses may play a role in Randall plaque formation through tissue damage and/or ROS-induced altered gene expression.

The inhibitory effect of an ethanol extract of the spores of Lygodium japonicum on ethylene glycol-induced kidney calculi in rats.

We investigated the effect of an ethanol extract of Lygodii spora (LS) as a preventive and therapeutic agent for experimentally induced calcium oxalate nephrolithiasis with ethylene glycol (EG) in rats. Male Wistar rats were randomly divided into preventive (n = 18, for 28 days) and therapeutic (n = 24, for 42 days) groups. The preventive group was further subdivided into three groups of six rats each: preventive control, preventive lithiatic control (EG) and preventive lithiatic LS (EG + 400 mg/kg LS). Similarly, the therapeutic group was subdivided into four groups of six rats each: therapeutic control, therapeutic lithiatic control, therapeutic lithiatic untreated, and therapeutic lithiatic LS. Lithiasis was induced by adding 0.75% EG to the drinking water of all groups except the preventive and therapeutic control groups. Preventive and therapeutic subjects also received the LS ethanol extract in drinking water at a dose of 400 mg/kg, since day 0 or day 28, respectively. At the end of the each experimental period, various biochemical parameters were measured in urine and kidney homogenates. The kidneys were subjected to histopathological analysis. The results revealed that treatment with the LS preventive protocol significantly decreased the levels of urinary calcium, oxalate and uric acid, and increased the levels of urinary citrate as compared to those in the EG control. No significant changes in the urinary parameters except oxalate and citrate levels were observed in the rats in the therapeutic protocol. In both preventive and therapeutic protocols, the extract significantly decreased kidney peroxides, renal calcium, oxalate content, and the number of kidney oxalate deposits as compared to those in the EG group. We conclude that LS is useful as a preventive and therapeutic agent against the formation of oxalate kidney stones.

Calcium oxalate nephrolithiasis and tubular necrosis in recent metamorphs of Rana sylvatica (Lithobates sylvaticus) fed spinach during the premetamorphic (tadpole) stage.

Amphibians in the family Ranidae (true frogs) seem highly susceptible to oxalosis, particularly when fed a diet high in oxalic acid during the premetamorphic (tadpole) stage. The authors describe the mortality of 150 captive-raised wood frogs (Rana sylvatica or Lithobates sylvaticus) from oxalate nephrolithiasis and renal tubular necrosis caused by consumption of boiled spinach during tadpole development. Renal lesions were due to intraluminal transparent crystals which were birefringent under polarized light and were identified morphologically and histochemically as composed of calcium oxalate. Evidence of early fibrosis or squamous metaplasia, and a presentation at least 2 weeks after spinach consumption had ended, suggested a subacute course. Tadpole-feeding protocols should avoid plants with high oxalate content (eg, spinach and rhubarb leaves), and any episode of high mortality in captive amphibians along with nephrolithiasis should prompt an evaluation of the feed sources for material with high oxalate content.

Osteopontin knockdown in the kidneys of hyperoxaluric rats leads to reduction in renal calcium oxalate crystal deposition

Osteopontin (OPN) expression is increased in kidneys of rats with ethylene glycol (EG) induced hyperoxaluria and calcium oxalate (CaOx) nephrolithiasis. The aim of this study is to clarify the effect of OPN knockdown by in vivo transfection of OPN siRNA on deposition of CaOx crystals in the kidneys. Hyperoxaluria was induced in 6-week-old male Sprague-Dawley rats by administering 1.5% EG in drinking water for 2 weeks. Four groups of six rats each were studied: Group A, untreated animals (tap water); Group B, administering 1.5% EG; Group C, 1.5% EG with in vivo transfection of OPN siRNA; Group D, 1.5% EG with in vivo transfection of negative control siRNA. OPN siRNA transfections were performed on day 1 and 8 by renal sub-capsular injection. Rats were killed at day 15 and kidneys were removed. Extent of crystal deposition was determined by measuring renal calcium concentrations and counting renal crystal deposits. OPN siRNA transfection resulted in significant reduction in expression of OPN mRNA as well as protein in group C compared to group B. Reduction in OPN expression was associated with significant decrease in crystal deposition in group C compared to group B. Specific suppression of OPN mRNA expression in kidneys of hyperoxaluric rats leads to a decrease in OPN production and simultaneously inhibits renal crystal deposition.

Antioxidant therapy prevents ethylene glycol-induced renal calcium oxalate crystal deposition in Wistar rats

Renal epithelial cell injury by reactive oxygen species is a prerequisite step in the pathogenesis of urolithiasis, and there is increasing evidence that reactive oxygen species is produced and oxidative stress (OS) is developed during idiopathic calcium oxalate nephrolithiasis. It appears that the administration of natural antioxidants has been used to protect against nephrolithiasis in human and experimental animals. Calcium oxalate urolithiasis was induced experimentally by administration of 0.75 % v/v ethylene glycol in drinking water of male Wistar rats weighing 150-200 g. Study was conducted in 4- and 8-week periods. In the 4-week period, Group 1 (control) was fed a standard commercial diet. Group 2 received the same diet with the addition of 0.75 % of ethylene glycol (EG). Group 3 received EG plus the diet, and water with additional antioxidant nutrients, and lemon juice as the dietary source of citrate (EG + AX). Group 4 was the same as Group 3, but with no EG in water. In the 8-week study protocol, Group 5 was fed the standard diet with EG in water for the first 28 days, followed with no EG. Group 6 (curative group) received the diet with EG for the first 28 days, followed by discontinuation of EG plus the addition of antioxidant nutrients. Group 7 was provided the diet with antioxidant nutrients for 8 weeks. Group 8 (preventive group) received the diet with antioxidant nutrients for 4 weeks, followed by antioxidant nutrients with EG for the next 4 weeks. Lime juice was given along the antioxidants. After treatment period, kidneys were removed and used for histopathological examination. In the 4-week study, the mean number of crystal deposits in Group 2 was significantly higher than that of animals in Group 3. After 8 weeks, animals given curative antioxidant supplementation within the second 4-week period developed fewer deposits in Group 6 as compared to Group 5 animals. In the other preventive AX loading Group 8, the number of crystal deposits was substantially less than that of either Group 2 or Group 5 animals (EG-treated rats). Results showed a beneficial effect on treating and superior renal protection for preventing stone deposition in the rat kidney. These results provide a scientific rationale for preventive and treatment roles of antioxidant nutrient complex in human kidney stone disease.

Animal Protein and the Risk of Kidney Stones: A Comparative Metabolic Study of Animal Protein Sources

We compared the effect of 3 animal protein sources on urinary stone risk. A total of 15 healthy subjects completed a 3-phase randomized, crossover metabolic study. During each 1-week phase subjects consumed a standard metabolic diet containing beef, chicken or fish. Serum chemistry and 24-hour urine samples collected at the end of each phase were compared using mixed model repeated measures analysis. Serum and urinary uric acid were increased for each phase. Beef was associated with lower serum uric acid than chicken or fish (6.5 vs 7.0 and 7.3 mg/dl, respectively, each p <0.05). Fish was associated with higher urinary uric acid than beef or chicken (741 vs 638 and 641 mg per day, p = 0.003 and 0.04, respectively). No significant difference among phases was noted in urinary pH, sulfate, calcium, citrate, oxalate or sodium. Mean saturation index for calcium oxalate was highest for beef (2.48), although the difference attained significance only compared to chicken (1.67, p = 0.02) but not to fish (1.79, p = 0.08). Consuming animal protein is associated with increased serum and urine uric acid in healthy individuals. The higher purine content of fish compared to beef or chicken is reflected in higher 24-hour urinary uric acid. However, as reflected in the saturation index, the stone forming propensity is marginally higher for beef compared to fish or chicken. Stone formers should be advised to limit the intake of all animal proteins, including fish.

Fibrinogen alpha chain precursor and apolipoprotein A-I in urine as biomarkers for noninvasive diagnosis of calcium oxalate nephrolithiasis: a proteomics study.

Calcium oxalate nephrolithiasis is the most common urological disease, but noninvasive and convenient methods of diagnosis are rarely available. Objective. The present study aimed to identify potential urine biomarkers for noninvasive diagnosis of CaOx nephrolithiasis. Methodology. Urine samples from 72 patients with CaOx nephrolithiasis and 30 healthy controls were collected and proteomics analysis was performed using matrix-assisted laser desorption/ionization-time of flight-mass spectrometer (MALDI-TOF-MS). Results. Thirteen proteins/peptides displayed statistically significant differences. The peptides of m/z 1207.23 and 2773.86 were selected by the genetic algorithm (GA) to build a possible diagnostic model. The area under the curve of m/z 1207.23 and 2773.86 was 0.936 and 0.987, respectively. The diagnostic model in distinguishing patients and healthy subjects showed 100% sensitivity and specificity. The peak at m/z 2773.86 was identified as fibrinogen alpha chain (FGA) with the sequence G.EGDFLAEGGGVR.G, and the peak at m/z 2773.86 was identified as apolipoprotein A-I (apoA-I) with the sequence L.PVLESFKVSFLSALEEYTKKLNTQ. Conclusion. The study results strongly suggested that urinary FGA and apoA-I are highly sensitive and specific biomarkers for noninvasive diagnosis of CaOx nephrolithiasis.

Ethylene glycol induced renal toxicity in female Wistar rats

The calcium oxalate nephrolithiasis due to ethylene glycol treatment has long been observed in male species but the prevalence of calcium oxalate nephrolithiasis in female species due to ethylene glycol has been still a subject of controversy. Ethylene glycol was administered in drinking water at three different doses (0.4%, 0.75% and 1.0%, v/v) for 28 days in female Wistar rats. Ethylene glycol treatment caused significant decrease in body weight and corresponding increase in relative organ weight with significant hypercalciuria, hyperoxaluria and proteinuria as well as increased retention of calcium, oxalate, phosphate and total protein in kidney in a dose — and time — dependent manner. However, calcium level was significantly reduced in the serum while an increase in total protein and phosphate level in serum was observed. Furthermore, there was a significant reduction in magnesium level in urine, serum and kidney due to ethylene glycol. The effects were also confirmed in histopathological studies.

Anti-urolithiatic effect of ethanolic extract of Pedalium murex linn. fruits on ethylene glycol-induced renal calculi.

To evaluate effect of ethanolic extract of Pedalium murex Linn. fruits on experimental model of calcium oxalate nephrolithiasis. Thirty-six male Wistar albino rats were randomly divided in 6 groups.Normal controls received distilled water for 28 days. Other five groups received ethylene glycol(1% v/v) in distilled water for 28 days. Pedalium murex ethanolic extract was given 200 mg/kg and 400 mg/kg orally in distilled water for 28 days in prophylactic groups (III and IV) and from 15th to 28th days in treatment groups (V and VI). The urea, creatinine, random blood sugar, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, bilirubin and calcium were measured on 28th day. 24 hr urinary oxalate and volume were measured on day 0 and 28. On day 28, kidneys were removed, weighed and subjected to histopathological examination. Calcium oxalate crystallization was evaluated by renal histopathology and in-vitro method of mineralization.All parameters were analyzed by Kruskal-Wallis or one-way ANOVA with post-hoc test. Pedalium murex showed significant improvement in renal function and kidney weight inprophylactic groups as compared to ethylene glycol controls. It did not show any effect on urinary oxalate, urine volume and any other serological parameters. Calcium oxalate crystallization was significantly reduced in all the Pedalium murex treated groups (P < .05). Calcium oxalate and phosphate mineralization were also inhibited by 33% and 57%. Ethanolic extract of Pedalium murex fruits possess significant activity for prevention of renal calculi.

Wu-Ling-San formula prophylaxis against recurrent calcium oxalate nephrolithiasis- a prospective randomized controlled trial

Wu-Ling-San (WLS) formula has been proved to prevent calcium oxalate nephrolithiasis both in vitro and in vivo. This is the first prospective, randomized and placebo-controlled clinical trial of WLS in calcium oxalate nephrolithiasis prevention. All patients who enrolled were asked to drink enough fluid to urinate at least 2 L daily during the study period. A 24-hour urine collection was performed to establish the baseline levels of multiple urinary parameters before taking the medicine. The patients were randomized and divided into two groups. The medication group took 2 gm WLS formula three times daily for 1 month. The control group took 2 gm placebo three times daily for 1 month. A 24-hour urine collection was performed to evaluate multiple urinary and serum parameters from all patients during the study period. A total of 39 patients were enrolled and 28 patients completed the study. Fourteen patients were allocated to WLS group and 14 patients to placebo group. After treatment, the mean urine output level increased to 2796.4 ± 525.7 ml/day (percentage of change, 13.9 %) in the WLS formula group. With placebo therapy, the mean decreased slightly to 2521.4 ± 762.7ml/day (percentage of change, -5.7 %). The percentage of change was significantly different between the two groups (independent t-test, P=0.02). No patient complained of side effects, such as fatigue, dizziness, musculoskeletal symptoms, or gastrointestinal disturbance. WLS formula is a promising adjunct to surgical and medical management of kidney stones. Active therapy with WLS formula has a positive effect on diuresis without leading to electrolyte imbalance.

Acute kidney injury in a patient with sarcoidosis: hypercalciuria and hypercalcemia leading to calcium phosphate deposition

Sarcoidosis can affect kidney function through many different mechanisms. We present a patient with sarcoidosis who developed acute kidney injury (AKI). He had a high 1,25-OH vitamin D level and hypercalciuria. As his renal function declined he developed hypercalcemia. A kidney biopsy showed acute tubular necrosis (ATN) with giant cell formation around calcium phosphate crystals. Calcium phosphate deposition is uncommon in sarcoid. We speculate that early interstitial calcium phosphate deposition may in time lead to the development of Randall's plaques and to the more typical calcium oxalate nephrolithiasis seen in sarcoidosis.

2086 OXALATE-INDUCED NADPH OXIDASE ACTIVATION IN RENAL EPITHELIAL CELLS: ROLE OF UPSTREAM MEDIATORS

You have accessJournal of UrologyStone Disease: Basic Research (I)1 Apr 20132086 OXALATE-INDUCED NADPH OXIDASE ACTIVATION IN RENAL EPITHELIAL CELLS: ROLE OF UPSTREAM MEDIATORS Vijayalakshmi Thamilselvan, Mani Menon, and Sivagnanam Thamilselvan Vijayalakshmi ThamilselvanVijayalakshmi Thamilselvan Detroit, MI More articles by this author , Mani MenonMani Menon Detroit, MI More articles by this author , and Sivagnanam ThamilselvanSivagnanam Thamilselvan Detroit, MI More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2013.02.2505AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Hyperoxaluria is one of the major risk factors for calcium oxalate nephrolithiasis and several in vitro and in vivo studies have shown that oxalate induced oxidative stress mediated cell injury contributes to renal stone formation. We have recently demonstrated that oxalate induces reactive oxygen species (ROS) via NADPH oxidase activation in renal epithelial cells. However, the upstream signaling mechanisms responsible for NADPH oxidase activation by oxalate are largely unknown. Therefore, we investigated the relevant upstream modulators of NADPH oxidase-dependent oxalate-induced cell injury in renal epithelial cells. METHODS Confluent monolayers of LLC-PK1 cells incubated with or without inhibitors of PKC (PKCα : inhibitor peptide, PKCδ: rottlerin), PI3-kinase (LY294002), tyrosine kinase (genistein), c-Src (PP2), p38 MAPK (SB203580), TGF-β1 (neutralizing antibody), NADPH oxidase (DPI), RNA synthesis (actinomycin-D), and protein synthesis (cycloheximide) for 30 min were then exposed to 0.5 to 1.0 mM oxalate for different time periods. The ROS production, cell injury (LDH released), and NADPH oxidase activity (lucigenin chemiluminescence) were determined. Phosphorylation of p40phox, activation of Rac1, and membrane translocation of p47phox were determined by Western analysis. RESULTS Exposure of LLC-PK1 cells to oxalate caused a dose and time dependent increase in ROS production, LDH release, and NADPH oxidase activity. Oxalate induced NADPH oxidase activity was associated with increased p40phox phosphorylation, p47phox membrane translocation, and Rac1 activation. Studies with actinomycin-D and cycloheximide indicated that NADPH oxidase activity was regulated at the transcriptional and translational levels. Inhibitors of PKC, PI3-kinase, tyrosine kinase, c-Src, and TGF-β1 significantly attenuated oxalate-induced NADPH oxidase activity, superoxide production, and LDH release. DPI pretreatment completely prevented oxalate-induced NADPH oxidase activity in LLC-PK1 cells, where as p38 MAPK inhibitor did not prevent oxalate-induced ROS, LDH release, and NADPH oxidase activity. CONCLUSIONS Our data demonstrates that oxalate-induced integrated signaling mechanisms leading to the activation of NADPH oxidase is responsible for sustained increase in ROS generation. The cellular injury as a result of increased ROS generation may lead to calcium oxalate kidney stone formation. Our study suggests that these signaling pathways may represent significant targets for therapeutic intervention in the treatment of recurrent kidney stones. © 2013 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 189Issue 4SApril 2013Page: e856 Advertisement Copyright & Permissions© 2013 by American Urological Association Education and Research, Inc.MetricsAuthor Information Vijayalakshmi Thamilselvan Detroit, MI More articles by this author Mani Menon Detroit, MI More articles by this author Sivagnanam Thamilselvan Detroit, MI More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Calcium oxalate nephrolithiasis and expression of matrix GLA protein in the kidneys

Polymorphism of the gene for matrix GLA protein (MGP), a calcification inhibitor, is associated with nephrolithiasis. However, experimental investigations of MGP role in stone pathogenesis are limited. We determined the effect of renal epithelial exposure to oxalate (Ox), calcium oxalate (CaOx) monohydrate (COM) or hydroxyapatite (HA) crystal on the expression of MGP. MDCK cells in culture were exposed to 0.3, 0.5 or 1 mM Ox and 33, 66 or 133-150 μg/cm(2) of COM/HA for 3-72 h. MGP expression and production were determined by Western blotting and densitometric analysis. Enzyme-linked immunosorbent assay was performed to determine MGP release into the medium. Hyperoxaluria was induced in male Sprague-Dawley rats by feeding hydroxyl-L-proline. Immunohistochemistry was performed to detect renal MGP expression. Exposure to Ox and crystals led to time- and concentration-dependent increase in expression of MGP in MDCK cells. Cellular response was quicker to crystal exposure than to the Ox, expression being significantly higher after 3-h exposure to COM or HA crystals and more than 6 h of exposure to Ox. MGP expression was increased in kidneys of hyperoxaluric rats particularly in renal peritubular vessels. We demonstrate increased expression of MGP in renal tubular epithelial cells exposed to Ox or CaOx crystals as well as the HA crystals. The most significant finding of this study is the increased staining seen in renal peritubular vessels of the hyperoxaluric rats, indicating involvement of renal endothelial cells in the synthesis of MGP.

Effect of astaxanthin on ethylene glycol induced nephrolithiasis

Nephrolithiasis is one of the most common and painful of urological disorders with a high prevalence rate. The role of calcium oxalate crystals, which are the predominant component of kidney stones in generating oxidative stress, have been clearly demonstrated in previous studies. Astaxanthin, found in marine organisms is a dietary xanthophyll carotenoid with enhanced antioxidative properties and pharmacological effects. In the present study, we have investigated the effect of this natural antioxidant, at a daily dose of 25mg/kg in experimental calcium oxalate nephrolithiasis in male Wistar rats. Liver function markers, hepatic antioxidants, albumin creatinine ratios, renal calcium content and changes in body and kidney weight have been studied to evaluate the effect of this carotenoid in vivo. The effect of citrate, a component of most pharmaceutical drugs for management of nephrolithiasis has also been evaluated for the purpose of comparison with astaxanthin treatment. Astaxanthin is seen to exert a protective effect on the liver and kidney tissues in ethylene glycol treated rats by improving the liver function, restoring the activity of the hepatic antioxidant enzymes, decreasing the albumin creatinine ratios and calcium levels and maintaining the organ to body weight ratio. Our results also indicate that astaxanthin administration is more beneficial than citrate treatment.