Metformin improves relevant disease parameters in an autosomal dominant polycystic kidney disease mouse model.

Autosomal dominant polycystic kidney disease (ADPKD), caused by mutations in the polycystin 1 (PKD1) or polycystin 2 genes, presents with progressive development of kidney cysts and eventual end-stage kidney disease with limited treatment options. Previous work has shown that metformin reduces cyst growth in rapid ADPKD mouse models via inhibition of cystic fibrosis transmembrane conductance regulator-mediated fluid secretion, mammalian target of rapamycin, and cAMP pathways. The present study importantly tested the effectiveness of metformin as a therapy for ADPKD in a more clinically relevant Pkd1RC/RC mouse model, homozygous for the R3277C knockin point mutation in the Pkd1 gene. This mutation causes ADPKD in humans. Pkd1RC/RC male and female mice, which have a slow progression to end-stage kidney disease, received metformin (300 mg/kg/day in drinking water vs. water alone) from 3 to 9 or 12 mo of age. As previously reported, Pkd1RC/RC females had a more severe disease phenotype as compared with males. Metformin treatment reduced the ratio of total kidney weight-to-body weight relative to age-matched and sex-matched untreated controls at both 9 and 12 mo and reduced the cystic index in females at 9 mo. Metformin also increased glomerular filtration rate, lowered systolic blood pressure, improved anemia, and lowered blood urea nitrogen levels relative to controls in both sexes. Moreover, metformin reduced gene expression of key inflammatory markers and both gene and protein expression of kidney injury marker-1 and cyclin-dependent kinase-1 versus untreated controls. Altogether, these findings suggest several beneficial effects of metformin in this highly relevant slowly progressive ADPKD mouse model, which may help inform new ADPKD therapies in patients.NEW & NOTEWORTHY Metformin treatment improved ADPKD disease severity in a relevant, slowly progressive ADPKD mouse model that recapitulates a PKD-associated PKD1 mutation. Relative to controls, metformin reduced kidney weight/body weight, cystic index and BUN levels, while improving GFR, blood pressure and anemia. Metformin also reduced key inflammatory and injury markers, along with cell proliferation markers. These findings suggest several beneficial effects of metformin in this ADPKD mouse model, which may help inform new ADPKD therapies in patients.

Semaporin3A inhibitor ameliorates renal fibrosis through the regulation of JNK signaling.

Renal fibrosis is the common pathological pathway in progressive renal diseases. In the present study, we analyzed the roles of semaphorin 3 A (SEMA3A) on renal fibrosis and the effect of SEMA3A inhibitor (SEMA3A-I) using a unilateral ureteral obstruction (UUO) mouse model. Expression of SEMA3A in the proximal tubulus and neuropilin-1, a recepor of SEMA3A, in fibloblast and tubular cells were increased in UUO kidneys. The expression of myofibroblast marker tenascin-C and fibronection as well as renal fibrosis were increased in UUO kidneys, all of which were ameliorated by SEMA3A-I. In addition, the JNK signaling pathway, known as the target of SEMA3A signaling, was activated in proximal tubular cells and fibroblast cells after UUO surgery, and SEMA3A-I significantly attenuated the activation. In vitro, treatments with SEMA3A as well as transforming growth factor-ß1 (TGF-ß1) in human proximal tubular cells lost epithelial cell characteristics, and SEMA3A-I significantly ameliorated this transformation. The JNK inhibitor SP600125 partially reversed SEMA3A and TGF-ß1-induced cell transformation, indicating that JNK signaling is involved in SEMA3A-induced renal fibrosis. In addition, treatment with SEMA3A in fibroblast cells activated expression of tenascin-C, collagen type I, and fibronection, indicating that SEMA3A may accelerate renal fibrosis through the activation of fibroblast cells. Analysis of human data revealed the positive correlation between urinary SEMA3A and urinary N-acetyl-ß-d-glucosaminidase, indicating the association between SEMA3A and tubular injury. In conclusion, SEMA3A signaling is involved in renal fibrosis through the JNK signaling pathway and SEMA3A-I might be a therapeutic option for protecting from renal fibrosis.NEW & NOTEWORTHY Renal fibrosis is the common pathological pathway in the progression of renal diseases. This study, using a unilateral ureteral obstruction (UUO) mouse model, indicated increased semaphorin3A (SEMA3A) signaling in renal tubular cells as well as fibroblast cells under UUO surgery, and SEMA3A inhibitor ameliorated UUO-induced renal fibrosis through the regulation of JNK signaling. The study proposes the potential therapeutic option of SEMA3A inhibitor to treat renal fibrosis.

Phase 1/2 Study of Lumasiran for Treatment of Primary Hyperoxaluria Type 1: A Placebo-Controlled Randomized Clinical Trial.

BACKGROUND AND OBJECTIVES: In the rare disease primary hyperoxaluria type 1, overproduction of oxalate by the liver causes kidney stones, nephrocalcinosis, kidney failure, and systemic oxalosis. Lumasiran, an RNA interference therapeutic, suppresses glycolate oxidase, reducing hepatic oxalate production. The objective of this first-in-human, randomized, placebo-controlled trial was to evaluate the safety, pharmacokinetic, and pharmacodynamic profiles of lumasiran in healthy participants and patients with primary hyperoxaluria type 1. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: This phase 1/2 study was conducted in two parts. In part A, healthy adults randomized 3:1 received a single subcutaneous dose of lumasiran or placebo in ascending dose groups (0.3-6 mg/kg). In part B, patients with primary hyperoxaluria type 1 randomized 3:1 received up to three doses of lumasiran or placebo in cohorts of 1 or 3 mg/kg monthly or 3 mg/kg quarterly. Patients initially assigned to placebo crossed over to lumasiran on day 85. The primary outcome was incidence of adverse events. Secondary outcomes included pharmacokinetic and pharmacodynamic parameters, including measures of oxalate in patients with primary hyperoxaluria type 1. Data were analyzed using descriptive statistics. RESULTS: Thirty-two healthy participants and 20 adult and pediatric patients with primary hyperoxaluria type 1 were enrolled. Lumasiran had an acceptable safety profile, with no serious adverse events or study discontinuations attributed to treatment. In part A, increases in mean plasma glycolate concentration, a measure of target engagement, were observed in healthy participants. In part B, patients with primary hyperoxaluria type 1 had a mean maximal reduction from baseline of 75% across dosing cohorts in 24-hour urinary oxalate excretion. All patients achieved urinary oxalate levels ≤1.5 times the upper limit of normal. CONCLUSIONS: Lumasiran had an acceptable safety profile and reduced urinary oxalate excretion in all patients with primary hyperoxaluria type 1 to near-normal levels. CLINICAL TRIAL REGISTRY NAME AND REGISTRATION NUMBER: Study of Lumasiran in Healthy Adults and Patients with Primary Hyperoxaluria Type 1, NCT02706886.

Phosphoproteomic identification of vasopressin V2 receptor-dependent signaling in the renal collecting duct.

Vasopressin controls water balance largely through PKA-dependent effects to regulate the collecting duct water channel aquaporin-2 (AQP2). Although considerable information has accrued regarding the regulation of water and solute transport in collecting duct cells, information is sparse regarding the signaling connections between PKA and transport responses. Here, we exploited recent advancements in protein mass spectrometry to perform a comprehensive, multiple-replicate analysis of changes in the phosphoproteome of native rat inner medullary collecting duct cells in response to the vasopressin V2 receptor-selective agonist 1-desamino-8D-arginine vasopressin. Of the 10,738 phosphopeptides quantified, only 156 phosphopeptides were significantly increased in abundance, and only 63 phosphopeptides were decreased, indicative of a highly selective response to vasopressin. The list of upregulated phosphosites showed several general characteristics: 1) a preponderance of sites with basic (positively charged) amino acids arginine (R) and lysine (K) in position -2 and -3 relative to the phosphorylated amino acid, consistent with phosphorylation by PKA and/or other basophilic kinases; 2) a greater-than-random likelihood of sites previously demonstrated to be phosphorylated by PKA; 3) a preponderance of sites in membrane proteins, consistent with regulation by membrane association; and 4) a greater-than-random likelihood of sites in proteins with class I COOH-terminal PDZ ligand motifs. The list of downregulated phosphosites showed a preponderance of those with proline in position +1 relative to the phosphorylated amino acid, consistent with either downregulation of proline-directed kinases (e.g., MAPKs or cyclin-dependent kinases) or upregulation of one or more protein phosphatases that selectively dephosphorylate such sites (e.g., protein phosphatase 2A). The phosphoproteomic data were used to create a web resource for the investigation of G protein-coupled receptor signaling and regulation of AQP2-mediated water transport.

The Effect of Tadalafil on Renal Fibrosis Induced by Ureteral Obstruction / Efecto del Tadalafil en la Fibrosis Renal Inducida por Obstrucción Uretral

ABSTRACT Objective: It has been reported that phosphodiesterase-5 (PDE-5) inhibitors improve kidney function during acute and chronic renal failure. This study aimed to determine the possible therapeutic effects of tadalafil, a specific PDE-5 inhibitor, on renal fibrosis induced by unilateral ureteral obstruction (UUO). Methods: Male Sprague-Dawley rats were used and randomly divided into three groups (n = 6) as sham-operated, UUO and tadalafil-treated (10 mg/72 hours, ig) UUO (UUO+T) groups. Unilateral ureteral obstruction was induced by complete ligation of the left ureter and 14 days after surgery creatinine clearance, urinary cyclic guanosine monophosphate (cGMP), renal alpha-smooth muscle actin (α-sma) and transforming growth factor βeta (TGF-β) levels, as well as histologic changes, were observed in all the animals. Results: Unilateral ureteral obstruction-induced renal fibrosis was confirmed by increased α-sma level, collagen deposition, tubular dilation, inflammatory cell infiltration and necrosis. An increased renal TGF-β level and decreased urinary cGMP level was also observed in obstructed animals in addition to reduced creatinine clearance. Tadalafil treatment, which restored the animals 'urinary cGMP level, significantly attenuated the fibrotic changes and TGF-β increase in their kidneys. Conclusion: This study suggests that tadalafil treatment ameliorates renal fibrosis by reducing TGF-β expression and may have important clinical relevance since tadalafil is currently used clinically to treat erectile dysfunction and pulmonary hypertension.

RESUMEN Objetivo: Se ha reportado que los inhibidores de la fosfodiesterasa-5 (PDE-5) mejoran las funciones renales durante la insuficiencia renal aguda y crónica. Este estudio tuvo por objetivo determinar los posibles efectos terapéuticos del tadalafil - un inhibidor específico de la PDE-5 - sobre la fibrosis renal inducida por una obstrucción ureteral unilateral (OUU). Métodos: Se utilizaron ratas machos Sprague-Dawley, divididas de manera aleatoria en tres grupos (n = 6): operación simulada, OUU y tratamiento con tadalafil (10 mg/72 horas, IG), y OUU (OUU+T). La obstrucción uretral unilateral fue inducida por una ligadura completa del uréter izquierdo y 14 días después de la cirugía, se observaron niveles de monofosfato de guanosina cíclico (GMP) urinario, alfa-actina de músculo liso (α-SMA), y factor de crecimiento transformante βeta (FCT-β), así como cambios histológicos en todos los animales. Resultados: La fibrosis renal inducida por obstrucción uretral unilateral fue confirmada por un aumento del nivel de α-SMA, deposición de colágeno, dilatación tubular, infiltración de células inflamatorias y necrosis. También se observó un aumento del nivel de FCT-β renal y una disminución del nivel de GMP urinario en los animales con obstrucción, además de una reducción del aclaramiento de la creatinina. El tratamiento con tadalafil, que restauró el nivel de GMP urinario de los animales, atenuó significativamente los cambios fibróticos y el aumento de FCT-β en los riñones. Conclusión: Este estudio sugiere que el tratamiento con tadalafil mejora la fibrosis renal al reducir la expresión de FCT-β y puede tener una importante relevancia clínica por cuanto el tadalafil se usa hoy día clínicamente para tratar la disfunción eréctil y la hipertensión pulmonar.

Discovery of Potent, Selective, and Short-Acting Peptidic V2 Receptor Agonists.

The vasopressin analogue desmopressin (desamino-d-arginine8 vasopressin, dDAVP, 1) is a potent vasopressin 2 (V2) receptor (V2R) agonist approved in many countries for the treatment of diabetes insipidus, primary nocturnal enuresis, nocturia, and coagulation disorders. Since 1 is primarily excreted via the kidneys, an age-related decline in kidney function leads to slower elimination, prolonged antidiuresis, and hyponatremia. In search of novel, potent, selective, and short-acting peptidic V2R agonists, we synthesized a series of C-terminally truncated analogues of [Val4]dDAVP, 2, modified in positions 2, 3, and 7 and/or at the disulfide bridge. The peptides were evaluated for in vitro potency at the human V2 receptor, selectivity versus the related receptors (human vasopressin 1a receptor, human vasopressin 1b receptor, and human oxytocin receptor), and pharmacokinetic profiles in rodents and other higher species. The truncated analogues show excellent potency at the V2R, increased systemic clearance, and shorter half-life in rats. Two compounds 19 (c(Bua-Cpa-Thi-Val-Asn-Cys)-Pro-Agm) and 38 (c(Bua-Cpa-Thi-Val-Asn-Cys)-Pro-d-Arg-NEt2) have been selected for clinical development for nocturia.

Drosophila Renal Organ as a Model for Identification of Targets and Screening of Potential Therapeutic Agents for Diabetic Nephropathy.

Diabetic nephropathy is one of the major causes of kidney failure, accounting for ~44% of all cases. In spite of the significant mortality rate of diabetic nephropathy, specific and effective treatment is still eluding. Identification of genetic determinants and understanding their role in the progression of disease are essential for developing diagnostic tools and effective therapy. Drosophila melanogaster is a genetically tractable model organism and is being used for understanding the genetic basis of several human diseases. Drosophila has a well developed renal system and shares conserved developmental and functional processes with humans. Apart from similarities in renal system, type 1 and type 2 diabetes can be induced in Drosophila following mechanisms similar to those in human. This review discusses the current therapies available for diabetic nephropathy and examines the potential of Drosophila renal system as a model for identifying drug targets for diabetic nephropathy and screening of the potential drugs for their efficacy.

Aquaporin-2 plays an important role in water transportation through the bladder wall in rats.

AIM: We investigated the role of the bladder wall in permeating water, focusing on aquaporins. METHODS: Female Sprague-Dawley rats weighing 300 g were used to investigate the role of the bladder wall in saline permeation. Changes in intravesical fluid volume and sodium concentration were measured in the desmopressin acetate hydrate-loaded and control groups 3 h after administration. Bladders were resected to measure aquaporin-1, 2, and 3 gene expression using qRT-PCR. Additionally, the change of aquaporin-2 expression was measured using Western blotting and immunohistochemistry in intravesical aquaporin-2 siRNA-treated and control groups. RESULTS: Although the intravesical fluid volume and sodium concentration significantly decreased from 0 to 3 h (1.00 ± 0.00 vs 0.83 ± 0.08 mL, 157.80 ± 1.30 vs 146.8 ± 1.92 mEq/mL, P < 0.01, respectively in the control group), administration of desmopressin did not affect the extent of volume change. Aquaporin-2 expression was significantly higher in the 3-h distended bladders than in the empty bladder. Aquaporin-2 siRNA treatment suppressed aquaporin-2 expression and the change of intravesical fluid volume from 0 to 3 h (1.00 ± 0.00 and 0.99 ± 0.02 mL), which was related to the suppression of sodium concentration change in comparison with control siRNA treatment (149.6 ± 2.4 vs 143.6 ± 3.67 mEq/mL, P < 0.05). CONCLUSIONS: The rat urinary bladder absorbs water and salts under the full-filled condition. Aquaporin-2 plays an important role in the transport of water, accompanied by sodium concentration change. We demonstrated a part of the bladder absorption mechanism, which may lead to development of a new method for regulating bladder storage function.

Salvianolic acid A ameliorates renal ischemia/reperfusion injury by activating Akt/mTOR/4EBP1 signaling pathway.

Salvianolic acid A (Sal A) has been shown to prevent and treat ischemic cardiovascular, as well as cerebral vascular diseases. However, little is known about Sal A in renal ischemia/reperfusion (I/R) injury. In this study, a renal I/R injury model in rats and a hypoxia/reoxygenation (H/R) model to damage proximal renal tubular cells (HK-2) were used to assess whether Sal A halts the development and progression of renal I/R injury. As compared with vehicle treatment, Sal A significantly attenuated kidney injury after renal I/R injury, accompanied by decreases in plasma creatinine, blood urea nitrogen levels, the number of apoptosis-positive tubular cells, and kidney oxidative stress. Sal A also activated phosphorylated protein kinase B (p-Akt) and phosphorylated-mammalian target of rapamycin (p-mTOR) compared with vehicle-treated I/R injury rats. In H/R-injured HK-2 cells, Sal A can reduce the levels of reactive oxygen species in a dose-related manner. Similar to the results from in vivo experiments, in vitro Sal A also increased the protein expression of phosphorylated-eukaryotic initiation factor 4E binding protein 1 (p-4EBP1) compared with vehicle. Furthermore, the cytoprotective activity of Sal A was inhibited by LY294002 and rapamycin. These findings indicate that Sal A can ameliorate renal I/R injury and promote tubular cell survival partly via the Akt/mTOR/4EBP1pathway. Sal A could be a candidate compound to prevent ischemic tissue damage.

Concomitant use of rapamycin and rosiglitazone delays the progression of polycystic kidney disease in Han:SPRD rats: a study of the mechanism of action.

Attributing to their antiproliferative effect, both rapamycin and peroxisome proliferator-activated receptor-γ (PPARγ) can halt the progression of autosomal dominant polycystic kidney disease (ADPKD). Whether combined use could enhance this effect is unknown. The present study used rapamycin and the PPARγ agonist rosiglitazone concomitantly to observe their combined effects on the proliferation of ADPKD cyst-lining epithelial cells and the progression of ADPKD in Han:SPRD rats. Concomitant use of the two drugs inhibited the proliferation of WT9-12 cells significantly through a superimposition effect. Rosiglitazone inhibited the phosphorylation of mammalian target of rapamycin p70S6K. Concomitant use of rosiglitazone and rapamycin further downregulated the p-p70S6K level. Rosiglitazone also inhibited the phosphorylation of Akt and antagonized the activation of Akt induced by rapamycin. Concomitant use of rosiglitazone and rapamycin significantly retarded the deterioration of renal function, decreased cyst cell proliferation and interstitial fibrosis in Han:SPRD rats. Rapamycin significantly increased cholesterol levels in the blood, whereas rosiglitazone mitigated rapamycin-induced hyperlipidemia. These results indicate that the effects of concomitant use of rosiglitazone and rapamycin in inhibiting the proliferation of WT9-12 cells and delaying the progression of ADPKD in Han:SPRD rats are stronger than those of either drug alone. The present study may provide a new strategy for the long-term treatment of ADPKD.

High-content screening assay-based discovery of paullones as novel podocyte-protective agents.

Podocyte dysfunction and loss is an early event and a hallmark of proteinuric kidney diseases. A podocyte's normal function is maintained via its unique cellular architecture that relies on an intracellular network of filaments, including filamentous actin (F-actin) and microtubules, that provides mechanical support. Damage to this filamentous network leads to changes in cellular morphology and results in podocyte injury, dysfunction, and death. Conversely, stabilization of this network protects podocytes and ameliorates proteinuria. This suggests that stabilization of podocyte architecture via its filamentous network could be a key therapeutic strategy for proteinuric kidney diseases. However, development of podocyte-directed therapeutics, especially those that target the cell's filamentous network, is still lacking, partly because of unavailability of appropriate cellular assays for use in a drug discovery environment. Here, we describe a new high-content screening-based methodology and its implementation on podocytes to identify paullone derivatives as a novel group of podocyte-protective compounds. We find that three compounds, i.e., kenpaullone, 1-azakenpaullone, and alsterpaullone, dose dependently protect podocytes from puromycin aminonucleoside (PAN)-mediated injury in vitro by reducing PAN-induced changes in both the filamentous actin and microtubules, with alsterpaullone providing maximal protection. Mechanistic studies further show that alsterpaullone suppressed PAN-induced activation of signaling downstream of GSK3ß and p38 mitogen-activated protein kinase. In vivo it reduced ADR-induced glomerular injury in a zebrafish model. Together, these results identify paullone derivatives as novel podocyte-protective agents for future therapeutic development.

Triptolide prevents extracellular matrix accumulation in experimental diabetic kidney disease by targeting microRNA-137/Notch1 pathway.

MicroRNAs (miRNAs) are involved in multiple biological functions via suppressing target genes. Triptolide is a monomeric compound isolated from a traditional Chinese herb, which exerts protective roles in many kinds of glomerular diseases. However, our understanding of the triptolide effect on miRNAome is still limited. In this study, we found that triptolide significantly decreased albuminuria and improved glomerulosclerosis in rats with diabetic kidney disease (DKD). And triptolide also inhibited extracellular matrix (ECM) protein accumulation and the notch1 pathway activation under diabetic conditions. MiR-137 was significantly decreased in the HG (high glucose)-treated HRMCs and in the kidney tissues of the diabetic rats, but was upregulated by triptolide. In addition, overexpression of miR-137 exerted similar effects to those of triptolide, while miR-137 inhibition aggravated ECM protein accumulation. Luciferase reporter assay results demonstrated that miR-137 directly targets Notch1. Furthermore, the miR-137-dependent effects were due to Notch1 suppression that in turn inhibited ECM protein expression, key mediators of glomerulosclerosis. Finally, downregulation of miR-137 reversed the ECM inhibition role of triptolide in HG cultured HRMCs. Taken together, these findings indicate that triptolide is a potential therapeutic option for DKD and that miR-137/Notch1 pathway play roles in the anti-glomerulosclerosis mechanism of triptolide.

Effects of the Vitamin D Analog 2AMD in Cyclosporine-Induced Nephrotoxicity: Dose-Response and Antifibrotic Activity.

OBJECTIVES: In this study, we aimed to ascertain the efficacy and determine the dose effects of a new analog of vitamin D, 2α-methyl-19-nor-(20S)-1α,25-dihydroxyvitamin D3 (2AMD), in decreasing fibrosis and improving renal function in a rat model of kidney disease. MATERIALS AND METHODS: Using the cyclosporine model of chronic kidney disease, we tested 4 dose regimens (2.5, 5, 10, and 20 ng/kg) of 2AMD by subcutaneous administration. The 2AMD analog was compared with another analog, 2-methylene-19-nor-(20S)-1α,25-dihydroxyvitamin D3 (2MD), given at 5 ng/kg. RESULTS: After 28 days of cyclosporine administration with 5 ng/kg 2AMD or 2MD, blood urea nitrogen levels were decreased by 20% and 30%, with no increase in serum calcium. This dose significantly decreased collagen levels by 50%, as determined by relative measurements of birefringence elicited under polarized light following picrosirius red staining of kidney tissues. The 20 ng/kg dose of 2AMD was hypercalcemic, with consequent deleterious effects on measured parameters; however, all doses of 2AMD tested decreased collagen as determined by picrosirius staining. In Western blot analysis of extracts from rat kidneys treated with cyclosporine and 5 ng/kg 2AMD, the fibrotic markers, fibronectin and vimentin, were decreased compared with animals treated only with cyclosporine. CONCLUSIONS: We found that both vitamin D analogs are potent inhibitors of kidney fibrosis with potential renoprotective activity.

Long-term treatment with chaethomellic acid A reduces glomerulosclerosis and arteriolosclerosis in a rat model of chronic kidney disease.

The high prevalence of end-stage renal disease emphasizes the failure to provide therapies to effectively prevent and/or reverse renal fibrosis. Therefore, the aim of this study was to evaluate the effect of long-term treatment with chaethomellic acid A (CAA), which selectively blocks Ha-Ras farnesylation, on renal mass reduction-induced renal fibrosis. Male Wistar rats were sham-operated (SO) or subjected to 5/6 renal mass reduction (RMR). One week after surgery, rats were placed in four experimental groups: SO:SO rats without treatment (n=13); SO+CAA: SO rats treated with CAA (n=13); RMR:RMR rats without treatment (n=14); and RMR+CAA:RMR rats treated with CAA (n=13). CAA was intraperitoneally administered in a dose of 0.23µg/kg three times a week for six months. Renal fibrosis was evaluated by two-dimensional ultrasonography and histopathological analysis. The kidneys of the RMR animals treated with CAA showed a significantly decrease in the medullary echogenicity (p<0.05) compared with the RMR rats that received no treatment. Glomerulosclerosis and arteriolosclerosis scores were significantly lower (p<0.001) in the RMR+CAA group when compared with the RMR group. There were no significant differences in interstitial fibrosis, interstitial inflammation and tubular dilatation scores between the RMR+CAA and RMR groups. These data suggest that CAA can be a potential future drug to attenuate the progression of chronic kidney disease.

Pirfenidone suppresses MAPK signalling pathway to reverse epithelial-mesenchymal transition and renal fibrosis.

AIM: Recent studies indicate that pirfenidone (PFD) may have anti-fibrotic effects in many tissues, but the potential molecular mechanism remains unknown. The purpose of this study is to investigate the potential effects of PFD on epithelial-to-mesenchymal transition (EMT) and renal fibrosis in a unilateral ureteral obstruction (UUO) rat model and the involved molecular mechanism related to cultured human renal proximal tubular epithelial cells (HK-2). METHODS: Sixty rats were randomly divided into three groups: sham-operated, vehicle-treated UUO, and PFD-treated UUO. Kidney specimens were collected at day 7 or 14 after UUO. PFD treatment was also performed for human HK-2. The tubulointerstitial injury, interstitial collagen deposition, and expression of type I and III collagen, α-SMA, S100A4, fibronection and E-cadherin were assessed. In addition, extracellular signal regulated kinase (ERK1/2), p38 MAPK (p38), and c-Jun N-terminal kinase/stress-activated protein kinase (JNK) were also detected. RESULTS: In vitro, PFD significantly attenuated TGF-ß1-induced EMT and extracellular matrix (ECM) synthesis, as determined by reducing expression of α-SMA, type I and III collagen, S100A4, fibronection, and increased expression of E-cadherin. PFD treatment attenuated TGF-ß1-induced up-regulation of phosphorylation of ERK1/2, p38 and JNK. In vivo, PFD reduced the degree of tubulointerstitial injury and renal fibrosis, which was associated with reduced expression of TGF-ß1, type III collagen, α-SMA, S100A4, fibronection, and increased expression of E-cadherin. CONCLUSION: These results suggest that pirfenidone is able to attenuate EMT and fibrosis in vivo and in vitro through antagonizing the MAPK pathway, providing a potential treatment to alleviate renal tubulointerstitial fibrosis.

Cenderitide: structural requirements for the creation of a novel dual particulate guanylyl cyclase receptor agonist with renal-enhancing in vivo and ex vivo actions.

AIMS: Cenderitide is a novel dual natriuretic peptide (NP) receptor chimeric peptide activator, which targets the particulate guanylyl cyclase B (pGC-B) receptor and pGC-A unlike native NPs. Cenderitide was engineered to retain the anti-fibrotic properties of C-type natriuretic peptide (CNP)/pGC-B with renal-enhancing actions facilitated by fusion to the carboxyl terminus of Dendroaspis NP (DNP), a pGC-A agonist, to CNP. Here, we address significance of the DNP carboxyl terminus in dual pGC receptor activation and actions of cenderitide compared with CNP on renal function and cyclic guanosine monophosphate (cGMP) in vivo and ex vivo in normal canines. METHODS AND RESULTS: In vitro, only cenderitide and not CNP or three CNP-based variants was a potent dual pGC-A/pGC-B activator of cGMP production (from 5 to 237 pmol/mL) in human embryonic kidney (HEK) 293 cells overexpressing human pGC-A while in pGC-B overexpressing cells cenderitide increased cGMP production (from 4 to 321 pmol/mL) while the three CNP-based variants were weak agonists. Based upon our finding that the DNP carboxyl terminus is a key structural requirement for dual pGC-A/pGC-B activation, we defined in vivo the renal-enhancing actions of cenderitide compared with CNP. Cenderitide increased urinary cGMP excretion (from 989 to 5977 pmol/mL), net generation of renal cGMP (821-4124 pmol/min), natriuresis (12-242 µEq/min), and glomerular filtration rate (GFR) (37-51 mL/min) while CNP did not. We then demonstrated the transformation of CNP ex vivo into a renal cGMP-activating peptide which increased cGMP in freshly isolated glomeruli eight-fold greater than CNP. CONCLUSION: The current study establishes that dual pGC-A and pGC-B activation with CNP requires the specific carboxyl terminus of DNP. In normal canines in vivo and in glomeruli ex vivo, the carboxyl terminus of DNP transforms CNP into a natriuretic and GFR-enhancing peptide. Future studies of cenderitide are warranted in cardiorenal disease states to explore its efficacy in overall cardiorenal homeostasis.

The next generation of therapeutics for chronic kidney disease.

Chronic kidney disease (CKD) represents a leading cause of death in the United States. There is no cure for this disease, with current treatment strategies relying on blood pressure control through blockade of the renin-angiotensin system. Such approaches only delay the development of end-stage kidney disease and can be associated with serious side effects. Recent identification of several novel mechanisms contributing to CKD development - including vascular changes, loss of podocytes and renal epithelial cells, matrix deposition, inflammation and metabolic dysregulation - has revealed new potential therapeutic approaches for CKD. This Review assesses emerging strategies and agents for CKD treatment, highlighting the associated challenges in their clinical development.

Targeted γ-Secretase Inhibition To Control the Notch Pathway in Renal Diseases.

Notch is a membrane inserted protein activated by the membrane-inserted γ-secretase proteolytic complex. The Notch pathway is a potential therapeutic target for the treatment of renal diseases but also controls the function of other cells, requiring cell-targeting of Notch antagonists. Toward selective targeting, we have developed the γ-secretase inhibitor-based prodrugs 13a and 15a as substrates for γ-glutamyltranspeptidase (γ-GT) and/or γ-glutamylcyclotransferase (γ-GCT) as well as aminopeptidase A (APA), which are overexpressed in renal diseases, and have evaluated them in experimental in vitro and in vivo models. In nondiseased mice, the cleavage product from Ac-γ-Glu-γ-secretase inhibitor prodrug 13a (γ-GT-targeting and γ-GCT-targeting) but not from Ac-α-Glu-γ-secretase inhibitor prodrug 15a (APA-targeting) accumulated in kidneys when compared to blood and liver. Potential nephroprotective effects of the γ-secretase inhibitor targeted prodrugs were investigated in vivo in a mouse model of acute kidney injury, demonstrating that the expression of Notch1 and cleaved Notch1 could be selectively down-regulated upon treatment with the Ac-γ-Glu-γ-secretase-inhibitor 13a.

Gum acacia mitigates genetic damage in adenine-induced chronic renal failure in rats.

BACKGROUND: Subjects with chronic renal failure (CRF) exhibit oxidative genome damage, which may predispose to carcinogenesis, and Gum acacia (GumA) ameliorates this condition in humans and animals. We evaluated here renal DNA damage and urinary excretion of four nucleic acid oxidation adducts namely 8-oxoguanine (8-oxoGua), 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), 8-oxoguanosine (8-oxoGuo) and 8-hydroxy-2-deoxyguanisone (8-OHdg) in rats with adenine (ADE)-induced CRF with and without GumA treatment. MATERIALS AND METHODS: Twenty-four rats were divided into four equal groups and treated for 4 weeks. The first group was given normal food and water (control). The second group was given normal food and GumA (15% w/v) in drinking water. The third group was fed powder diet containing adenine (ADE) (0·75% w/w in feed). The fourth group was fed like in the third group, plus GumA in drinking water (15%, w/v). RESULTS: ADE feeding induced CRF (as measured by several physiological, biochemical and histological indices) and also caused a significant genetic damage and significant decreases in urinary 8-oxo Gua and 8-oxoGuo, but not in the other nucleic acids. However, concomitant GumA treatment reduced the level of genetic damage in kidney cells as detected by Comet assay and significantly reversed the effect of adenine on urinary 8-oxoGuo. CONCLUSIONS: Treatment with GumA is able to mitigate genetic damage in renal tissues of rats with ADE-induced CRF.

Naringenin ameliorates daunorubicin induced nephrotoxicity by mitigating AT1R, ERK1/2-NFκB p65 mediated inflammation.

Inflammation and oxidative stress play important roles in the progression of renal damage. The natural polyphenol naringenin is known to exert potent antioxidant and anti-inflammatory effects. In this study, we have investigated the effect of naringenin on kidney dysfunction, fibrosis, endoplasmic reticulum (ER) stress, angiotensin II type I receptor (AT1R) expression and inflammation in daunorubicin (DNR) induced nephrotoxicity model. Nephrotoxicity was induced in rats by intravenous injection of DNR at a cumulative dose of 9 mg/kg. After 1 week, naringenin (20mg/kg/day. p.o) was administered daily for 6 weeks. Biochemical studies were performed to evaluate renal function. Western blotting was performed to measure the protein levels of AT1R, endothelin (ET)1, ET receptor type A (ETAR), extracellular signal-regulated kinase (ERK)1/2, nuclear factor (NF)κB p65, peroxisome proliferator activated receptor (PPAR)γ, oxidative/ER stress, apoptosis, and inflammatory markers in the kidney of DNR treated rats. Histopathological analysis was done using hemotoxylin eosin and Masson trichrome stained renal sections to investigate the structural abnormalities and fibrosis. DNR treated rats suffered from nephrotoxicity as evidenced by worsened renal function, increased blood urea nitrogen, serum creatinine levels in renal tissues and histopathogical abnormalities. Treatment with naringenin mitigated these changes. Furthermore, naringenin up regulated PPARγ and down regulated AT1R, ET1, ETAR, p-ERK1/2, p-NFκB p65, ER stress, apoptosis, and inflammatory markers. Our results suggest that naringenin has an ability to improve renal function and attenuates AT1R, ERK1/2-NFκB p65 signaling pathway in DNR induced nephrotoxicity in rats.