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Background: Lipid-metabolizing enzymes and their metabolites affect inflammation and fibrosis, but their roles in chronic kidney disease (CKD) have not been completely understood. Methods: To clarify their role in CKD, we measured the mRNA levels of major lipid-metabolizing enzymes in 5/6 nephrectomized (Nx) kidneys of C57BL/6 J mice. Mediator lipidomics was performed to reveal lipid profiles of CKD kidneys. Results: In 5/6 Nx kidneys, both mRNA and protein levels of Alox15 were higher when compared with those in sham kidneys. With respect to in situ hybridization, the mRNA level of Alox15 was higher in renal tubules of 5/6 Nx kidneys. To examine the role of Alox15 in CKD pathogenesis, we performed 5/6 Nx on Alox15−/− mice. Alox15−/− CKD mice exhibited better renal functions than wild-type mice. Interstitial fibrosis was also inhibited in Alox15−/− CKD mice. Mediator lipidomics revealed that Alox15−/− CKD mouse kidneys had significantly higher levels of PGD2 than the control. To investigate the effects of PGD2 on renal fibrosis, we administered PGD2 to TGF-β1-stimulated NRK-52E cells and HK-2 cells, which lead to a dose-dependent suppression of type I collagen and αSMA in both cell lines. Conclusion: Increased PGD2 in Alox15−/− CKD mouse kidneys could inhibit fibrosis, thereby resulting in CKD improvement. Thus, Alox15 inhibition and PGD2 administration may be novel therapeutic targets for CKD. [ABSTRACT FROM AUTHOR]

Background: Pseudohypoaldosteronism type II (PHAII) is a hereditary hypertensive disease caused by mutations in four genes: WNK1, WNK4, Kelch-like3 (KLHL3), and cullin3 (CUL3). Recently, it was revealed that CUL3-KLHL3 E3 ligase complex ubiquitinates WNK1 and WNK4, leading to their degradation, and that a common pathogenesis of PHAII is defective WNK degradation due to CUL3-KLHL3 E3 ligase complex impairment. PHAII-causing CUL3 mutations mediate exon9 skipping, producing a CUL3 protein with a 57-amino acid deletion (Δ403-459). However, the pathogenic effects of KLHL3, an adaptor protein that links WNKs with CUL3, in PHAII caused by CUL3 mutation remain unclear.Methods: To clarify detailed pathophysiological mechanisms underlying PHAII caused by CUL3 mutation in vivo, we generated and analyzed knock-in mice carrying the same CUL3 exon9 deletion (CUL3WT/Δex9) as that reported in PHAII patients.Results: CUL3WT/Δex9 mice exhibited a PHAII-like phenotype. Interestingly, we confirmed markedly decreased KLHL3 expression in CUL3WT/Δex9 mice by confirming the true KLHL3 band in vivo. However, the expression of other KLHL family proteins, such as KLHL2, was comparable between WT and mutant mice.Conclusion: KLHL3 expression was decreased in CUL3WT/Δex9 mice. However, expression levels of other KLHL family proteins were comparable between the wild-type and mutant mice. These findings indicate that the decreased abundance of KLHL3 is a specific phenomenon caused by mutant CUL3 (Δexon9). Our findings would improve our understanding of the pathogenesis of PHAII caused by CUL3 mutation in vivo. [ABSTRACT FROM AUTHOR]

Background: Elevated white blood cell (WBC) count is a well-known predictor of chronic kidney disease (CKD) progression. However, elderly patients commonly fail to develop a high WBC count in response to several diseased states and may instead present a low WBC count. Therefore, we hypothesized that low WBC count, in addition to high WBC count, is associated with CKD progression in the elderly.Methods: We conducted a prospective cohort study using 3-year follow-up data from the CKD Research of Outcomes in Treatment and Epidemiology study. In the present study, participants aged over 60 years with pre-dialysis CKD stages G2-G5 were eligible. Patients were stratified into three groups according to WBC count using tertiles (T). The primary outcome was a composite of end-stage renal disease and a 50% reduction in estimated glomerular filtration rate. Data were analyzed using Cox proportional hazard models with adjustments for covariates.Results: We enrolled 697 patients (males, 69%). The median WBC count was 6100 cells/µl (T1, <5400, n =  222; T2, 5400-6900, n =  235; T3, ≥6900, n = 240). During a median follow-up of 868 days, the primary outcome was observed in 170 patients, whereas 54 patients died. T1 and T3 had significantly higher hazard ratios (HR) than T2 (T1, HR 1.69, 95% confidence interval 1.14-2.51; T3, HR 1.63, 95% confidence interval 1.10-2.41). Moreover, T1 had a significantly higher adjusted HR (1.54, 95% confidence interval 1.00-2.37).Conclusion: Low WBC count is independently associated with CKD progression in the elderly. [ABSTRACT FROM AUTHOR]

Background: Vitamin D analogs have generally been recommended for treatment of mineral bone disease in chronic kidney disease (CKD). However, the association between this treatment and CKD progression has not yet been established. Methods: We designed a post hoc propensity score-matched cohort analysis derived from 3-year follow-up data of a prospective cohort. Adult participants with pre-dialysis CKD stages 4-5 who had newly been prescribed active vitamin D analogs during the observation period were eligible as matched cases. Then, matched controls were extracted from participants who had never been prescribed active vitamin D analogs. The primary outcome was a composite of end-stage renal disease or a 50 % reduction in estimated glomerular filtration rate (eGFR). A Cox proportional hazards model evaluated the association between the use of vitamin D analogs and the primary outcome. Results: We enrolled 240 patients (males, 65 %). The number of matched cases and controls was 30 and 210, respectively. The primary outcome was observed in 94 patients, whereas 25 patients died. The mean ± standard deviation age and eGFR were 69 ± 12 years and 17 ± 5.7 ml/min/1.73 m, respectively. In a Cox proportional hazard model, the use of vitamin D analogs was independently associated with a lower risk of the primary outcome (crude hazard ratio 0.41; 95 % confidence interval 0.19, 0.89; adjusted hazard ratio 0.38; 95 % confidence interval 0.17, 0.88). Conclusion: The use of vitamin D analogs is independently associated with the preservation of renal function in patients with pre-dialysis CKD stages 4-5. [ABSTRACT FROM AUTHOR]

Background: The relationship between protein-energy wasting and chronic kidney disease (CKD) progression is unknown. In the present prospective cohort study, we evaluated the hypothesis that a combination of low body mass index (BMI) and serum albumin level is associated with rapid CKD progression. Methods: The study cohort comprised 728 predialysis Japanese patients with CKD (stages 2-5) enrolled from 2010 to 2011. Patients were categorized into four groups according to their serum albumin levels and BMI: group 1, low serum albumin level (<4 g/dL) and low BMI (<23.5 kg/m); group 2, high serum albumin level (≥4 g/dL) and low BMI; group 3, low serum albumin level and high BMI (≥23.5 kg/m); and group 4, high serum albumin level and high BMI. The primary outcome was a 30 % decline in estimated glomerular filtration rate (eGFR) or start of dialysis within 2 years. The secondary outcome was an annual GFR decline (mL/min/1.73 m/year). Results: Logistic regression analysis adjusted for baseline characteristics (reference, group 4) showed that only group 1 was associated with a significant risk of CKD progression, with adjusted odds ratio of 3.51 [95 % confidence interval (CI) (1.63, 7.56)]. A multivariate linear regression analysis adjusted for baseline characteristics showed a significant difference in annual eGFR decline between groups 1 and 4 [coefficients β (standard error) −2.62 (0.75), p = 0.001]. Conclusion: This study suggests that combined effects of low BMI (<23.5 kg/m) and serum albumin level (<4 g/dL) are associated with CKD progression. [ABSTRACT FROM AUTHOR]

Background: Electrolyte abnormalities, particularly dysnatremia, are independent predictors of adverse outcome in individuals with and without renal failure. However, the association of serum chloride level (Cl) with mortality or risk of cardiovascular (CV) events in chronic kidney disease (CKD) remains unclear. Methods: This prospective cohort study included 923 pre-dialysis CKD G2-G5 patients among the participants of the CKD Research of Outcomes in Treatment and Epidemiology (CKD-ROUTE) study, who newly visited 16 nephrology centers. The primary outcome was a composite of overall death and CV events, and the secondary outcome was overall death. Data were analyzed using the Cox hazards model with adjustment for potential confounders. Results: Median Cl was 106.0 mEq/L at enrollment [quartile (Q) 1: ≤103.9, n = 207; Q2: 104.0-105.9, n = 207; Q3: 106.0-108.0, n = 289; Q4: ≥108.1, n = 220]. During a median follow-up of 33 months, there were 98 CV events, 66 deaths, and 154 composite outcomes. The hazard ratio (HR) for the composite outcome was higher for Q1 than Q3 [HR 1.72; 95 % confidence interval (CI) 1.08-2.72; P = 0.022]. As a continuous variable in a subset of patients whose Cl was ≤106.0 mEq/L, higher Cl was associated with lower risk of the composite outcome (HR 0.93; 95 % CI 0.87-0.99; P = 0.023). HR for all-cause mortality was also higher for Q1 than Q3 (HR 2.48; 95 % CI 1.22-5.03; P = 0.012). Conclusion: Low Cl was associated with increased mortality and risk of CV events in pre-dialysis CKD patients. Cl may be an additional prognostic indicator in CKD. [ABSTRACT FROM AUTHOR]

Background: Gene identification of hereditary kidney diseases by DNA sequencing is important for precise diagnosis, treatment, and genetic consultations. However, the conventional Sanger sequencing is now practically powerless in the face of ever increasing numbers of reported causative genes of various hereditary diseases. The advent of next-generation sequencing technology has enabled large-scale, genome-wide, simultaneous sequence analyses of multiple candidate genes. Methods: We designed and verified a comprehensive diagnosis panel for approximately 100 major inherited kidney diseases, including 127 known genes. The panel was named Simple, sPEedy and Efficient Diagnosis of Inherited KIdney Diseases (SPEEDI-KID). We applied the panel to 73 individuals, clinically diagnosed with an inherited kidney disease, from 56 families. Results: The panel efficiently covered the candidate genes and allowed a prompt and accurate genetic diagnosis. Moreover, 18 unreported mutations suspected as the disease causes were detected. All these mutations were validated by Sanger sequencing, with 100 % concordance. Conclusion: In conclusion, we developed a powerful diagnostic method, focusing on inherited kidney diseases, using a custom panel, SPEEDI-KID, allowing a fast, easy, and comprehensive diagnosis regardless of the disease type. [ABSTRACT FROM AUTHOR]

Background: We found that a mechanism of hypertension in pseudohypoaldosteronism type II (PHAII) caused by a WNK4 missense mutation (D561A) was activation of the WNK-OSR1/SPAK-NCC signal cascade. However, the pathogenic effect of intronic deletions in WNK1 genes also observed in PHAII patients remains unclear. To understand the pathophysiological roles of WNK1 in vivo, WNK1 mice have been analyzed, because homozygous WNK1 knockout is embryonic lethal. Although WNK1 mice have been reported to have hypotension, detailed analyses of the WNK signal cascade in the kidney and other organs of WNK1 mice have not been performed. Method: We assess the effect of heterozygous deletion of WNK1 on the WNK-OSR1/SPAK-NCC/NKCC1/NKCC2 signal cascade in the kidney and blood vessels. Results: Contrary to the previous report, the blood pressure of WNK1 mice was not decreased, even under a low-salt diet. Under a WNK4 background, the heterozygous deletion of the WNK1 gene did not reduce the high blood pressure either. We then evaluated the phosphorylation status of OSR1, SPAK, NCC, NKCC1, and NKCC2 in the kidney, but no significant decrease in the phosphorylation was observed in WNK1 mice or WNK1WNK4 mice. In contrast, a significant decrease in NKCC1 phosphorylation in the aorta and a decreased pressure-induced myogenic response in the mesenteric arteries were observed in WNK1 mice. Conclusion: The contribution of WNK1 to total WNK kinase activity in the kidney may be small, but that WNK1 may play a substantial role in the regulation of blood pressure in the arteries. [ABSTRACT FROM AUTHOR]

Background: Mutations of WNK kinase genes were identified as the cause of a hereditary hypertensive disease, pseudohypoaldosteronism type II; however, little is known about the regulation of WNK kinases. In the present study, we focused on anisosmotic conditions as the initial clues for clarifying a stimulating factor for WNK kinase activity. Methods: Endogenous WNK kinase activity in COS7 cells was monitored by the phosphorylation of its substrate, OSR1. Results: Knockdown experiments revealed that WNK1 was a major WNK kinase in COS7 cells. In contrast to the transient increase in WNK1 activity caused by hypertonic medium, hypotonic medium increased the phosphorylation of OSR1 for 24 h, suggesting that the hypotonic medium included a signal for continuously stimulating WNK1 kinase activity. To identify the signal, ion substitution experiments were performed. Surprisingly, even isotonic media with low Cl or low K was found to increase OSR1 phosphorylation as well as the hypotonic medium. Furthermore, WNK1 activation by the hypotonic medium was completely blocked by quinine (500 μM) but not by 5-nitro-2-(3-phenylpropylamino) benzoic acid (100 μM), and this inhibition was closely correlated with the inhibition of Rb (=K) efflux but not with the inhibition of I (=Cl) efflux. These results suggest that K, rather than hypotonicity or low Cl, may be an important regulator for WNK1 activation. Finally, we confirmed that high K and low K media under the physiological range decreased and increased WNK1 activity, respectively. Conclusion: Extracellular K is an important regulator of WNK1 kinase activity. [ABSTRACT FROM AUTHOR]