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Nephrotoxicity, including electrolytic disorders and acute kidney injury (AKI), limits the clinical dosage and utility of platinated antineoplastics such as cisplatin. Cisplatin nephrotoxicity embodies a tubulopathy involving the medullary S2 and S3 segments of the proximal and the distal tubules. Higher dosage extends damage over the cortical S1 segment and intensifies overall injury. However, the standard diagnosis based on plasma creatinine as well as novel injury biomarkers lacks enough pathophysiological specificity. Further granularity in the detection of renal injury would help understand the implications of individual damage patterns needed for personalized patient handling. In this article, we studied the association of urinary ganglioside GM2 activator protein (GM2AP) with the patterns of tubular damage produced by 5 and 10 mg/kg cisplatin in rats. Our results show that GM2AP appears in the urine only following damage to the cortical segment of the proximal tubule. The information provided by GM2AP is not redundant with but distinct and complementary to that provided by urinary neutrophil gelatinase-associated lipocalin (NGAL). Similarly, treatment with 150 mg/kg/day gentamicin damages the renal cortex and increases GM2AP urinary excretion; whereas renal ischemia, which does not affect the cortex, has no effect on GM2AP. Because of the key role of the cortical proximal tubule in renal function, we contend GM2AP as a potential diagnostic biomarker to stratify AKI patients according to the underlying damage and follow their evolution and prognosis. Prospectively, urinary GM2AP may help grade the severity of platinated antineoplastic nephrotoxicity by forming part of a non-invasive liquid biopsy., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Competing interests The authors declare no competing interests., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The chemotherapeutic agent cisplatin accumulates in the kidneys, leading to acute kidney injury (AKI). Preclinical and clinical studies have demonstrated sex-dependent outcomes of cisplatin-AKI. Deranged histone deacetylase (HDAC) activity is hypothesized to promote the pathogenesis of male murine cisplatin-AKI; however, it is unknown whether there are sex differences in the kidney HDACs. We hypothesized that there would be sex-specific Hdac expression, localization, or enzymatic activity, which may explain sexual dimorphic responses to cisplatin-AKI. In normal human kidney RNA samples, HDAC10 was significantly greater in the kidneys of women compared with men, whereas HDAC1 , HDAC6 , HDAC10 , and HDAC11 were differentially expressed between the kidney cortex and medulla, regardless of sex. In a murine model of cisplatin-AKI (3 days after a 15 mg/kg injection), we found few sex- or cisplatin-related differences in Hdac kidney transcripts among the mice. Although Hdac9 was significantly greater in female mice compared with male mice, HDAC9 protein localization did not differ. Hdac7 transcripts were greater in the inner medulla of cisplatin-AKI mice, regardless of sex, and this agreed with a greater HDAC7 abundance. HDAC activity within the cortex, outer medulla, and inner medulla was significantly lower in cisplatin-AKI mice but did not differ between the sexes. In agreement with these findings, a class I HDAC inhibitor did not improve kidney injury or function. In conclusion, even though cisplatin-AKI was evident and there were transcript level differences among the different kidney regions in this model, there were few sex- or cisplatin-dependent effects on kidney HDAC localization or activity. NEW & NOTEWORTHY Kidney histone deacetylases (HDACs) are abundant in male and female mice, and the inner medulla has the greatest HDAC activity. A low dose of cisplatin caused acute kidney injury (AKI) in these mice, but there were few changes in kidney HDACs at the RNA/protein/activity level. A class I HDAC inhibitor failed to improve AKI outcomes. Defining the HDAC isoform, cellular source, and interventional timing is necessary to determine whether HDAC inhibition is a therapeutic strategy to prevent cisplatin-AKI in both sexes.

Renal iron overload is a common complication of diabetes that leads to oxidative stress and mitochondrial dysfunction in the kidneys. This study investigated the effects of iron chelation using deferiprone on mitochondrial dysfunction and oxidative stress in the renal cortex of a murine model of type 2 diabetes. Diabetic rats were treated with deferiprone (50 mg/kg BW) for 16 weeks. Our results show that iron chelation with deferiprone significantly increased the nuclear accumulation of Nrf2, a transcription factor that regulates the expression of antioxidant enzymes. This led to enhanced antioxidant capacity, reduced production of reactive oxygen species, and improved mitochondrial bioenergetic function in diabetic rats. However, chronic iron chelation led to altered mitochondrial respiration and increased oxidative stress in non-diabetic rats. In conclusion, our findings suggest that iron chelation with deferiprone protects mitochondrial bioenergetics and mitigates oxidative stress in the renal cortex, involving the NRF2 pathway in type 2 diabetes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

A high-salt (HS) diet leads to metabolic disorders in Dahl salt-sensitive (SS) rats, and promotes the development of hypertension. According to the changes in the metabolites of SS rats, a set of combined dietary supplements containing amino acids and organic acids (AO) were designed. The purpose of the present study was to evaluate the effect of AO supplementation on the blood pressure of SS rats after the HS diet and clarify the mechanism of AO by metabolomics and biochemical analyses. The results showed that AO supplementation avoided the elevation of blood pressure induced by the HS diet in SS rats, increased the renal antioxidant enzyme activities (catalase, superoxide dismutase, glutathione reductase, and glutathione S-transferase), reduced the H 2 O 2 and MDA levels, and restored the normal antioxidant status of the serum and kidneys. AO also reversed the decrease in the nitric oxide (NO) levels and NO synthase activity induced by the HS feed, which involved the L-arginine/NO pathway. Metabolomics analysis showed that AO administration increased the levels of amino acids such as cysteine, glycine, hypotaurine, and lysine in the renal medulla and the levels of leucine, isoleucine, and serine in the renal cortex. Of note, lysine, hypotaurine and glycine had higher metabolic centrality in the metabolic correlation network of the renal medulla after AO administration. In conclusion, AO intervention could prevent HS diet-induced hypertension in SS rats by restoring the metabolic homeostasis of the kidneys. Hence, AO has the potential to become a functional food additive to improve salt-sensitive hypertension.

Diabetic nephropathy (DN), the leading cause of end-stage renal disease, has become a massive global health burden. Despite considerable efforts, the underlying mechanisms have not yet been comprehensively understood. In this study, a systematic approach was utilized to identify the microRNA signature in DN and to introduce novel drug targets (DTs) in DN. Using microarray profiling followed by qPCR confirmation, 13 and 6 differentially expressed (DE) microRNAs were identified in the kidney cortex and medulla, respectively. The microRNA-target interaction networks for each anatomical compartment were constructed and central nodes were identified. Moreover, enrichment analysis was performed to identify key signaling pathways. To develop a strategy for DT prediction, the human proteome was annotated with 65 biochemical characteristics and 23 network topology parameters. Furthermore, all proteins targeted by at least one FDA-approved drug were identified. Next, mGMDH-AFS, a high-performance machine learning algorithm capable of tolerating massive imbalanced size of the classes, was developed to classify DT and non-DT proteins. The sensitivity, specificity, accuracy, and precision of the proposed method were 90%, 86%, 88%, and 89%, respectively. Moreover, it significantly outperformed the state-of-the-art (P-value ≤ 0.05) and showed very good diagnostic accuracy and high agreement between predicted and observed class labels. The cortex and medulla networks were then analyzed with this validated machine to identify potential DTs. Among the high-rank DT candidates are Egfr, Prkce, clic5, Kit, and Agtr1a which is a current well-known target in DN. In conclusion, a combination of experimental and computational approaches was exploited to provide a holistic insight into the disorder for introducing novel therapeutic targets., (© 2021. The Author(s).)

Diabetic nephropathy (DN) is a primary cause of end‑stage renal disease. Despite the beneficial effects of astragaloside IV (AS)‑IV on renal disease, the underlying mechanism of its protective effects against DN has not been fully determined. The aims of the present study were to assess the effects of AS‑IV against DN in db/db mice and to explore the mechanism of AS‑IV involving the NLR family pyrin domain containing 3 (NLRP3), caspase‑1 and interleukin (IL)‑1β pathways. The 8‑week‑old db/db mice received 40 mg/kg AS‑IV once a day for 12 weeks via intragastric administration. Cultured mouse podocytes were used to further confirm the underlying mechanism in vitro . AS‑IV effectively reduced weight gain, hyperglycemia and the serum triacylglycerol concentration in db/db mice. AS‑IV also reduced urinary albumin excretion, urinary albumin‑to‑creatinine ratio and creatinine clearance rate, as well as improved renal structural changes, accompanied by the upregulation of the podocyte markers podocin and synaptopodin. AS‑IV significantly inhibited the expression levels of NLRP3, caspase‑1 and IL‑1β in the renal cortex, and reduced the serum levels of tumor necrosis factor (TNF)‑α and monocyte chemoattractant protein‑1. In high glucose‑induced podocytes, AS‑IV significantly improved the expression levels of NLRP3, pro‑caspase‑1 and caspase‑1, and inhibited the cell viability decrease in a dose‑dependent manner, while NLRP3 overexpression eliminated the effect of AS‑IV on podocyte injury and the inhibition of the NLRP3 and caspase‑1 pathways. The data obtained from in vivo and in vitro experiments demonstrated that AS‑IV ameliorated renal functions and podocyte injury and delayed the development of DN in db/db mice via anti‑NLRP3 inflammasome‑mediated inflammation.

High saturated fat diets have been associated with the development of obesity and hypertension, along with other pathologies related to the metabolic syndrome. In contrast, the Mediterranean diet, characterized by its high content of monounsaturated fatty acids, has been proposed as a dietary factor capable of positively regulating cardiovascular function. These effects have been linked to changes in the local renal renin angiotensin system (RAS) and the activity of the sympathetic nervous system. The main goal of this study was to analyze the role of two dietary fat sources on aminopeptidases activities involved in local kidney RAS. Male Wistar rats (six months old) were fed during 24 weeks with three different diets: the standard diet (S), the standard diet supplemented with virgin olive oil (20%) (VOO), or the standard diet enriched with butter (20%) plus cholesterol (0.1%) (Bch). Kidney samples were separated in medulla and cortex for aminopeptidase activities (AP) assay. Urine samples were collected for routine analysis by chemical tests. Aminopeptidase activities were determined by fluorometric methods in soluble (sol) and membrane-bound (mb) fractions of renal tissue, using arylamide derivatives as substrates. After the experimental period, the systolic blood pressure (SBP) values were similar in standard and VOO animals, and significantly lower than in the Bch group. At the same time, a significant increase in GluAP and IRAP activities were found in renal medulla of Bch animals. However, in VOO group the increase of GluAP activity in renal medulla was lower, while AspAP activity decreased in the renal cortex. Furthermore, the VOO diet also affected other aminopeptidase activities, such as TyrAP and pGluAP, related to the regulation of the sympathetic nervous system and the metabolic rate. These results support the beneficial effect of VOO in the regulation of SBP through changes in local AP activities of the kidney.

Diabetic kidney disease (DKD) is a prevalent and progressive comorbidity of diabetes mellitus that increases one's risk of developing renal failure. Progress toward development of better DKD therapeutics is limited by an incomplete understanding of forces driving and connecting the various features of DKD, which include renal steatosis, fibrosis, and microvascular dysfunction. Herein we review the literature supporting roles for bioactive ceramides as inducers of local and systemic DKD pathology. In rodent models of DKD, renal ceramides are elevated, and genetic and pharmacological ceramide-lowering interventions improve kidney function and ameliorate DKD histopathology. In humans, circulating sphingolipid profiles distinguish human DKD patients from diabetic controls. These studies highlight the potential for ceramide to serve as a central and therapeutically tractable lipid mediator of DKD., Competing Interests: SS is a consultant and shareholder with Centaurus Therapeutics. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Nicholson, Pezzolesi and Summers.)

Oxidative stress (OS)-induced glutathione (GSH) depletion plays an essential role in several kidney diseases such as chronic kidney disease and nephrotoxicity. The OS-dependent activation of TRPM2 cation channel in several neurons and cells were modulated by the concentration of intracellular GSH. However, the effects of GSH alteration on TRPM2 activation, OS, and apoptosis in the cortical collecting duct (mpkCCD c14 ) cells still remain elusive. We investigated the effects of GSH supplementation on OS-induced TRPM2 activation, mitochondrial oxidative stress, and apoptosis in the human embryonic kidney 293 (HEK293) and mpkCCD c14  cells treated with buthionine-sulfoximine (BSO), a GSH synthase inhibitor. The HEK293 and mpkCCD c14  cells were divided into five groups as control, GSH (10 mM for 2 h), BSO (0.5 mM for 6 h), BSO + GSH, and BSO + TRPM2 channel blockers. Apoptosis, cell death, mitochondrial OS, caspase -3, caspase -9, cytosolic free Zn 2+ , and Ca 2+ concentrations were increased in the BSO group of the TRPM2 expressing mpkCCD c14  cells, although they were diminished by the treatments of GSH, PARP-1 inhibitors (PJ34 and DPQ), and TRPM2 blockers (ACA and 2-APB). The BSO-induced decreases in the levels of cell viability and cytosolic GSH were increased by the treatments of GSH, ACA, and 2-APB. However, the effects of BSO and GSH were not observed in the non-TRPM2 expressing HEK293 cells. Current results show that maintaining GSH homeostasis is not only important for quenching OS in the cortical collecting duct cells but equally critical to modulate TRPM2 activation. Thus, suppressing apoptosis and mitochondrial OS responses elicited by oxidant action of GSH depletion., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Hypertension associated with hyperhomocysteinemia (HHcy) is associated with a high risk of vascular diseases. However, the mechanisms of HHcy-associated hypertensive renal damage and the efficacy of folic acid (FA) as a treatment have not been fully elucidated. The aim of the present study was to evaluate whether lowering the plasma homocysteine (Hcy) level using different doses of FA can reduce HHcy-associated glomerular injury in spontaneously hypertensive rats (SHRs) and to clarify the potential mechanisms of such effects. SHRs were randomized into a control group, HHcy group, HHcy + low-dose FA (LFA) group, and HHcy + high-dose FA (HFA) group. Compared with the control group, the HHcy group had reduced serum superoxide dismutase and GFR levels and elevated serum malondialdehyde and urinary albumin creatinine ratio levels. Increased extracellular matrix of the glomerulus and an increased glomerular sclerosis index, podocyte foot process effacement and fusion, as well as increased podocyte apoptosis, were observed in the HHcy group compared with the control group; these effects were associated with increased expression of NOX2 and NOX4 and decreased nephrin expression in renal tissue from SHRs with HHcy. HHcy-induced changes were counteracted by LFA and HFA treatment. Apart from lower levels of NOX2 in the HHcy + HFA group, there were no significant differences in other indicators between the HHcy + LFA and HHcy + HFA groups. These results suggest that even at a low dose, FA can reduce plasma Hcy and attenuate HHcy-induced glomerular injury by inhibiting oxidative stress and apoptosis.

A number of metabolic disorders, including hyperlipidemia, potentially cause chronic kidney disease (CKD), one of their major chronic complications and comorbidities. Rosuvastatin is one of the widely used antiatherogenic drugs among hyperlipidemic patients. Meanwhile, sarpogrelate is not only a 5-hydroxytryptamine receptor antagonist but also an antiplatelet agent, inhibiting platelet-stimulated blood coagulation and improving peripheral circulation. In this study, a combination of sarpogrelate and/or rosuvastatin was used on CKD mice induced by a high-fat diet for 8 weeks. The mice were tested for pathological changes using histological evaluation. Tremendous alterations were found, including a remarked increase in total cholesterol and low-density lipoprotein cholesterol levels, glomerular endothelial proliferation, and mesangial expansion. Also, tubular damage and extracellular matrix accumulation occurred, namely, a marked increase in the macula densa, scattered and apoptotic loss of the apical brush border with vacuolated basophilic cytoplasm and heavily stained nuclei, and expanded Bowman's space, which were at least partially ameliorated by sarpogrelate and/or rosuvastatin treatment. The analysis of expression profiles at both the RNA and protein levels, using real-time quantitative polymerase chain reaction and Western blot analysis, indicated that LDL-R/CD68/LOX-1-positive monocyte/macrophage-mediated enhanced proinflammatory activation, including the significant upregulation of tumor necrosis factor- α and interleukin-6, was actually attenuated by sarpogrelate and/or rosuvastatin treatment. The findings indicated that sarpogrelate and/or rosuvastatin treatment potentially ameliorates CKD progression in patients with the aforementioned comorbid metabolic disorders., Competing Interests: All authors declare that they have no conflict of interests., (Copyright © 2020 Jingyi Xu et al.)

Bumetanide, a sulfamyl loop diuretic, is used for the treatment of edema in association with congestive heart failure. Being a polar, anionic compound at physiologic pH, bumetanide uptake and efflux into different tissues is largely transporter-mediated. Of note, organic anion transporters (SLC22A) have been extensively studied in terms of their importance in transporting bumetanide to its primary site of action in the kidney. The contribution of one of the less-studied bumetanide transporters, monocarboxylate transporter 6 (MCT6; SLC16A5), to bumetanide pharmacokinetics (PK) and pharmacodynamics (PD) has yet to be characterized. The affinity of bumetanide for murine Mct6 was evaluated using Mct6-transfected Xenopus laevis oocytes. Furthermore, bumetanide was intravenously and orally administered to wild-type mice (Mct6 +/+ ) and homozygous Mct6 knockout mice (Mct6 -/- ) to elucidate the contribution of Mct6 to bumetanide PK/PD in vivo. We demonstrated that murine Mct6 transports bumetanide at a similar affinity compared with human MCT6 (78 and 84 μM, respectively, at pH 7.4). After bumetanide administration, there were no significant differences in plasma PK. Additionally, diuresis was significantly decreased by ∼55% after intravenous bumetanide administration in Mct6 -/- mice. Kidney cortex concentrations of bumetanide were decreased, suggesting decreased Mct6-mediated bumetanide transport to its site of action in the kidney. Overall, these results suggest that Mct6 does not play a major role in the plasma PK of bumetanide in mice; however, it significantly contributes to bumetanide's pharmacodynamics due to changes in kidney concentrations. SIGNIFICANCE STATEMENT: Previous evidence suggested that MCT6 transports bumetanide in vitro; however, no studies to date have evaluated the in vivo contribution of this transporter. In vitro studies indicated that mouse and human MCT6 transport bumetanide with similar affinities. Using Mct6 knockout mice, we demonstrated that murine Mct6 does not play a major role in the plasma pharmacokinetics of bumetanide; however, the pharmacodynamic effect of diuresis was attenuated in the knockout mice, likely because of the decreased bumetanide concentrations in the kidney., (Copyright © 2020 by The American Society for Pharmacology and Experimental Therapeutics.)

Cell cultures are often used to study physiological processes in health and disease. It is well-known that cells change their gene expression in vitro compared to in vivo, but it is rarely experimentally addressed. High glucose is a known trigger of apoptosis in proximal tubular cells (PTC). Here we used RNA-seq to detect differentially expressed genes in cultures of primary rat PTC, 3 days old, compared to cells retrieved directly from rat outer renal cortex and between PTC exposed to 15 mM glucose and control for 8 h. The expression of 6,174 genes was significantly up- or downregulated in the cultures of PTC compared to the cells in the outer renal cortex. Most altered were mitochondrial and metabolism related genes. Gene expression of proapoptotic proteins were upregulated and gene expression of antiapoptotic proteins were downregulated in PTC. Expression of transporter related genes were generally downregulated. After 8 h, high glucose had not altered the gene expression in PTC. The current study provides evidence that cells alter their gene expression in vitro compared to in vivo and suggests that short-term high glucose exposure can trigger apoptosis in PTC without changing the gene expression levels of apoptotic proteins.

Background: Few studies examined the effect of long-acting nitrates on renal function in chronic heart failure (CHF). Thus, we aimed to investigate the effect of long-acting nitrate on the expression of adrenoceptors (AR) and angiotensin II receptor (ATR) subtypes of the renal cortex, in rats with myocardial infarction-induced CHF., Methods: Rats were randomly divided into the following groups: control, sham-operated, CHF, low- and high-dose nitrate, positive drug control (olmesartan), and high-dose of long-acting nitrate + olmesartan. Ultrasound echocardiography markers were compared, and the levels of AR subtypes, AT 1 R, and AT 2 R were measured using reverse transcription-polymerase chain reaction and western blot analysis. Histopathology of the kidney was determined on hematoxylin and eosin-stained sections., Results: CHF significantly increased plasma renin activity (PRA) and angiotensin II levels, upregulated AT 1 R expression and downregulated α 1A -, β 1 -, β 2 -AR, and AT 2 R expression compared to the sham control. High-dose nitrate or olmesartan alone, and especially in combination, decreased the levels of PRA and angiotensin II and downregulated the CHF-induced expression of AT 1 R, α 1A -, β 1 -, and β 2 -AR, and AT 2 R. CHF resulted in significant impairment of the renal tissue, including inflammatory cells infiltration to the tubular interstitium and surrounding the renal glomerulus, and tubular necrosis, which was alleviated in all treatment groups to different degrees., Conclusions: Long-acting nitrates could reverse CHF-induced changes in AR and ATR subtypes in the kidney, and improve cardiac function to protect renal function. Compared with monotherapy, the combination of nitrates and olmesartan shows more significant benefits in regulating AR and ATR subtypes.

Objective To explore the expression of renal cortex bone morphogenetic protein and activating protein membrane binding inhibitor (BAMBI), NLR family pyrin domain containing 3 (NLRP3), apoptosis-associated speck like protein containing a CARD-2 (ASC-2), caspase-1, inflammatory mediators and related fibrosis markers in renal cortex of diabetic mellitus (DM) rats treated with insulin. Methods SD rats were randomly divided into 3 groups: a normal control group, a DM group and an insulin treatment group. Each group included 8 rats. The DM rat model was established by streptozotocin (STZ). Four weeks after STZ injection, the rats in the insulin treatment group was subcutaneously injected with insulin at the dose of 8 U/kg once, three times each day for successive 6 weeks, and then sacrificed finally. The expression of BAMBI, NLRP3 and epithelial cadherin (E-cadherin) in the renal cortex were detected by immunohistochemistry. The expression of BAMBI, NLRP3, ASC-2, caspase-1, α-smooth muscle actin (α-SMA), fibronectin (FN) and type IV collagen (Col4) in the renal cortex were detected by Western blot analysis. The mRNA levels of BAMBI and NLRP3 in the renal cortex were measured by real-time quantitative PCR, and the levels of interleukin-1β (IL-1β) and IL-18 in the renal cortex were tested by ELISA. The correlation between BAMBI and NLRP3 was analyzed by Spearman method. Results Compared with the normal control group, the expression of NLRP3, ASC-2, caspase-1, α-SMA, FN and Col4 significantly increased, but BAMBI and E-cadherin significantly decreased in the renal cortex of DM group. However, the expression of NLRP3, ASC-2, caspase-1, α-SMA, FN, Col4, IL-1β and IL-18 in the insulin-treated group were lower than those in the DM group, but the expression of BAMBI and E-cadherin were higher. According to the correlation analysis, there was a negative correlation between the expression of BAMBI and NLRP3 in the DM group and insulin treatment group. Conclusion Insulin can restore the expression of BAMBI in the renal cortex of DM rats, and then inhibit the activation of NLRP3, thus reducing the renal inflammatory response in the kidney of DM rats and preventing or delaying the progression of diabetic nephropathy.

The rodent collecting duct (CD) expresses a 24p3/NGAL/lipocalin-2 (LCN2) receptor (SLC22A17) apically, possibly to mediate high-affinity reabsorption of filtered proteins by endocytosis, although its functions remain uncertain. Recently, we showed that hyperosmolarity/-tonicity upregulates SLC22A17 in cultured mouse inner-medullary CD cells, whereas activation of toll-like receptor 4 (TLR4), via bacterial lipopolysaccharides (LPS), downregulates SLC22A17. This is similar to the upregulation of Aqp2 by hyperosmolarity/-tonicity and arginine vasopressin (AVP), and downregulation by TLR4 signaling, which occur via the transcription factors NFAT5 (TonEBP or OREBP), cAMP-responsive element binding protein (CREB), and nuclear factor-kappa B, respectively. The aim of the study was to determine the effects of osmolarity/tonicity and AVP, and their associated signaling pathways, on the expression of SLC22A17 and its ligand, LCN2, in the mouse (m) cortical collecting duct cell line mCCD(cl.1). Normosmolarity/-tonicity corresponded to 300 mosmol/L, whereas the addition of 50-100 mmol/L NaCl for up to 72 h induced hyperosmolarity/-tonicity (400-500 mosmol/L). RT-PCR, qPCR, immunoblotting and immunofluorescence microscopy detected Slc22a17 /SLC22A17 and Lcn2 /LCN2 expression. RNAi silenced Nfat5 , and the pharmacological agent 666-15 blocked CREB. Activation of TLR4 was induced with LPS. Similar to Aqp2 , hyperosmotic/-tonic media and AVP upregulated Slc22a17 /SLC22A17, via activation of NFAT5 and CREB, respectively, and LPS/TLR4 signaling downregulated Slc22a17 /SLC22A17. Conversely, though NFAT5 mediated the hyperosmolarity/-tonicity induced downregulation of Lcn2 /LCN2 expression, AVP reduced Lcn2 /LCN2 expression and predominantly apical LCN2 secretion, evoked by LPS, through a posttranslational mode of action that was independent of CREB signaling. In conclusion, the hyperosmotic/-tonic upregulation of SLC22A17 in mCCD(cl.1) cells, via NFAT5, and by AVP, via CREB, suggests that SLC22A17 contributes to adaptive osmotolerance, whereas LCN2 downregulation could counteract increased proliferation and permanent damage of osmotically stressed cells.

GLUT9 is generally considered to be associated with the uric acid transport, which plays an important role in the regulation of serum uric acid level. In this study, the expression level of miR-143-3p was significantly decreased in hyperuricemia mice model group compared with the normal control by miRNA microarray, the same results were confirmed in the hyperuricemia patients and the healthy control group. It is predicted that GLUT9 may be the target gene of miR-143-3p by target scan and other net-software. GLUT9 as the downstream target gene of miR-143-3p was determinated by fluorescence enzyme activity assay. Western blotting and qRT-PCR indicated that the expression of GLUT9 in human renal tubular epithelial cells transfected with miR-143-3p mimics was significantly reduced. Meanwhile inflammatory factors IL-1β and MCP-1 significantly decreased. In conclusion, miR-143-3p can reduce uric acid reabsorption by inhibiting its downstream target gene GLUT9., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Aim: Disturbances of renal medullary perfusion and metabolism have been implicated in the pathogenesis of kidney disease and hypertension. Furosemide, a loop diuretic, is widely used to prevent renal medullary hypoxia in acute kidney disease by uncoupling sodium metabolism, but its effects on medullary perfusion in humans are unknown. We performed quantitative imaging of both renal perfusion and oxygenation using Magnetic Resonance Imaging (MRI) before and during furosemide. Based on the literature, we hypothesized that furosemide would increase medullary oxygenation, decrease medullary perfusion, but cause minor changes (<10%) in renal artery flow (RAF)., Methods: Interleaved measurements of RAF, oxygenation (T 2 *) and perfusion by arterial spin labelling in the renal cortex and medulla of 9 healthy subjects were acquired before and after an injection of 20 mg furosemide. They were preceded by measurements made during isometric exercise (5 minutes handgrip bouts), which are known to induce changes in renal hemodynamics, that served as a control for the sensitivity of the hemodynamic MRI measurements. Experiments were repeated on a second day to establish that the measurements and the induced changes were reproducible., Results: After furosemide, T 2 * values in the medulla increased by 53% (P < 0.01) while RAF and perfusion remained constant. After hand-grip exercise, T 2 * values in renal medulla increased by 22% ± 9% despite a drop in medullary perfusion of 7.2% ± 4.7% and a decrease in renal arterial flow of 17.5% ± 1.7% (P < 0.05). Mean coefficients of variation between repeated measurements for all parameters were 7%., Conclusion: Furosemide induced the anticipated increase in renal medullary oxygenation, attributable exclusively to a decrease in renal oxygen consumption, since no change of RAF, cortical or medullary perfusion could be demonstrated. All measures and the induced changes were reproducible., (© 2019 The Authors. Acta Physiologica published by John Wiley & Sons Ltd on behalf of Scandinavian Physiological Society.)

Diabetes mellitus (DM) remains a great health problem with approximate 30% of patients with DM eventually suffering from diabetic nephropathy. The search for exogenous protective factors has recently received wide attention. The current study aimed to investigate the protective effects of Dendrobium candidum (DC) on kidneys in diabetic rats. Initially, streptozotocin-induced diabetic rats were established and randomly divided into the model group, DC group (0.2, 0.4, and 0.8 g/kg) and irbesartan group (17.5 mg/kg). The biochemical indexes, pathological changes, and the expressions of vascular endothelial growth factor (VEGF), GLUT-1, and CTGF were examined. It was found that as compared with the model group, the kidney index, serum creatine, blood urea nitrogen, 24-hour urine protein, and VEGF of DC treatment groups were significantly decreased, and pathological changes in kidney were improved in the DC groups and irbesartan group ( P < 0.05 for each parameter). The protein and messenger RNA levels of GLUT-1 and CTGF in treatment groups were significantly lower than those in rats' renal cortex without treatment. Our data suggest that DC may protect the kidneys of diabetic rats via regulating expression of VEGF, GLUT-1, and CTGF., (© 2019 Wiley Periodicals, Inc.)

The kidney uses specialized G protein-coupled receptors, including olfactory receptors (ORs), to act as sensors of molecules and metabolites. In the present study, we cloned and studied seven renal ORs, which we previously found to be expressed in the murine renal cortex. As most ORs are orphan receptors, our goal was to identify ligands for these ORs in the hope that this will guide future research into their functional roles. We identified novel ligands for two ORs: Olfr558 and Olfr90. For Olfr558, we confirmed activation by previously reported ligands and identified 16 additional carboxylic acids that activated this OR. The strongest activation of Olfr558 was produced by butyric, cyclobutanecarboxylic, isovaleric, 2-methylvaleric, 3-methylvaleric, 4-methylvaleric, and valeric acids. The primary in vivo source of both butyric and isovaleric acids is gut microbial metabolism. We also identified 14 novel ligands that activated Olfr90, the strongest of which were 2-methyl-4-propyl-1,3-oxathiane, 1-octen-3-ol, 2-octanol, and 3-octanol. Interestingly, 8 of these 14 ligands are of fungal origin. We also investigated the tissue distribution of these receptors and found that they are each found in a subset of "nonsensory" tissues. Finally, we examined the putative human orthologs of Olfr558 and Olfr90 and found that the human ortholog of Olfr558 (OR51E1) has a similar ligand profile, indicating that the role of this OR is likely evolutionarily conserved. In summary, we examined seven novel renal ORs and identified new ligands for Olfr558 and Olfr90, which imply that both of these receptors serve to detect metabolites produced by microorganisms.

Repeated administration of meloxicam can cause kidney damage in cats by mechanisms that remain unclear. Metabolomics and lipidomics are powerful, noninvasive approaches used to investigate tissue response to drug exposure. Thus, the objective of this study was to assess the effects of meloxicam on the feline kidney using untargeted metabolomics and lipidomics approaches. Female young-adult purpose-breed cats were allocated into the control (n = 4) and meloxicam (n = 4) groups. Cats in the control and meloxicam groups were treated daily with saline and meloxicam at 0.3 mg/kg subcutaneously for 17 days, respectively. Renal cortices and medullas were collected at the end of the treatment period. Random forest and metabolic pathway analyses were used to identify metabolites that discriminate meloxicam-treated from saline-treated cats and to identify disturbed metabolic pathways in renal tissue. Our results revealed that the repeated administration of meloxicam to cats altered the kidney metabolome and lipidome and suggest that at least 40 metabolic pathways were altered in the renal cortex and medulla. These metabolic pathways included lipid, amino acid, carbohydrate, nucleotide and energy metabolisms, and metabolism of cofactors and vitamins. This is the first study using a pharmacometabonomics approach for studying the molecular effects of meloxicam on feline kidneys., (© 2019 John Wiley & Sons Ltd.)

Nandrolone decanoate is an anabolic androgenic steroid that is abused worldwide by young athletes and bodybuilders to enhance their physical performance. Many clinical reports among those abusers demonstrated a variety of renal disorders. Lycopene is one of the dietary carotenoids found in fruits like tomato, watermelon, and grapefruit and has attracted considerable attention as an antioxidant. Therefore, the present study was designed to evaluate the protective effect of lycopene against nandrolone decanoate induced renal cortical damage. Forty adult male rats were equally divided into four main groups: group I served as the control, group II received lycopene 4 mg/kg/day, group III received nandrolone 10 mg/kg/week, and group IV received nandrolone and lycopene at a dose similar to the previous groups. At the end of the experiment, urea, creatinine and oxidative stress indicators were measured, then the kidneys were sampled for histopathological and immunohistochemical studies. Sections of the group (ПI) showed variable histopathological alterations in the form of distorted shrunken glomeruli and almost complete loss of the glomerular capillaries, in addition to vacuolation and shedding of the tubular epithelium. In conclusion, these results showed that nandrolone decanoate induced toxic effects in the kidney of rats and lycopene had protective effects versus such evoked renal damage., (Copyright © 2019 Elsevier GmbH. All rights reserved.)

Tolvaptan, a vasopressin type 2 receptor antagonist initially developed to increase free-water diuresis, has been approved for the treatment of autosomal dominant polycystic kidney disease in multiple countries. Furthermore, tolvaptan has been shown to improve the renal functions in rodent models of chronic kidney disease (CKD); however, the underlying molecular mechanisms remain unknown. CKD is characterized by increased levels of oxidative stress, and an antioxidant transcription factor-nuclear factor erythroid 2-related factor 2 (Nrf2)-has been gaining attention as a therapeutic target. Therefore, we investigated the effects of tolvaptan and a well-known Nrf2 activator, bardoxolone methyl (BARD) on Nrf2. To determine the role of tolvaptan, we used a renal cortical collecting duct (mpkCCD) cell line and mouse kidneys. Tolvaptan activated Nrf2 and increased mRNA and protein expression of antioxidant enzyme heme oxygenase-1 (HO-1) in mpkCCD cells and the outer medulla of mouse kidneys. In contrast to BARD, tolvaptan regulated the antioxidant systems via a unique mechanism. Tolvaptan activated the Nrf2/HO-1 antioxidant pathway through phosphorylation of protein kinase RNA-like endoplasmic reticulum kinase (PERK). As a result, tolvaptan and BARD could successfully generate synergistic activating effects on Nrf2/HO-1 antioxidant pathway, suggesting that this combination therapy can contribute to the treatment of CKD.

Aims: β-Hydroxybutyrate (βOHB) is a metabolic intermediate that constitutes about 70% of ketone bodies produced in liver from oxidation of fatty acids released from adipose tissue. A recent study showed that βOHB inhibits HDAC1, 3 and 4 (classes I and IIa) in human embryonic kidney 293 (HEK293) cells. Therefore, βOHB could regulate epigenetics via modulating HDACs. However, little is known about the protective effect of βOHB on renal cells through epigenetics. The aim of this study is to investigate whether βOHB reduces cisplatin-induced nephrotoxicity in human renal cortical epithelial (HRCE) cells by modulating HDACs., Main Methods: In this study, we used human renal cortical epithelial (HRCE) cells. The anti-apoptotic effect of βOHB was evaluated using flow cytometry analysis. The expression of apoptosis-related proteins and HDACs was evaluated by western immunoblot., Key Findings: The results showed that βOHB significantly reduced cisplatin-induced apoptosis in HRCE cells. Furthermore, βOHB significantly reduced cisplatin-induced cleavage of caspase-3, acetylation of histone H3, and phosphorylation of AMP-activated kinase. This anti-apoptotic effect of βOHB was markedly attenuated by an inhibitor of HDAC4/5, and βOHB-mediated suppression of cleavage of caspase3 was significantly blocked by siRNA-induced gene silencing of HDAC5., Significance: βOHB attenuates cisplatin-induced apoptosis by activation of HDAC5 in HRCE cells, suggesting that βOHB may be a new therapeutic agent for cisplatin nephropathy., (Copyright © 2019 Elsevier Inc. All rights reserved.)

This study aimed to investigate the effects of various types of exercise on organic anion transporter 3 (Oat3) function, a major transporter that plays a role in the secretion of a variety of drugs and endogenous compounds. Male Wistar rats were randomly allocated to non-exercise, exhaustive, acute and training exercise groups. The function of Oat3 was assessed by the uptake of [ 3 H]-estrone sulfate ([ 3 H]-ES) into rat renal cortical slices. Acute and training exercises had no effect on [ 3 H]-ES uptake whereas a marked reduction in [ 3 H]-ES uptake occurred immediately after exhaustive exercise. However, the reduction in Oat3 function was gradually recovered at 6 and 24 h after the exercise session. Importantly, the impairment of Oat3 function was associated with a decrease in renal Oat3 protein expression. Our results indicate that exhaustive exercise produces a significant impact on renal organic anion transport function, which in turn could alter the plasma level of drugs and compounds in the body.

The increasing application of carbon nanotubes (CNTs) within environmental, occupational and consumer settings has raised concerns regarding their biosafety and adverse effects on human health. The present study was designed to investigate the possible adverse effect of pristine and functionalized (amylated and polyethelene glycol coated) multi-walled (MW) CNTs on rat kidney with special concern to the histological alterations and the associated oxidative stress, apoptosis and inflammation. Healthy male albino rats (n = 40) were randomly divided into 4 groups: group I (control), group II (pristine MWCNTs), group III (amylated MWCNTs) and group IV [polyethelene glycol (PEG)-coated MWCNTs]. Animals of groups II, III and IV received a single dose of 1 mg/kg body weight of MWCNTs via intra-tracheal (IT) instillation at the beginning of the experiment and all rats were sacrificed after 30 days. Rats in groups II and III showed, nearly similar, renal tissue damage (evidenced by thin collapsed glomeruli, packed mesangial and endothelial cells as well as edematous hemorrhagic glomeruli with apoptotic changes) and functional disruptions (indicated by high serum levels of urea and creatinine) probably through induction of oxidative stress [revealed by high level of the lipid peroxidation marker malondialdehyde (MDA) and lower levels of the antioxidant enzymes superoxide dismutase (SOD) and glutathione peroxidase (GPx)], apoptosis (indicated by high caspase 3 activity), and inflammation (evidenced by high level of IL1β). However, PEG-coated MWCNTs-treated group (group IV) showed nearly normal renal structure and function. It could be concluded that pristine and functionalized amylated MWCNTs have nephrotoxic effect, while PEG-coated MWCNTs had lowest, or none, toxic effects making them safer for therapy and diagnosis of a variety of diseases., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

Acute renal failure complicates renal ischemia-reperfusion (I/R) owing to reactive oxygen species production. Atorvastatin (ATO) has anti-inflammatory and antioxidant properties. The current study investigated whether ATO alleviated damage induced by renal I/R injury in nondiabetic versus diabetic rat models. Thirty-six rats were equally divided into 6 groups: group A1 (nondiabetic sham), group A2 (nondiabetic I/R), group A3 (nondiabetic ATO + I/R), group B1 (diabetic sham), group B2 (diabetic I/R), and group B3 (diabetic ATO + I/R). All groups experienced 45 min of bilateral renal ischemia followed by 24 h of reperfusion. Groups A3 and B3 were treated with single intraperitoneal doses of ATO (10 mg/kg) 30 min before ischemia. Histological analysis of kidney tissues, kidney function tests, and analyses of caspase-3 and CD44 expression and oxidative stress markers were performed to assess tubular injury. Histological analysis revealed marked tubular damage in groups A2 and B2 but improvement in groups A3 and B3. Improvements were also found in groups A3 and B3 for caspase-3 and CD44 expression, kidney function tests, and oxidative stress markers. Our results suggest ATO may ameliorate renal I/R injury differently between nondiabetic and diabetic rats.

Aims: In this animal study, we tried to test the hypothesis that apocynin could play an anti-inflammation role by inhibiting NLRP3/X-linked inhibitor of apoptosis protein (XIAP) signalling and have an effect on antifibrosis in rats with diabetic nephropathy., Main Methods: Diabetic nephropathy rats were induced by tail-vein injection of streptozotocin at 60 mg/kg body weight in sodium citrate buffer (0.01 M, pH 4.5) with unrestricted access to food/water for 12 weeks, and rats with blood glucose levels above 18.0 mM were considered diabetic; the damage index for glomerular mesangial cells damage index was calculated by morphological examinations; protein and mRNA changes were analysed by western blotting immunohistochemistry and real-time quantitative polymerase chain reaction; interstitial fibrosis was assessed and scored using Masson's staining., Key Findings: In rats with diabetic nephropathy, apocynin (1) reduced renal injury and improved renal function; (2) downregulated the expression of NLRP3 in renal cortex; (3) downregulated the expression of XIAP in renal cortex; and (4) attenuated renal fibrosis., Significance: As an inhibitor of reactive oxygen species (ROS), apocynin could downregulate the expression of NLRP3 and XIAP, and alleviate renal fibrosis, which meant not only that ROS was one type of ligands of NLRP3, but also that ROS mechanism and NLRP3 activation might be therapeutic targets in the treatment of diabetic nephropathy in the future., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Glomerular mesangial cell (MC) apoptosis is one of the important mechanisms of glomerulosclerosis, which induces an increased severity of albuminuria and promotes the development of diabetic nephropathy (DN). However, the mechanism by which high glucose (HG) induces MCs apoptosis is not fully understood. In the present study, we investigated the effects of mTOR signalling on apoptosis in cultured MCs exposed to HG and in type I diabetes, and tried to clarify the specific mechanisms underlying these effects. In vitro, exposure of MCs to HG stimulated ROS production, decreased the antioxidant enzyme superoxide dismutase (SOD) activity and glutathione (GSH) level, increased nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity, upregulated P53 expression and Bax/Bcl-2 ratio and enhanced cleavage of caspase 3, resulting in an increase in programmed cell death. Pretreatment of the cells with rapamycin ameliorated oxidative stress, reduced the number of apoptotic cells induced by HG and caused the downstream effects of mTOR activation. In vivo, compared with control rats, diabetic rats had more apoptotic cells in glomeruli. Induction of diabetes increased the level of MDA and NADPH oxidase activity, decreased the SOD activity and GSH level, elevated the Bax/Bcl ratio and P53 expression and activated caspase 3. mTOR inhibitor rapamycin treatment prevented these changes further alleviated albuminuria and improved renal function. Taken together, our data suggest that mTOR plays a key role in mediating ROS-induced MC apoptosis in diabetic nephropathy, and these effects have been associated with the promotion of ROS production by upregulating the antioxidant enzyme and downregulating the NADPH oxidase activity., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The role of autophagy in the kidney and many nephrological diseases has gained prominence in recent years. Much of this research has been focused on markers of autophagy that are static and reveal little about the state of this dynamic pathway. Other mechanistic investigations are limited to in vitro studies, that often provide circumstantial evidence of autophagic flux. Here we describe a method for measuring autophagic flux ex vivo that allows more direct observations to be made in situ regarding the state of autophagic flux within the renal cortex of a single animal., (© 2018 Asian Pacific Society of Nephrology.)

Aims: Cisplatin is a major therapeutic drug for solid tumors, but can cause severe nephrotoxicity. However, the role and therapeutic potential of hydrogen sulfide (H 2 S), an endogenous gasotransmitter, in cisplatin-induced nephrotoxicity remain to be defined., Results: Cisplatin led to the impairment of H 2 S production in vitro and in vivo by downregulating the expression level of cystathionine γ-lyase (CSE), which may contribute to the subsequent renal proximal tubule (RPT) cell death and thereby renal toxicity. H 2 S donors NaHS and GYY4137, but not AP39, mitigated cisplatin-induced RPT cell death and nephrotoxicity. The mechanisms underlying the protective effect of H 2 S donors included the suppression of intracellular reactive oxygen species generation and downstream mitogen-activated protein kinases by inhibiting NADPH oxidase activity, which may be possibly through persulfidating the subunit p47phox. Importantly, GYY4137 not only ameliorated cisplatin-caused renal injury but also added on more anticancer effect to cisplatin in cancer cell lines. Innovation and Conclusion: Our study provides a comprehensive understanding of the role and therapeutic potential of H 2 S in cisplatin-induced nephrotoxicity. Our results indicate that H 2 S may be a novel and promising therapeutic target to prevent cisplatin-induced nephrotoxicity. Antioxid. Redox Signal. 29, 455-470.

Background Renal hypoxia, implicated as crucial factor in onset and progression of chronic kidney disease, may be attributed to reduced nitric oxide because nitric oxide dilates vasculature and inhibits mitochondrial oxygen consumption. We hypothesized that chronic nitric oxide synthase inhibition would induce renal hypoxia. Methods and Results Oxygen-sensitive electrodes, attached to telemeters, were implanted in either renal cortex (n=6) or medulla (n=7) in rats. After recovery and stabilization, baseline oxygenation ( pO 2 ) was recorded for 1 week. To inhibit nitric oxide synthase, N-ω-nitro-l-arginine (L-NNA; 40 mg/kg/day) was administered via drinking water for 2 weeks. A separate group (n=8), instrumented with blood pressure telemeters, followed the same protocol. L-NNA rapidly induced hypertension (165±6 versus 108±3 mm Hg; P<0.001) and proteinuria (79±12 versus 17±2 mg/day; P<0.001). Cortical pO 2 , after initially dipping, returned to baseline and then increased. Medullary pO 2 decreased progressively (up to -19±6% versus baseline; P<0.05). After 14 days of L-NNA, amplitude of diurnal medullary pO 2 was decreased (3.7 [2.2-5.3] versus 7.9 [7.5-8.4]; P<0.01), whereas amplitudes of blood pressure and cortical pO 2 were unaltered. Terminal glomerular filtration rate (1374±74 versus 2098±122 μL/min), renal blood flow (5014±336 versus 9966±905 μL/min), and sodium reabsorption efficiency (13.0±0.8 versus 22.8±1.7 μmol/μmol) decreased (all P<0.001). Conclusions For the first time, we show temporal development of renal cortical and medullary oxygenation during chronic nitric oxide synthase inhibition in unrestrained conscious rats. Whereas cortical pO 2 shows transient changes, medullary pO 2 decreased progressively. Chronic L-NNA leads to decreased renal perfusion and sodium reabsorption efficiency, resulting in progressive medullary hypoxia, suggesting that juxtamedullary nephrons are potentially vulnerable to prolonged nitric oxide depletion.

Background: Diabetes is a proinflammatory state. Fibrosis of the renal glomerulus is the most common cause of end-stage renal disease. Glomerulosclerosis is caused by the accumulation of extracellular matrix (ECM) proteins in the mesangial interstitial space. Mesangial cells are unique stromal cells in the renal glomerulus that form the vascular pole of the renal corpuscle along with the mesangial matrix. Myeloid-derived suppressor cells (MDSCs) are heterogeneous immature myeloid cells that rapidly expand to regulate host immunity during inflammation, infection, and cancer. High concentrations of granulocyte-macrophage colony-stimulating factor (GM-CSF) alone or in combination with other molecules represent the most common ex-vivo protocol for differentiating MDSCs from bone marrow or from peripheral blood mononuclear cells. In this study, we analyzed and characterized the functions of MDSCs under the influence of mouse mesangial cells (MMCs) in a hyperglycemic environment and investigated whether cytokine-induced MDSCs ameliorated renal glomerulosclerosis in diabetic mice., Methods: Cytokine-induced MDSCs were propagated from bone marrow cells cultured with mouse recombinant GM-CSF, IL-6, and IL-1β. Diabetic mice were induced with streptozotocin (STZ) and maintained at a blood glucose concentration exceeding 350 mg/dl. The ECM of the renal cortex and fibronectin expression of MMCs were analyzed through immunohistochemistry and western blotting. Arginase 1 and inducible NO synthase expressions of MDSCs were evaluated using quantitative reverse-transcriptase PCR. Cytokines released from MMCs were examined using a cytokine array assay., Results: MDSCs in the diabetic mice were redistributed from the bone marrow into peripheral organs. An increase in fibronectin production was also observed in the renal glomerulus. MMCs in vitro produced more fibronectin and proinflammatory cytokines, such as macrophage inflammatory protein-2, RANTES, and stromal-cell-derived factor-1, under hyperglycemic conditions. The adoptive transfer of cytokine-induced MDSCs into STZ-induced mice normalized the glomerular filtration rate to reduce the kidney to body weight ratio and decrease fibronectin production in the renal glomerulus, ameliorating renal fibrosis. These results demonstrate the anti-inflammatory properties of cytokine-induced MDSCs and offer an alternative immunotherapy protocol for the management of diabetic nephropathy., Conclusions: The application of cytokine-induced MDSCs provides a promising treatment for renal fibrosis and the prevention of diabetic nephropathy.

Pharmacological inhibition or genetic loss of function defects of the renin angiotensin aldosterone system (RAAS) causes compensatory renin cell hyperplasia and hyperreninemia. The triggers for the compensatory stimulation of renin synthesis and secretion in this situation may be multimodal. Since cyclooxygenase-2 (COX-2) expression in the macula densa is frequently increased in states of a defective RAAS, we have investigated a potential role of COX-2 and its derived prostaglandins for renin expression and secretion in aldosterone synthase-deficient mice (AS -/- ) as a model for a genetic defect of the RAAS. In comparison with wild-type mice (WT), AS -/- mice had 9-fold and 30-fold increases of renin mRNA and of plasma renin concentrations (PRC), respectively. Renin immunoreactivity in the kidney cortex of AS -/- mice was 10-fold higher than in WT. Macula densa COX-2 expression was 5-fold increased in AS -/- kidneys relative to WT kidneys. Treatment of AS -/- mice with the COX-2 inhibitor SC-236 for 1 week lowered both renal renin mRNA and PRC by 70%. Hyperplastic renin cells in AS -/- kidneys were found to express the prostaglandin E 2 receptors EP2 and EP4. Global deletion of EP2 receptors did not alter renin mRNA nor PRC values in AS -/- mice. Renin cell-specific inducible deletion of the EP4 receptor lowered renin mRNA and PRC by 25% in AS -/- mice. Renin cell-specific inducible deletion of the EP4 receptor in combination with global deletion of the EP2 receptor lowered renin mRNA and PRC by 70-75% in AS -/- mice. Lineage tracing of renin-expressing cells revealed that deletion of EP2 and EP4 leads to a preferential downregulation of perivascular renin expression. Our findings suggest that increased macula densa COX-2 activity in AS -/- mice triggers perivascular renin expression and secretion via prostaglandin E 2 .

Lead (Pb), a widely distributed environmental pollutant, is known to induce mitochondrial damage as well as autophagy in vitro and in vivo. In this study, we found that Pb could trigger mitophagy in both HEK293 cells and the kidney cortex of male Kunming mice. However, whether ataxia telangiectasis mutated (ATM) which is reported to be linked with PTEN-induced putative kinase 1 (PINK1)/Parkin pathway (a well-characterized mitophagic pathway) participates in the regulation of Pb-induced mitophagy and its exact role remains enigmatic. Our results indicated that Pb activated ATM in vitro and in vivo, and further in vitro studies showed that ATM could co-localize with PINK1 and Parkin in cytosol and interact with PINK1. Knockdown of ATM by siRNA blocked Pb-induced mitophagy even under the circumstance of enhanced accumulation of PINK1 and mitochondrial Parkin. Intriguingly, elevation instead of reduction in phosphorylation level of PINK1 and Parkin was observed in response to ATM knockdown and Pb did not contribute to the further increase of their phosphorylation level, implying that ATM indirectly regulated PINK1/Parkin pathway. These findings reveal a novel mechanism for Pb toxicity and suggest the regulatory importance of ATM in PINK1/Parkin-mediated mitophagy., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Signaling via the receptor of advanced glycation end products (RAGE)-though complex and not fully elucidated in the setting of diabetes-is considered a key injurious pathway in the development of diabetic nephropathy (DN). We report here that RAGE deletion resulted in increased expression of fibrotic markers (collagen I and IV, fibronectin) and the inflammatory marker MCP-1 in primary mouse mesangial cells (MCs) and in kidney cortex. RNA sequencing analysis in MCs from RAGE -/- and wild-type mice confirmed these observations. Nevertheless, despite these gene expression changes, decreased responsiveness to transforming growth factor-β was identified in RAGE -/- mice. Furthermore, RAGE deletion conferred a more proliferative phenotype in MCs and reduced susceptibility to staurosporine-induced apoptosis. RAGE restoration experiments in RAGE -/- MCs largely reversed these gene expression changes, resulting in reduced expression of fibrotic and inflammatory markers. This study highlights that protection against DN in RAGE knockout mice is likely to be due in part to the decreased responsiveness to growth factor stimulation and an antiapoptotic phenotype in MCs. Furthermore, it extends our understanding of the role of RAGE in the progression of DN, as RAGE seems to play a key role in modulating the sensitivity of the kidney to injurious stimuli such as prosclerotic cytokines., (© 2018 by the American Diabetes Association.)

Background: Chronic kidney disease (CKD) is common in geriatric cats, and the most prevalent pathology is chronic tubulointerstitial inflammation and fibrosis. The cell type predominantly responsible for the production of extra-cellular matrix in renal fibrosis is the myofibroblast, and fibroblast to myofibroblast differentiation is probably a crucial event. The cytokine TGF-β1 is reportedly the most important regulator of myofibroblastic differentiation in other species. The aim of this study was to isolate and characterise renal fibroblasts from cadaverous kidney tissue of cats with and without CKD, and to investigate the transcriptional response to TGF-β1., Results: Cortical fibroblast cultures were successfully established from the kidney tissue of cats with normal kidney function (FCF) and cats with chronic kidney disease (CKD-FCF). Both cell types expressed the mesenchymal markers vimentin, CD44 and CD29, and were negative for the epithelial marker cytokeratin, mesangial cell marker desmin and endothelial cell marker vWF. Only CKD-FCF expressed VCAM-1, a cell marker associated with inflammation. Incubation with TGF-β1 (0-10 ng/ml) induced a concentration dependent change in cell morphology, and upregulation of myofibroblast marker gene α-SMA expression alongside collagen 1α1, fibronectin, TGF-β1 and CTGF mRNA. These changes were blocked by the TGF-β1 receptor 1 antagonist SB431542 (5 μM)., Conclusions: FCF and CKD-FCF can be cultured via a simple method and represent a model for the investigation of the progression of fibrosis in feline CKD. The findings of this study suggest TGF-β1 may be involved in fibroblast-myofibroblast transition in feline CKD, as in other species.

Obesity-related glomerulopathy (ORG) is morphologically characterized by glomerulomegaly with or without observable focal segmental glomerulosclerosis under light microscope, with decreased podocyte density and number, and with increased foot‑process width observed under electron microscope. The severity of podocyte injury is correlated with the degree of proteinuria and renal dysfunction. However, the pathogenesis of ORG is not well understood. The aim of the present study was to explore the possible pathogenic role of aldosterone (ALDO) in ORG. In the in vivo animal experiments, body weight, Lee's obesity index, abdominal fat index, urinary protein excretion, average glomerular diameter were significantly increased, the mRNA and protein expression of podocyte‑associated molecules including nephrin, podocin, podoplanin and podocalyxin were significantly reduced, and the Wnt/β‑catenin signaling pathway was activated in ORG model mice compared with the Control mice, whereas the administration of spironolactone significantly ameliorated these effects. In the in vitro experiments on cultured podocytes, the mRNA and protein expression levels of the aforementioned podocyte‑associated molecules were significantly downregulated and the Wnt/β‑catenin signaling pathway was activated following ALDO stimulation, whereas eplerenone significantly attenuated all the above effects. Dickkopf‑related protein 1 (DKK1), an inhibitor of Wnt/β‑catenin signaling pathway, also reduced the effects of ALDO exposure on the expression of podocyte‑associated molecules. The present study hypothesized that ALDO may be involved in the pathogenesis of ORG through the activation of Wnt/β‑catenin signaling pathway in podocytes.

This study aimed at investigating the possible ameliorative effects of sitagliptin in mice with gentamicin (GEN) nephrotoxicity. Sitagliptin was given to the animals at an oral dose of 10mgkg -1 per day for 10days, and in some of these mice, GEN was injected intraperitoneally at a dose of 100mgkg -1 per day during the last seven days of the treatment. Nephrotoxicity was evaluated histopathologically by light microscopy and biochemically by measuring several indices in plasma, urine and renal cortex homogenates. GEN treatment induced nephrotoxicity as evidenced by significantly (P<0.0001) increasing the plasma concentrations of urea, creatinine, circulatory cytokines, cystatin C, sclerostin, and TNFα. Treatment with GEN also significantly elevated urinary N-acetyl-β-d glucosaminidase (NAG) concentration (P<0.0001). Moreover, GEN caused significant increase in oxidative stress in the kidneys (P<0.0001). Histopathological examination revealed massive tubular injury, necrosis, infiltration of inflammatory cells and intraluminal hyaline casts in mice treated with GEN. Sitagliptin alone did not significantly affect any of the indices measured. However, concomitant treatment with sitagliptin and GEN significantly mitigated most of the nephrotoxic actions of GEN. Pending further studies, sitagliptin may potentially be useful as a nephroprotectant agent., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)

Therapeutic use of cisplatin (CP), an effective anticancer drug, is limited by dose dependent nephrotoxicity. Thymoquinone (TQ), the major Nigella sativa seed oil constituent has been shown to prevent progression of various renal disorders. The present study investigates the protective effect of TQ on CP-induced nephrotoxicity. Rats were divided into six groups viz. control, CP, CPTQ 1 , CPTQ 2 , CPTQ 3 , and TQ alone group. Animals in CP and TQ combination groups were administered TQ (0.5, 1.5, and 3 mg/kg bwt, orally) with single intraperitoneal dose of CP (6 mg/kg bwt). The effect of TQ administration was determined on CP-induced alterations in various serum/urine parameters and on the enzymes of brush border membrane enzyme (BBM), carbohydrate metabolism, and antioxidant defense system in renal cortex and medulla. Oral administration of TQ in all the three doses prior to and following a single dose CP treatment caused significant recovery of serum creatinine and blood urea nitrogen levels; however, maximum recovery was seen in CPTQ 2 group. TQ administration averted CP-induced decline in BBM activities, both in the cortical and medullary homogenates and in isolated BBM vesicles. TQ administration also ameliorated CP-induced impairments in renal metabolic and antioxidant status. Histopathological studies supported these biochemical findings. TQ ameliorates CP-induced oxidative damage owing to its intrinsic antioxidant properties.

The potential toxic effects of silver nanoparticles (AgNPs), administered by a single intratracheal instillation (i.t), was assessed in a rat model using commercial physico-chemical characterized nanosilver. Histopathological changes, overall toxic response and oxidative stress (kidney and plasma protein carbonylation), paralleled by ultrastructural observations (TEM), were evaluated to examine renal responses 7 and 28 days after i.t. application of a low AgNP dose (50 µg/rat), compared to an equivalent dose of ionic silver (7 µg AgNO₃/rat). The AgNPs caused moderate renal histopathological and ultrastructural alteration, in a region-specific manner, being the cortex the most affected area. Notably, the bulk AgNO₃, caused similar adverse effects with a slightly more marked extent, also triggering apoptotic phenomena. Specifically, 7 days after exposure to both AgNPs and AgNO₃, dilatation of the intercapillary and peripheral Bowman's space was observed, together with glomerular shrinkage. At day 28, these effects still persisted after both treatments, accompanied by an additional injury involving the vascular component of the mesangium, with interstitial micro-hemorrhages. Neither AgNPs nor AgNO₃ induced oxidative stress effects in kidneys and plasma, at either time point. The AgNP-induced moderate renal effects indicate that, despite their benefits, novel AgNPs employed in consumer products need exhaustive investigation to ensure public health safety., Competing Interests: The authors declare no conflict of interest.

This study examined the mechanism of release of endogenous acetylcholine (ACh) in rabbit renal cortex by applying a microdialysis technique. In anesthetized rabbits, a microdialysis probe was implanted into the renal cortex and perfused with Ringer's solution containing high potassium concentration, high sodium concentration, a Na + /K + -ATPase inhibitor (ouabain), or an epithelial Na + channel blocker (benzamil). Dialysate samples were collected at baseline and during exposure to each agent, and ACh concentrations in the samples were measured by high-performance liquid chromatography. High potassium had no effect on renal ACh release. High sodium increased dialysate ACh concentrations significantly. Ouabain increased dialysate ACh concentration significantly. Benzamil decreased dialysate ACh concentrations significantly both at baseline and under high sodium. The finding that high potassium-induced depolarization does not increase ACh release suggests that endogenous ACh is released in renal cortex mainly by non-neuronal mechanism. Sodium ion transport may be involved in the non-neuronal ACh release.

Acute kidney injury (AKI), resulting from chemotherapeutic agents such as cisplatin, remains an obstacle in the treatment of cancer. Cisplatin-induced AKI involves apoptotic and necrotic cell death, pathways regulated by sphingolipids such as ceramide and glucosylceramide. Results from this study indicate that C57BL/6J mice treated with cisplatin had increased ceramide and hexosylceramide levels in the renal cortex 72 h following cisplatin treatment. Pretreatment of mice with inhibitors of acid sphingomyelinase and de novo ceramide synthesis (amitriptyline and myriocin, respectively) prevented accumulation of ceramides and hexosylceramide in the renal cortex and protected from cisplatin-induced AKI. To determine the role of ceramide metabolism to hexosylceramides in kidney injury, we treated mice with a potent and highly specific inhibitor of glucosylceramide synthase, the enzyme responsible for catalyzing the glycosylation of ceramides to form glucosylceramides. Inhibition of glucosylceramide synthase attenuated the accumulation of the hexosylceramides and exacerbated ceramide accumulation in the renal cortex following treatment of mice with cisplatin. Increasing ceramides and decreasing glucosylceramides in the renal cortex sensitized mice to cisplatin-induced AKI according to markers of kidney function, kidney injury, inflammation, cell stress, and apoptosis. Under conditions of high ceramide generation, data suggest that metabolism of ceramides to glucosylceramides buffers kidney ceramides and helps attenuate kidney injury.-Dupre, T. V., M. A. Doll, P. P. Shah, C. N. Sharp, D. Siow, J. Megyesi, J. Shayman, A. Bielawska, J. Bielawski, L. J. Beverly, M. Hernandez-Corbacho, C. J. Clarke, A. J. Snider, R. G. Schnellmann, L. M. Obeid, Y. A. Hannun, and L. J. Siskind. Inhibiting glucosylceramide synthase exacerbates cisplatin-induced acute kidney injury. J. Lipid Res 2017. 58: 1439-1452.

Lysophosphatidic acid (LPA) is known to regulate various biological responses by binding to LPA receptors. The serum level of LPA is elevated in diabetes, but the involvement of LPA in the development of diabetes and its complications remains unknown. Therefore, we studied LPA signaling in diabetic nephropathy and the molecular mechanisms involved. The expression of autotaxin, an LPA synthesis enzyme, and LPA receptor 1 was significantly increased in both mesangial cells (SV40 MES13) maintained in high-glucose media and the kidney cortex of diabetic db/db mice. Increased urinary albumin excretion, increased glomerular tuft area and volume, and mesangial matrix expansion were observed in db/db mice and reduced by treatment with ki16425, a LPA receptor 1/3 antagonist. Transforming growth factor (TGF)β expression and Smad-2/3 phosphorylation were upregulated in SV40 MES13 cells by LPA stimulation or in the kidney cortex of db/db mice, and this was blocked by ki16425 treatment. LPA receptor 1 siRNA treatment inhibited LPA-induced TGFβ expression, whereas cells overexpressing LPA receptor 1 showed enhanced LPA-induced TGFβ expression. LPA treatment of SV40 MES13 cells increased phosphorylated glycogen synthase kinase (GSK)3β at Ser9 and induced translocation of sterol regulatory element-binding protein (SREBP)1 into the nucleus. Blocking GSK3β phosphorylation inhibited SREBP1 activation and consequently blocked LPA-induced TGFβ expression in SV40 MES13 cells. Phosphorylated GSK3β and nuclear SREBP1 accumulation were increased in the kidney cortex of db/db mice and ki16425 treatment blocked these pathways. Thus, LPA receptor 1 signaling increased TGFβ expression via GSK3β phosphorylation and SREBP1 activation, contributing to the development of diabetic nephropathy., (Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Gentamicin nephrotoxicity accounts for 10%-15% of all cases of acute renal failure. Several natural antioxidants were found to be effective against drug-induced toxicity. The possible protective effects of lycopene (Lyc) and rosmarinic acid (RA) alone or combined on gentamicin (Gen) induced renal cortical oxidative stress, apoptosis, and autophagy were evaluated. Sixty-three rats were randomly divided into seven groups named: control, group II received RA 50 mg/kg/day, group III received Lyc 4 mg/kg/day, group IV received Gen 100 mg/kg/day, group V (RA + Gen), group VI (Lyc + Gen), and group VII (RA + Lyc + Gen). At the end of the experiment, kidney functions were estimated then the kidneys were sampled for histopathological, immunohistochemistry, and biochemical studies. Administration of rosmarinic acid and lycopene decreased elevated serum creatinine, blood urea nitrogen, renal malondialdehyde and immunoexpression of the proapoptotic protein (Bax), autophagic marker protein (LC3/B), and inducible nitric oxide synthase (iNOS) induced by gentamicin. They increased reduced glutathione, glutathione peroxidase, superoxide dismutase, and immunoexpression of the antiapoptotic protein (Bcl2). They also improved the histopathological changes induced by gentamicin. The combination therapy of rosmarinic acid and lycopene shows better protective effects than the corresponding monotherapy. Anat Rec, 300:1137-1149, 2017. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Recently, gold nanoparticles (GNPs) have shown promising applications in targeted drug delivery and contrast imaging. Although in vitro cytotoxicity of GNPs has been thoroughly studied, there are limited data on in vivo toxicity of GNPs. In this study, we evaluated the effects of intraperitoneally injected 10 nm and 50 nm GNPs (5 μg/animal) on the expression of proinflammatory cytokines (IL-1β, IL-6 and TNF-α) on day 1 and day 5, post-exposure. The results of immunohistochemistry showed that both 10 nm and 50 nm GNPs induced an acute phase expression of proinflammatory cytokines in renal cortex and medulla. This proinflammatory response was comparatively more intense in renal medulla than cortex. All the three cytokines were undetectable in control cortex and medulla. In conclusion, both 10 nm and 50 nm GNPs caused an acute phase induction of proinflammatory cytokines in cortex and medulla of rat kidneys. An intense immunostaining of proinflammatory cytokines in renal medulla warrants further studies to evaluate the nephrotoxicity of GNPs to validate the safe application of GNPs for contrast imaging in renal insufficiency.

This study determined the comparative nephrotoxic potential of four trichloronitrobenzenes (TCNBs) (2,3,4-; 2,4,5-; 2,4,6-; and 3,4,5-TCNB) and explored the effects of antioxidants and biotransformation inhibitors on TCNB-induced cytotoxicity in isolated renal cortical cells (IRCC) from male Fischer 344 rats. IRCC were incubated with a TCNB up to 1.0 mM for 15-120 min. Pretreatment with an antioxidant or cytochrome P450 (CYP), flavin monooxygenase (FMO), or peroxidase inhibitor was used in some experiments. Among the four TCNBs, the order of decreasing nephrotoxic potential was approximately 3,4,5- > 2,4,6- > 2,3,4- > 2,4,5-TCNB. The four TCNBs exhibited a similar profile of attenuation of cytotoxicity in response to antioxidant pretreatments. 2,3,4- and 3,4,5-TCNB cytotoxicity was attenuated by most of the biotransformation inhibitors tested, 2,4,5-TCNB cytotoxicity was only inhibited by isoniazid (CYP 2E1 inhibitor), and 2,4,6-TCNB-induced cytotoxicity was inhibited by one CYP inhibitor, one FMO inhibitor, and one peroxidase inhibitor. All of the CYP specific inhibitors tested offered some attenuation of 3,4,5-TCNB cytotoxicity. These results indicate that 3,4,5-TCNB is the most potent nephrotoxicant, free radicals play a role in the TCNB cytotoxicity, and the role of biotransformation in TCNB nephrotoxicity in vitro is variable and dependent on the position of the chloro groups., Competing Interests: The authors declare no conflict of interest.

Short-chain fatty acids (SCFAs), such as acetate, propionate, and butyrate, are produced predominantly by gut microbiota fermentation of dietary fiber. SCFAs are newly identified as endogenous ligands of two orphan G protein-coupled receptors, GPR41 and GPR43, which have the potential to modulate inflammation. Therefore, GPR41 and GPR43 may mediate the link between the gut microbiome status and various disease conditions including renal inflammation. This study aimed at investigating whether SCFAs activate GPR41 and GPR43, and thereby exert anti-inflammatory effects in human renal cortical epithelial cells (HRCEs) as a main component of kidney tissue. Immunohistochemical analyses of human renal biopsy specimens revealed the expression of GPR41 and GPR43 protein in the distal renal tubules and collecting tubules. TNF-α increased the expression of monocyte chemoattractant protein-1 (MCP-1), a potential fibrotic inducer, at least partly via enhancing phosphorylation of p38 and JNK in HRCEs. SCFAs, especially propionate, attenuated TNF-α- stimulated MCP-1 expression by inhibiting the phosphorylation of p38 and JNK. This inhibitory effect was considerably attenuated by an inactivator of the Gi/o-type G protein and a Gβγ (i/o) blocker, but not by a Gα (i/o) blocker. Furthermore, SCFA-mediated inhibition of MCP-1 expression was significantly blocked by siRNA-induced gene silencing of GPR41 and GPR43. In conclusion, SCFAs lowered TNF-α-induced MCP-1 expression by reducing phosphorylation of p38 and JNK in a GPR41/43-dependent manner in HRCEs, suggesting that SCFA modification may be a new therapeutic tool for preventing progression of renal inflammation and fibrosis., (Copyright © 2017 Elsevier Inc. All rights reserved.)