Nicorandil protects podocytes via modulation of antioxidative capacity in acute puromycin aminonucleoside-induced nephrosis in rats.

Nephrotic syndrome, characterized by proteinuria and hypoalbuminemia, results from the dysregulation of glomerular podocytes and is a significant cause of end-stage kidney disease. Patients with idiopathic nephrotic syndrome are generally treated with immunosuppressive agents; however, these agents produce various adverse effects. Previously, we reported the renoprotective effects of a stimulator of the mitochondrial ATP-dependent K+ channel (MitKATP), nicorandil, in a remnant kidney model. Nonetheless, the cellular targets of these effects remain unknown. Here, we examined the effect of nicorandil on puromycin aminonucleoside-induced nephrosis (PAN) rats, a well-established model of podocyte injury and human nephrotic syndrome. PAN was induced using a single intraperitoneal injection. Nicorandil was administered orally at 30 mg/kg/day. We found that proteinuria and hypoalbuminemia in PAN rats were significantly ameliorated following nicorandil treatment. Immunostaining and ultrastructural analysis under electron microscopy demonstrated that podocyte injury in PAN rats showed a significant partial attenuation following nicorandil treatment. Nicorandil ameliorated the increase in the oxidative stress markers nitrotyrosine and 8-hydroxy-2-deoxyguanosine in glomeruli. Conversely, nicorandil prevented the decrease in levels of the antioxidant enzyme manganese superoxide dismutase in PAN rats. We found that mitochondrial Ca2+ uniporter levels in glomeruli were higher in PAN rats than in control rats, and this increase was significantly attenuated by nicorandil. We conclude that stimulation of MitKATP by nicorandil reduces proteinuria by attenuating podocyte injury in PAN nephrosis, which restores mitochondrial antioxidative capacity, possibly through mitochondrial Ca2+ uniporter modulation. These data indicate that MitKATP may represent a novel target for podocyte injury and nephrotic syndrome.NEW & NOTEWORTHY Our findings suggest that the mitochondrial Ca2+ uniporter may be an upstream regulator of manganese superoxide dismutase and indicate a biochemical basis for the interaction between the ATP-sensitive K+ channel and Ca2+ signaling. We believe that our study makes a significant contribution to the literature because our results indicate that the ATP-sensitive K+ channel may be a potential therapeutic target for podocyte injury and nephrotic syndrome.

Xanthine Oxidoreductase Inhibitors Suppress the Onset of Exercise-Induced AKI in High HPRT Activity Urat1-Uox Double Knockout Mice.

BACKGROUND: Hereditary renal hypouricemia type 1 (RHUC1) is caused by URAT1/SLC22A12 dysfunction, resulting in urolithiasis and exercise-induced AKI (EIAKI). However, because there is no useful experimental RHUC1 animal model, the precise pathophysiologic mechanisms underlying EIAKI have yet to be elucidated. We established a high HPRT activity Urat1-Uox double knockout (DKO) mouse as a novel RHUC1 animal model for investigating the cause of EIAKI and the potential therapeutic effect of xanthine oxidoreductase inhibitors (XOIs). METHODS: The novel Urat1-Uox DKO mice were used in a forced swimming test as loading exercise to explore the onset mechanism of EIAKI and evaluate related purine metabolism and renal injury parameters. RESULTS: Urat1-Uox DKO mice had uricosuric effects and elevated levels of plasma creatinine and BUN as renal injury markers, and decreased creatinine clearance observed in a forced swimming test. In addition, Urat1-Uox DKO mice had increased NLRP3 inflammasome activity and downregulated levels of Na+-K+-ATPase protein in the kidney, as Western blot analysis showed. Finally, we demonstrated that topiroxostat and allopurinol, XOIs, improved renal injury and functional parameters of EIAKI. CONCLUSIONS: Urat1-Uox DKO mice are a useful experimental animal model for human RHUC1. The pathogenic mechanism of EIAKI was found to be due to increased levels of IL-1ß via NLRP3 inflammasome signaling and Na+-K+-ATPase dysfunction associated with excessive urinary urate excretion. In addition, XOIs appear to be a promising therapeutic agent for the treatment of EIAKI.

Mineralocorticoid Receptor Antagonists for Preventing Chronic Kidney Disease Progression: Current Evidence and Future Challenges.

Regulation and action of the mineralocorticoid receptor (MR) have been the focus of intensive research over the past 80 years. Genetic and physiological/biochemical analysis revealed how MR and the steroid hormone aldosterone integrate the responses of distinct tubular cells in the face of environmental perturbations and how their dysregulation compromises fluid homeostasis. In addition to these roles, the accumulation of data also provided unequivocal evidence that MR is involved in the pathophysiology of kidney diseases. Experimental studies delineated the diverse pathological consequences of MR overactivity and uncovered the multiple mechanisms that result in enhanced MR signaling. In parallel, clinical studies consistently demonstrated that MR blockade reduces albuminuria in patients with chronic kidney disease. Moreover, recent large-scale clinical studies using finerenone have provided evidence that the non-steroidal MR antagonist can retard the kidney disease progression in diabetic patients. In this article, we review experimental data demonstrating the critical importance of MR in mediating renal injury as well as clinical studies providing evidence on the renoprotective effects of MR blockade. We also discuss areas of future investigation, which include the benefit of non-steroidal MR antagonists in non-diabetic kidney disease patients, the identification of surrogate markers for MR signaling in the kidney, and the search for key downstream mediators whereby MR blockade confers renoprotection. Insights into these questions would help maximize the benefit of MR blockade in subjects with kidney diseases.

[A Case of Double Cancer of Squamous Cell Carcinoma of the Rectum and Adenocarcinoma of the Sigmoid Colon].

A 72-year-old male was transported to our hospital with complaints of heart palpitations and dyspnea since a month earlier and was immobile. Blood examination showed severe anemia, and colonoscopy revealed circumferential tumors in the rectum and the sigmoid colon. Histopathologic examination revealed the tumors as squamous cell carcinoma of the rectum and adenocarcinoma of the sigmoid colon. Therefore, they were diagnosed as double colorectal cancers. CT and MRI showed that rectal cancer invaded the seminal vesicles and the prostate; therefore, the patient underwent neoadjuvant chemoradiotherapy(oral capecitabine and concomitant radiation therapy: a total dose of 50.4 Gy/28 Fr)followed by total pelvic exenteration. Subsequent specimen pathology revealed a tumor regression grading of Grade 2 for the rectal and sigmoid colon cancers, and both were staged as ypT3N0M0, ypStage Ⅱa. Herein, we report a rare case of double cancer of adenocarcinoma of the sigmoid colon and squamous cell carcinoma of the rectum with a literature review.

Calcineurin dephosphorylates Kelch-like 3, reversing phosphorylation by angiotensin II and regulating renal electrolyte handling.

Calcineurin is a calcium/calmodulin-regulated phosphatase known for its role in activation of T cells following engagement of the T cell receptor. Calcineurin inhibitors (CNIs) are widely used as immunosuppressive agents; common adverse effects of CNIs are hypertension and hyperkalemia. While previous studies have implicated activation of the Na-Cl cotransporter (NCC) in the renal distal convoluted tubule (DCT) in this toxicity, the molecular mechanism of this effect is unknown. The renal effects of CNIs mimic the hypertension and hyperkalemia that result from germ-line mutations in with-no-lysine (WNK) kinases and the Kelch-like 3 (KLHL3)-CUL3 ubiquitin ligase complex. WNK4 is an activator of NCC and is degraded by binding to KLHL3 followed by WNK4's ubiquitylation and proteasomal degradation. This binding is prevented by phosphorylation of KLHL3 at serine 433 (KLHL3S433-P) via protein kinase C, resulting in increased WNK4 levels and increased NCC activity. Mechanisms mediating KLHL3S433-P dephosphorylation have heretofore been unknown. We now demonstrate that calcineurin expressed in DCT is a potent KLHL3S433-P phosphatase. In mammalian cells, the calcium ionophore ionomycin, a calcineurin activator, reduces KLHL3S433-P levels, and this effect is reversed by the calcineurin inhibitor tacrolimus and by siRNA-mediated knockdown of calcineurin. In vivo, tacrolimus increases levels of KLHL3S433-P, resulting in increased levels of WNK4, phosphorylated SPAK, and NCC. Moreover, tacrolimus attenuates KLHL3-mediated WNK4 ubiquitylation and degradation, while this effect is absent in KLHL3 with S433A substitution. Additionally, increased extracellular K+ induced calcineurin-dependent dephosphorylation of KLHL3S433-P These findings demonstrate that KLHL3S433-P is a calcineurin substrate and implicate increased KLHL3 phosphorylation in tacrolimus-induced pathologies.

A novel I551F variant of the Na+/HCO3- cotransporter NBCe1-A shows reduced cell surface expression, resulting in diminished transport activity.

Homozygous mutations in SLC4A4, which encodes the electrogenic Na+/[Formula: see text] cotransporter (NBCe1), cause proximal renal tubular acidosis associated with extrarenal symptoms. Although 17` mutated sites in SLC4A4 have thus far been identified among patients with proximal renal tubular acidosis, the physiological significance of other nonsynonymous single-nucleotide variants (SNVs) remains largely undetermined. Here, we investigated the functional properties of SNVs in NBCe1. From the National Center for Biotechnology Information dbSNP database, we identified 13 SNVs that have not previously been characterized in the highly conserved, transmembrane domains of NBCe1-A. Immunocytochemical analysis revealed that the I551F variant was present predominantly in the cytoplasm in human embryonic kidney (HEK)-293 cells, whereas all other SNVs did not show as dramatic a change in subcellular distribution. Western blot analysis in HEK-293 cells demonstrated that the I551F variant showed impaired glycosylation and a 69% reduction in cell surface levels. To determine the role of I551 in more detail, we examined the significance of various artificial mutants in both nonpolarized HEK-293 cells and polarized Madin-Darby canine kidney cells, which indicated that only I551F substitution resulted in cytoplasmic retention. Moreover, functional analysis using Xenopus oocytes demonstrated that the I551F variant had a significantly reduced activity corresponding to 39% of that of the wild-type, whereas any other SNVs and artificial I551 mutants did not show significant changes in activity. Finally, immunofluorescence experiments in HEK-293 cells indicated that the I551F variant retained wild-type NBCe1-A in the cytoplasm. These data demonstrate that the I551F variant of NBCe1-A shows impaired transport activity predominantly through cytoplasmic retention and suggest that the variant can have a dominant negative effect by forming complexes with wild-type NBCe1-A.NEW & NOTEWORTHY Electrogenic Na+/[Formula: see text] cotransporter 1-A (NBCe1-A) in the proximal tubule regulates the acid/base balance and fluid volume homeostasis. From the National Center for Biotechnology Information dbSNP database, we identified the I551F variant of NBCe1-A, which showed reduced glycosylation, cell surface expression, and transport activity. We also found that the I551F variant can exert a dominant negative effect on wild-type NBCe1-A, suggesting its physiological significance.

Upregulation of renal Na-K-2Cl cotransporter 2 in obese diabetes mellitus via a vasopressin receptor 2-dependent pathway.

Na-K-2Cl cotransporter 2 (NKCC2) in thick ascending limb (TAL) in the kidney plays a central role in tubuloglomerular feedback (TGF) system by sensing NaCl delivery to the distal tubules. Although accumulating data indicate that dysregulated TGF contributes to the progression of diabetic complications, the regulation of NKCC2 in diabetes mellitus (DM) remains unclear. We here show that NKCC2 is overactivated via a vasopressin receptor 2 (V2R)-dependent mechanism in db/db mice, a mouse model of obese DM. Compared with db/+ mice, we found that both aquaporin 2 and NKCC2 levels were significantly increased in the kidney in db/db mice. Immunohistochemical analysis of V2R and NKCC2 in the kidney demonstrated that V2R is present in the TAL, as well as in the collecting duct. Moreover, the administration of tolvaptan, a selective V2R antagonist, sharply decreased aquaporin 2 and NKCC2 in db/db mice, confirming the causal role of V2R signaling in NKCC2 induction in this model. Although tolvaptan reduced aquaporin 2 abundance also in db/+ mice, its effect on NKCC2 was modest compared with db/db mice. In total kidney lysates, uromodulin expression was not altered between db/+ and db/db mice, suggesting that V2R signaling alters NKCC2 without altering uromodulin levels. These data implicate the dysregulation of NKCC2 in the pathophysiology of type 2 DM, and underscore the complex nature of fluid volume disorders in diabetic kidney disease.

Phosphorylation by PKC and PKA regulate the kinase activity and downstream signaling of WNK4.

With-no-lysine kinase 4 (WNK4) regulates electrolyte homeostasis and blood pressure. WNK4 phosphorylates the kinases SPAK (Ste20-related proline alanine-rich kinase) and OSR1 (oxidative stress responsive kinase), which then phosphorylate and activate the renal Na-Cl cotransporter (NCC). WNK4 levels are regulated by binding to Kelch-like 3, targeting WNK4 for ubiquitylation and degradation. Phosphorylation of Kelch-like 3 by PKC or PKA downstream of AngII or vasopressin signaling, respectively, abrogates binding. We tested whether these pathways also affect WNK4 phosphorylation and activity. By tandem mass spectrometry and use of phosphosite-specific antibodies, we identified five WNK4 sites (S47, S64, S1169, S1180, S1196) that are phosphorylated downstream of AngII signaling in cultured cells and in vitro by PKC and PKA. Phosphorylation at S64 and S1196 promoted phosphorylation of the WNK4 kinase T-loop at S332, which is required for kinase activation, and increased phosphorylation of SPAK. Volume depletion induced phosphorylation of these sites in vivo, predominantly in the distal convoluted tubule. Thus, AngII, in addition to increasing WNK4 levels, also modulates WNK4 kinase activity via phosphorylation of sites outside the kinase domain.

Inhibition of Sodium Glucose Cotransporter 2 Attenuates the Dysregulation of Kelch-Like 3 and NaCl Cotransporter in Obese Diabetic Mice.

BACKGROUND: Mechanisms underlying the frequent association between salt-sensitive hypertension and type 2 diabetes remain obscure. We previously found that protein kinase C (PKC) activation phosphorylates Kelch-like 3 (KLHL3), an E3 ubiquitin ligase component, at serine 433. We investigated whether impaired KLHL3 activity results in increased renal salt reabsorption via NaCl cotransporter (NCC). METHODS: We used the db/db diabetes mouse model to explore KLHL3's role in renal salt handling in type 2 diabetes and evaluated mechanisms of KLHL3 dysregulation in cultured cells. RESULTS: We observed PKC activity in the db/db mouse kidney and phosphorylation of serine 433 in KLHL3 (KLHL3S433-P). This modification prevents binding of with-no-lysine (WNK) kinases; however, total KLHL3 levels were decreased, indicating severely impaired KLHL3 activity. This resulted in WNK accumulation, activating NCC in distal convoluted tubules. Ipragliflozin, a sodium glucose cotransporter 2 (SGLT2) inhibitor, lowered PKC activity in distal convoluted tubule cells and reduced KLHL3S433-P and NCC levels, whereas the thiazolidinedione pioglitazone did not, although the two agents similarly reduced in blood glucose levels. We found that, in human embryonic kidney cells expressing KLHL3 and distal convoluted tubule cells, cellular glucose accumulation increased KLHL3S433-P levels through PKC. Finally, the effect of PKC inhibition in the kidney of db/db mice confirmed PKC's causal role in KLHL3S433-P and NCC induction. CONCLUSIONS: Dysregulation of KLHL3 is involved in the pathophysiology of type 2 diabetes. These data offer a rationale for use of thiazide in individuals with diabetes and provide insights into the mechanism for cardiorenal protective effects of SGLT2 inhibitors.

Role of the Ubiquitin Proteasome System in the Regulation of Blood Pressure: A Review.

The kidney and the vasculature play crucial roles in regulating blood pressure. The ubiquitin proteasome system (UPS), a multienzyme process mediating covalent conjugation of the 76-amino acid polypeptide ubiquitin to a substrate protein followed by proteasomal degradation, is involved in multiple cellular processes by regulating protein turnover in various tissues. Increasing evidence demonstrates the roles of UPS in blood pressure regulation. In the kidney, filtered sodium is reabsorbed through diverse sodium transporters and channels along renal tubules, and studies conducted till date have provided insights into the complex molecular network through which ubiquitin ligases modulate sodium transport in different segments. Components of these pathways include ubiquitin ligase neuronal precursor cell-expressed developmentally downregulated 4-2, Cullin-3, and Kelch-like 3. Moreover, accumulating data indicate the roles of UPS in blood vessels, where it modulates nitric oxide bioavailability and vasoconstriction. Cullin-3 not only regulates renal salt reabsorption but also controls vascular tone using different adaptor proteins that target distinct substrates in vascular smooth muscle cells. In endothelial cells, UPS can also contribute to blood pressure regulation by modulating endothelial nitric oxide synthase. In this review, we summarize current knowledge regarding the role of UPS in blood pressure regulation, focusing on renal sodium reabsorption and vascular function.

PGI2 Analog Attenuates Salt-Induced Renal Injury through the Inhibition of Inflammation and Rac1-MR Activation.

Renal inflammation is known to be involved in salt-induced renal damage, leading to end-stage renal disease. This study aims to evaluate the role of inflammation in anti-inflammatory and renoprotective effects of beraprost sodium (BPS), a prostaglandin I2 (PGI2) analog, in Dahl salt-sensitive (DS) rats. Five-week-old male DS rats were fed a normal-salt diet (0.5% NaCl), a high-salt diet (8% NaCl), or a high-salt diet plus BPS treatment for 3 weeks. BPS treatment could inhibit marked proteinuria and renal injury in salt-loaded DS rats with elevated blood pressure, accompanied by renal inflammation suppression. Notably, high salt increased renal expression of active Rac1, followed by increased Sgk1 expressions, a downstream molecule of mineralocorticoid receptor (MR) signal, indicating salt-induced activation of Rac1-MR pathway. However, BPS administration inhibited salt-induced Rac1-MR activation as well as renal inflammation and damage, suggesting that Rac1-MR pathway is involved in anti-inflammatory and renoprotective effects of PGI2. Based upon Rac1 activated by inflammation, moreover, BPS inhibited salt-induced activation of Rac1-MR pathway by renal inflammation suppression, resulting in the attenuation of renal damage in salt-loaded DS rats. Thus, BPS is efficacious for the treatment of salt-induced renal injury.

Group I metabotropic glutamate receptor activation induces TRPC6-dependent calcium influx and RhoA activation in cultured human kidney podocytes.

Researches have shown that mice lacking the metabotropic glutamate receptor 1 (mGluR) showed albuminuria, remodeling of F-actin, with loss of stress fibers. Selective group I mGluRs agonist (S)-3,5-dihydroxyphenylglycine (DHPG) attenuated albuminuria in several rodent models of nephrotic syndrome. However, the molecular mechanism is obscure. Using a human podocyte cell line, we here investigated the molecular mechanisms of group I mGluRs-induced calcium influx and the formation of stress fibers. Our data showed that group I mGluRs activation by DHPG induced a significant calcium influx, and promoted cytoskeletal stress fiber formation and focal adhesions in podocytes. Pre-incubating podocytes with non-selective inhibitor of transient receptor potential channels (TRPC), or the knockdown of TRPC6 attenuated the calcium influx and the stress fiber formation induced by DHPG. Further, DHPG resulted in an increase of active RhoA expression. However, the knockdown of RhoA by siRNA abolished the DHPG-induced increase in stress fibers. Additionally, nonselective inhibitors of TRPC or TRPC6 knockdown clearly inhibited RhoA activation induced by DHPG, as assessed by Glutathione-S-transferase pull-down assay followed by Western blotting. Taken together, our findings suggest TRPC6 regulates actin stress fiber formation and focal adhesions via the RhoA pathway in response to group I mGluRs activation. Our data can potentially explain the mechanism of protective action of group I mGluRs in glomerular podocyte injury.

Electrolyte transport in the renal collecting duct and its regulation by the renin-angiotensin-aldosterone system.

Distal nephron of the kidney plays key roles in fluid volume and electrolyte homeostasis by tightly regulating reabsorption and excretion of Na+, K+, and Cl- Studies to date demonstrate the detailed electrolyte transport mechanisms in principal cells of the cortical collecting duct, and their regulation by renin-angiotensin-aldosterone system (RAAS). In recent years, however, accumulating data indicate that intercalated cells, another cell type that is present in the cortical collecting duct, also play active roles in the regulation of blood pressure. Notably, pendrin in ß-intercalated cells not only controls acid/base homeostasis, but is also one of the key components controlling salt and K+ transport in distal nephron. We have recently shown that pendrin is regulated by the co-ordinated action of angiotensin II (AngII) and aldosterone, and at the downstream of AngII, mammalian target of rapamycin (mTOR) signaling regulates pendrin through inhibiting the kinase unc51-like-kinase 1 and promoting dephosphorylation of mineralocorticoid receptor (MR). In this review, we summarize recent advances in the current knowledge on the salt transport mechanisms in the cortical collecting duct, and their regulation by the RAAS.

Aldosterone Is Essential for Angiotensin II-Induced Upregulation of Pendrin.

The renin-angiotensin-aldosterone system has an important role in the control of fluid homeostasis and BP during volume depletion. Dietary salt restriction elevates circulating angiotensin II (AngII) and aldosterone levels, increasing levels of the Cl-/HCO3- exchanger pendrin in ß-intercalated cells and the Na+-Cl- cotransporter (NCC) in distal convoluted tubules. However, the independent roles of AngII and aldosterone in regulating these levels remain unclear. In C57BL/6J mice receiving a low-salt diet or AngII infusion, we evaluated the membrane protein abundance of pendrin and NCC; assessed the phosphorylation of the mineralocorticoid receptor, which selectively inhibits aldosterone binding in intercalated cells; and measured BP by radiotelemetry in pendrin-knockout and wild-type mice. A low-salt diet or AngII infusion upregulated NCC and pendrin levels, decreased the phosphorylation of mineralocorticoid receptor in ß-intercalated cells, and increased plasma aldosterone levels. Notably, a low-salt diet did not alter BP in wild-type mice, but significantly decreased BP in pendrin-knockout mice. To dissect the roles of AngII and aldosterone, we performed adrenalectomies in mice to remove aldosterone from the circulation. In adrenalectomized mice, AngII infusion again upregulated NCC expression, but did not affect pendrin expression despite the decreased phosphorylation of mineralocorticoid receptor. By contrast, AngII and aldosterone coadministration markedly elevated pendrin levels in adrenalectomized mice. Our results indicate that aldosterone is necessary for AngII-induced pendrin upregulation, and suggest that pendrin contributes to the maintenance of normal BP in cooperation with NCC during activation of the renin-angiotensin-aldosterone system by dietary salt restriction.

Insulin stimulates uric acid reabsorption via regulating urate transporter 1 and ATP-binding cassette subfamily G member 2.

Accumulating data indicate that renal uric acid (UA) handling is altered in diabetes and by hypoglycemic agents. In addition, hyperinsulinemia is associated with hyperuricemia and hypouricosuria. However, the underlying mechanisms remain unclear. In this study, we aimed to investigate how diabetes and hypoglycemic agents alter the levels of renal urate transporters. In insulin-depleted diabetic rats with streptozotocin treatment, both UA excretion and fractional excretion of UA were increased, suggesting that tubular handling of UA is altered in this model. In the membrane fraction of the kidney, the expression of urate transporter 1 (URAT1) was significantly decreased, whereas that of ATP-binding cassette subfamily G member 2 (ABCG2) was increased, consistent with the increased renal UA clearance. Administration of insulin to the diabetic rats decreased UA excretion and alleviated UA transporter-level changes, while sodium glucose cotransporter 2 inhibitor (SGLT2i) ipragliflozin did not change renal UA handling in this model. To confirm the contribution of insulin in the regulation of urate transporters, normal rats received insulin and separately, ipragliflozin. Insulin significantly increased URAT1 and decreased ABCG2 levels, resulting in increased UA reabsorption. In contrast, the SGLT2i did not alter URAT1 or ABCG2 levels, although blood glucose levels were similarly reduced. Furthermore, we found that insulin significantly increased endogenous URAT1 levels in the membrane fraction of NRK-52E cells, the kidney epithelial cell line, demonstrating the direct effects of insulin on renal UA transport mechanisms. These results suggest a previously unrecognized mechanism for the anti-uricosuric effects of insulin and provide novel insights into the renal UA handling in the diabetic state.

Uric Acid in the Follow-Up Determines 30% Decline in Estimated GFR Over 2 Years: a Propensity Score Analysis.

BACKGROUND/AIMS: Higher level of serum uric acid (SUA) predicts early entry to dialysis in chronic kidney disease (CKD) patients. However, a short-term effect of SUA remains to be elucidated using a novel surrogate endpoint. METHODS: Japanese CKD stage 3 to 4 patients were retrospectively examined (n= 701). The follow-up level of SUA was estimated as time-averaged uric acid (TA-UA). A propensity score for 6.0, 6.5 or 7.0 mg/dL of TA-UA was respectively calculated using baseline 23 covariates. The time-to-event analysis was performed for 30% decline in estimated GFR over 2 years. RESULTS: Incidence rates over 2 years were 90 of 440 in men and 36 of 261 in women (p = 0.03). Despite the negative result of baseline SUA, stratified Cox regression on the quintiles of the estimated propensity score showed that higher TA-UA of the three thresholds were all significant (crude HR 2.10 to 2.44) even after adjusting for the confounders. Kaplan-Meier analysis after propensity score matching likewise showed worse survival in the patients with the higher TA-UA (HR 3.11 to 4.26). CONCLUSION: Higher SUA increases likelihood of reaching a surrogate endpoint over 2 years. Early intervention for SUA less than 6.0 mg/dL is recommended for slowing CKD progression.

Time to target uric acid to retard CKD progression.

Uric acid (UA) remains a possible risk factor of chronic kidney disease (CKD) but its potential role should be elucidated given a fact that multidisciplinary treatments assure a sole strategy to inhibit the progression to end-stage renal disease (ESRD). In clinical setting, most observational studies showed that elevation of serum uric acid (SUA) independently predicts the incidence and the development of CKD. The meta-analysis showed that SUA-lowering therapy with allopurinol may retard the progression of CKD but did not reach conclusive results due to small-sized studies. Larger scale, randomized placebo-controlled trials to assess SUA-lowering therapy are needed. Our recent analysis by propensity score methods has shown that the threshold of SUA should be less than 6.5 mg/dL to abrogate ESRD. In animal models an increase in SUA by the administration of oxonic acid, uricase inhibitor, or nephrectomy can induce glomerular hypertension, arteriolosclerosis including afferent arteriolopathy and tubulointerstitial fibrosis. The ever-growing discoveries of urate transporters prompt us to learn UA metabolism in the kidney and intestine. One example is that the intestinal ABCG2 may play a compensatory role at face of decreased renal clearance of UA in nephrectomized rats, the trigger of which is not a uremic toxin but SUA itself. This review will summarize the recent knowledge on the relationship between SUA and the kidney and try to draw a conclusion when and how to treat asymptomatic hyperuricemia accompanied by CKD. Finally we will address a future perspective on UA study including a Mendelian randomization approach.

Unique proximal tubular cell injury and the development of acute kidney injury in adult patients with minimal change nephrotic syndrome.

BACKGROUND: Adult patients with minimal change nephrotic syndrome (MCNS) are often associated with acute kidney injury (AKI). To assess the mechanisms of AKI, we examined whether tubular cell injuries unique to MCNS patients exist. METHODS: We performed a retrospective analysis of clinical data and tubular cell changes using the immunohistochemical expression of vimentin as a marker of tubular injury and dedifferentiation at kidney biopsy in 37 adult MCNS patients. AKI was defined by the criteria of the Kidney Disease: Improving Global Outcomes (KDIGO) Clinical Practice Guidelines for AKI. RESULTS: Thirteen patients (35.1%) were designated with AKI at kidney biopsy. No significant differences in age, history of hypertension, chronic kidney disease, diuretics use, proteinuria, and serum albumin were noted between the AKI and non-AKI groups. Urinary N-acetyl-ß-D-glucosaminidase (uNAG) and urinary alpha1-microglobulin (uA1MG) as markers of tubular injury were increased in both groups, but the levels were significantly increased in the AKI group compared with the non-AKI group. The incidence of vimentin-positive tubules was comparable between AKI (84.6%) and non-AKI (58.3%) groups, but vimentin-positive tubular area per interstitial area was significantly increased in the AKI group (19.8%) compared with the non-AKI group (6.8%) (p = 0.011). Vimentin-positive injured tubules with tubular simplification (loss of brush-border of the proximal tubule/dilated tubule with flattening of tubular epithelium) were observed in the vicinity of glomeruli in both groups, suggesting that the proximal convoluted tubules were specifically injured. Two patients exhibited relatively severe tubular injuries with vimentin positivity and required dialysis within 2 weeks after kidney biopsy. The percentage of the vimentin-positive tubular area was positively correlated with uNAG but not with uA1MG in the non-AKI group. CONCLUSIONS: Proximal tubular injuries with increased uNAG exist in MCNS patients without renal dysfunction and were more severe in the AKI group than they were in the non-AKI group. The unique tubular injuries probably due to massive proteinuria might be a predisposing factor for the development of severe AKI in adult MCNS patients.

Angiotensin II signaling via protein kinase C phosphorylates Kelch-like 3, preventing WNK4 degradation.

Hypertension contributes to the global burden of cardiovascular disease. Increased dietary K(+) reduces blood pressure; however, the mechanism has been obscure. Human genetic studies have suggested that the mechanism is an obligatory inverse relationship between renal salt reabsorption and K(+) secretion. Mutations in the kinases with-no-lysine 4 (WNK4) or WNK1, or in either Cullin 3 (CUL3) or Kelch-like 3 (KLHL3)--components of an E3 ubiquitin ligase complex that targets WNKs for degradation--cause constitutively increased renal salt reabsorption and impaired K(+) secretion, resulting in hypertension and hyperkalemia. The normal mechanisms that regulate the activity of this ubiquitin ligase and levels of WNKs have been unknown. We posited that missense mutations in KLHL3 that impair binding of WNK4 might represent a phenocopy of the normal physiologic response to volume depletion in which salt reabsorption is maximized. We show that KLHL3 is phosphorylated at serine 433 in the Kelch domain (a site frequently mutated in hypertension with hyperkalemia) by protein kinase C in cultured cells and that this phosphorylation prevents WNK4 binding and degradation. This phosphorylation can be induced by angiotensin II (AII) signaling. Consistent with these in vitro observations, AII administration to mice, even in the absence of volume depletion, induces renal KLHL3(S433) phosphorylation and increased levels of both WNK4 and the NaCl cotransporter. Thus, AII, which is selectively induced in volume depletion, provides the signal that prevents CUL3/KLHL3-mediated degradation of WNK4, directing the kidney to maximize renal salt reabsorption while inhibiting K(+) secretion in the setting of volume depletion.

Potassium depletion stimulates Na-Cl cotransporter via phosphorylation and inactivation of the ubiquitin ligase Kelch-like 3.

Kelch-like 3 (KLHL3) is a component of an E3 ubiquitin ligase complex that regulates blood pressure by targeting With-No-Lysine (WNK) kinases for degradation. Mutations in KLHL3 cause constitutively increased renal salt reabsorption and impaired K+ secretion, resulting in hypertension and hyperkalemia. Although clinical studies have shown that dietary K+ intake affects blood pressure, the mechanisms have been obscure. In this study, we demonstrate that the KLHL3 ubiquitin ligase complex is involved in the low-K+-mediated activation of Na-Cl cotransporter (NCC) in the kidney. In the distal convoluted tubules of mice eating a low-K+ diet, we found increased KLHL3 phosphorylation at S433 (KLHL3S433-P), a modification that impairs WNK binding, and also reduced total KLHL3 levels. These changes are accompanied by the accumulation of the target substrate WNK4, and activation of the downstream kinases SPAK (STE20/SPS1-related proline-alanine-rich protein kinase) and OSR1 (oxidative stress-responsive 1), resulting in NCC phosphorylation and its accumulation at the plasma membrane. Increased phosphorylation of S433 was explained by increased levels of active, phosphorylated protein kinase C (but not protein kinase A), which directly phosphorylates S433. Moreover, in HEK cells expressing KLHL3 and WNK4, we showed that the activation of protein kinase C by phorbol 12-myristate 13-acetate induces KLHL3S433-P and increases WNK4 levels by abrogating its ubiquitination. These data demonstrate the role of KLHL3 in low-K+-mediated induction of NCC; this physiologic adaptation reduces distal electrogenic Na+ reabsorption, preventing further renal K+ loss but promoting increased blood pressure.