Circulating Extracellular Vesicle-Propagated microRNA Signature as a Vascular Calcification Factor in Chronic Kidney Disease.

BACKGROUND: Chronic kidney disease (CKD) accelerates vascular calcification via phenotypic switching of vascular smooth muscle cells (VSMCs). We investigated the roles of circulating small extracellular vesicles (sEVs) between the kidneys and VSMCs and uncovered relevant sEV-propagated microRNAs (miRNAs) and their biological signaling pathways. METHODS AND RESULTS: We established CKD models in rats and mice by adenine-induced tubulointerstitial fibrosis. Cultures of A10 embryonic rat VSMCs showed increased calcification and transcription of osterix (Sp7), osteocalcin (Bglap), and osteopontin (Spp1) when treated with rat CKD serum. sEVs, but not sEV-depleted serum, accelerated calcification in VSMCs. Intraperitoneal administration of a neutral sphingomyelinase and biogenesis/release inhibitor of sEVs, GW4869 (2.5 mg/kg per 2 days), inhibited thoracic aortic calcification in CKD mice under a high-phosphorus diet. GW4869 induced a nearly full recovery of calcification and transcription of osteogenic marker genes. In CKD, the miRNA transcriptome of sEVs revealed a depletion of 4 miRNAs, miR-16-5p, miR-17~92 cluster-originated miR-17-5p/miR-20a-5p, and miR-106b-5p. Their expression decreased in sEVs from CKD patients as kidney function deteriorated. Transfection of VSMCs with each miRNA-mimic mitigated calcification. In silico analyses revealed VEGFA (vascular endothelial growth factor A) as a convergent target of these miRNAs. We found a 16-fold increase in VEGFA transcription in the thoracic aorta of CKD mice under a high-phosphorus diet, which GW4869 reversed. Inhibition of VEGFA-VEGFR2 signaling with sorafenib, fruquintinib, sunitinib, or VEGFR2-targeted siRNA mitigated calcification in VSMCs. Orally administered fruquintinib (2.5 mg/kg per day) for 4 weeks suppressed the transcription of osteogenic marker genes in the mouse aorta. The area under the curve of miR-16-5p, miR-17-5p, 20a-5p, and miR-106b-5p for the prediction of abdominal aortic calcification was 0.7630, 0.7704, 0.7407, and 0.7704, respectively. CONCLUSIONS: The miRNA transcriptomic signature of circulating sEVs uncovered their pathologic role, devoid of the calcification-protective miRNAs that target VEGFA signaling in CKD-driven vascular calcification. These sEV-propagated miRNAs are potential biomarkers and therapeutic targets for vascular calcification.

LRBA is essential for urinary concentration and body water homeostasis.

Protein kinase A (PKA) directly phosphorylates aquaporin-2 (AQP2) water channels in renal collecting ducts to reabsorb water from urine for the maintenance of systemic water homeostasis. More than 50 functionally distinct PKA-anchoring proteins (AKAPs) respectively create compartmentalized PKA signaling to determine the substrate specificity of PKA. Identification of an AKAP responsible for AQP2 phosphorylation is an essential step toward elucidating the molecular mechanisms of urinary concentration. PKA activation by several compounds is a novel screening strategy to uncover PKA substrates whose phosphorylation levels were nearly perfectly correlated with that of AQP2. The leading candidate in this assay proved to be an AKAP termed lipopolysaccharide-responsive and beige-like anchor protein (LRBA). We found that LRBA colocalized with AQP2 in vivo, and Lrba knockout mice displayed a polyuric phenotype with severely impaired AQP2 phosphorylation. Most of the PKA substrates other than AQP2 were adequately phosphorylated by PKA in the absence of LRBA, demonstrating that LRBA-anchored PKA preferentially phosphorylated AQP2 in renal collecting ducts. Furthermore, the LRBA-PKA interaction, rather than other AKAP-PKA interactions, was robustly dissociated by PKA activation. AKAP-PKA interaction inhibitors have attracted attention for their ability to directly phosphorylate AQP2. Therefore, the LRBA-PKA interaction is a promising drug target for the development of anti-aquaretics.

Absence of ULK1 decreases AMPK activity in the kidney, leading to chronic kidney disease progression.

AMP-activated protein kinase (AMPK) inactivation in chronic kidney disease (CKD) leads to energy status deterioration in the kidney, constituting the vicious cycle of CKD exacerbation. Unc-51-like kinase 1 (ULK1) is considered a downstream molecule of AMPK; however, it was recently reported that the activity of AMPK could be regulated by ULK1 conversely. We demonstrated that AMPK and ULK1 activities were decreased in the kidneys of CKD mice. However, whether and how ULK1 is involved in the underlying mechanism of CKD exacerbation remains unknown. In this study, we investigated the ULK1 involvement in CKD, using ULK1 knockout mice. The CKD model of Ulk1-/- mice exhibited significantly exacerbated renal function and worsening renal fibrosis. In the kidneys of the CKD model of Ulk1-/- mice, reduced AMPK and its downstream ß-oxidation could be observed, leading to an energy deficit of increased AMP/ATP ratio. In addition, AMPK signaling in the kidney was reduced in control Ulk1-/- mice with normal renal function compared to control wild-type mice, suggesting that ULK1 deficiency suppressed AMPK activity in the kidney. This study is the first to present ULK1 as a novel therapeutic target for CKD treatment, which regulates AMPK activity in the kidney.

A case of unexpected diagnosis of fibronectin glomerulopathy with histological features of membranoproliferative glomerulonephritis.

Fibronectin (FN) glomerulopathy (FNG), a rare autosomal hereditary renal disease, is characterized by proteinuria resulting from the massive accumulation of FN in the glomeruli. It typically affects individuals aged 10-50 years. In this report, we describe the case of a 57-year-old man who was diagnosed with FNG through genetic analysis and histological examination that revealed membranoproliferative glomerulonephritis. Despite treatment with prednisolone, the therapeutic response was unsatisfactory. Prednisolone was subsequently tapered and discontinued because the patient had pulmonary thromboembolism. Subsequent comprehensive genetic testing, which was initially not conducted because the patient's parents did not have a history of kidney disease, identified a known disease-causing variant in the FN1 gene, indicating a de novo variant. FNG was further confirmed by positive staining of glomeruli with FN using an IST-4 antibody. Although corticosteroid therapy is commonly employed as the initial treatment for MPGN, its appropriateness depends on the underlying etiology. Thus, clinicians must be aware of potential rare genetic causes underlying MPGN.

Associations Between Dietary Potassium Intake From Different Food Sources and Hyperkalemia in Patients With Chronic Kidney Disease.

OBJECTIVE: Previous studies reported mixed results on associations between dietary potassium intake and hyperkalemia in patients with chronic kidney disease (CKD). This study investigated the association between potassium intake from different food sources and hyperkalemia in patients with non-dialysis-dependent CKD. METHODS: A total of 285 patients were recruited at a university hospital and 2 city hospitals in Tokyo. Dietary potassium intake was estimated by a validated diet history questionnaire. Associations of potassium intake from all foods and individual food groups with serum potassium were examined by multivariable linear regression among potassium binder nonusers. An association between tertile groups of potassium intake and hyperkalemia, defined as serum potassium ≥5.0 mEq/L, was evaluated by multivariable logistic regression. RESULTS: Among 245 potassium binder nonusers, total potassium intake was weakly associated with serum potassium (regression coefficient = 0.147, 95% confidence interval (CI): 0.018-0.277), while an association with hyperkalemia was not observed (first vs third tertile: adjusted odds ratio = 0.98, 95% CI: 0.29-3.26). As for food groups, potassium intakes from potatoes, pulses, and green/yellow vegetables were positively associated with serum potassium. Patients in the highest tertile of potassium intake from potatoes had higher odds of hyperkalemia as compared to those in the lowest tertile (adjusted odds ratio = 4.12, 95% CI: 1.19-14.34). CONCLUSION: Total potassium intake was weakly associated with serum potassium, but not with hyperkalemia. Potassium intake from potatoes was associated with hyperkalemia. These findings highlight the importance of considering food sources of potassium in the management of hyperkalemia in CKD.

LRBA signalosomes activate vasopressin-induced AQP2 trafficking at recycling endosomes.

Aquaporin-2 (AQP2) water channels are proteins that are recycled between intracellular vesicles and the apical plasma membrane in renal collecting ducts. Lipopolysaccharide-responsive beige-like anchor protein (LRBA) is a protein kinase A (PKA) anchoring protein that creates compartmentalized PKA signalling responsible for AQP2 phosphorylation. In response to increased plasma osmolality, vasopressin/cyclic adenosine monophosphate (cAMP)/PKA signalling phosphorylates AQP2, promoting AQP2 trafficking into the apical plasma membrane and increasing water reabsorption from urine. However, the molecular mechanisms by which LRBA mediates vasopressin-induced AQP2 phosphorylation remain unknown. To investigate AQP2 intracellular localization and phosphorylation status in vivo, a density gradient ultracentrifugation technique was combined with an in situ proximity ligation assay, super-resolution structured illumination microscopy and immunoelectron microscopy. Most of the AQP2 was localized on the recycling endosome in the presence of tolvaptan, a vasopressin type 2 receptor (V2R) antagonist. Desmopressin, a V2R agonist, phosphorylated AQP2, translocating it from the recycling endosome to the apical plasma membrane. In contrast, LRBA was constitutively localized at the recycling endosome. Therefore, LRBA and AQP2 were well colocalized in the absence of vasopressin stimulation. The loss of LRBA/PKA signalling by Lrba knockout impaired vasopressin-induced AQP2 phosphorylation, resulting in AQP2 retention at the recycling endosome. Defective AQP2 trafficking caused low urinary concentrating ability in Lrba-/- mice. The LRBA-PKA complex created compartmentalized PKA signalling at the recycling endosome, which facilitated AQP2 phosphorylation in response to vasopressin. KEY POINTS: Membrane proteins are continuously internalized into the endosomal system via endocytosis, after which they are either recycled back to the plasma membrane or degraded at the lysosome. In T cells, lipopolysaccharide-responsive beige-like anchor protein (LRBA) binds directly to the cytotoxic T lymphocyte antigen 4 (CTLA-4), a checkpoint immune molecule, to prevent CTLA-4 lysosomal degradation and promote its vesicle recycling. LRBA has different physiological functions in renal collecting ducts. LRBA and aquaporin-2 (AQP2) water channels were colocalized on the recycling endosome in vivo in the absence of the anti-diuretic hormone vasopressin. LRBA promoted vasopressin-induced AQP2 trafficking, increasing water reabsorption from urine via AQP2. LRBA determined renal responsiveness to vasopressin at recycling endosomes. LRBA is a ubiquitously expressed anchor protein. LRBA signalosomes might regulate membrane trafficking of several constitutively recycled proteins at recycling endosomes.

Rapidly progressive IgA nephropathy with membranoproliferative glomerulonephritis-like lesions in an elderly man following the third dose of an mRNA COVID-19 vaccine: a case report.

BACKGROUND: As messenger RNA (mRNA)-based vaccines for coronavirus disease 2019 (COVID-19) have been administered to millions of individuals worldwide, cases of de novo and relapsing glomerulonephritis after mRNA COVID-19 vaccination are increasing in the literature. While most previous publications reported glomerulonephritis after the first or second dose of an mRNA vaccine, few reports of glomerulonephritis occurring after the third dose of an mRNA vaccine currently exist. CASE PRESENTATION: We report a case of rapidly progressive glomerulonephritis in a patient following the third dose of an mRNA COVID-19 vaccine. A 77-year-old Japanese man with a history of hypertension and atrial fibrillation was referred to our hospital for evaluation of anorexia, pruritus, and lower extremity edema. One year before referral, he received two mRNA vaccines (BNT162b2) for COVID-19. Three months before the visit, he received a third mRNA vaccine (mRNA-1273) for COVID-19. On admission, the patient presented severe renal failure with a serum creatinine level of 16.29 mg/dL, which had increased from 1.67 mg/dL one month earlier, prompting us to initiate hemodialysis. Urinalysis showed nephrotic-range proteinuria and hematuria. Renal biopsy revealed mild mesangial proliferation and expansion, a lobular appearance, and double contours of the glomerular basement membrane. Renal tubules had severe atrophy. Immunofluorescence microscopy showed strong mesangial staining for IgA, IgM, and C3c. Electron microscopy exhibited mesangial and subendothelial electron-dense deposits, leading to a diagnosis of IgA nephropathy with membranoproliferative glomerulonephritis-like changes. The kidney function remained unchanged after steroid therapy. CONCLUSIONS: Although the link between renal lesions and mRNA vaccines remains unclear, a robust immune response induced by mRNA vaccines may play a role in the pathogenesis of glomerulonephritis. Further studies of the immunological effects of mRNA vaccines on the kidney are warranted.

The Effect of Antihypertensive Therapy on Skeletal Muscle Mass and Bone Mineral Density in Patients With End-Stage Kidney Disease.

OBJECTIVE: Sarcopenia and osteoporosis substantially influence health and lifespan. However, the variables affecting skeletal muscle mass (SMM) or bone mineral density (BMD) remain unknown. DESIGN AND METHODS: From August 1, 2018 to July 31, 2019, we conducted a single-center, observational cohort study with 291 Japanese adult patients on maintenance hemodialysis due to end-stage kidney disease, who had their femoral neck BMD measured using dual-energy X-ray absorptiometry. After 1-year follow-up, we measured annual changes of BMD (ΔBMD) and SMM (ΔSMM), which were calculated through a modified creatinine index (mg/kg/day) using age, sex, serum creatinine, and single-pooled Kt/V for urea. The factors associated with ΔSMM/ΔBMD or progressive loss of SMM/BMD, defined as ΔSMM/ΔBMD < 0 per year, respectively, were analyzed with multivariable, linear regression or logistic regression models. RESULTS: The median age of the patients was 66 years and 33% were female. Dialysis vintage and ß-blocker-use were inversely correlated to ΔSMM. In comparison to nonusers, ß-blockers users had 2.5-fold higher SMM loss odd ratios [95% confidence interval, 1.3-4.8]. The risk for SMM loss caused by ß-blockers was not increased in users of renin-angiotensin system inhibitors. The ΔBMD was negatively correlated to the usage of calcium channel blockers. The risk of developing osteosarcopenia, which was defined as annual loss of both SMM and BMD, increased in calcium channel blockers users. CONCLUSIONS: The use of ß-blockers is associated with an elevated risk of developing sarcopenia, whereas renin-angiotensin system inhibitors may minimize this effect in patients with end-stage kidney disease. Use of calcium channel blocker therapy was associated with a faster decline of BMD.

Phenotypic differences of mutation-negative cases in Gitelman syndrome clinically diagnosed in adulthood.

Gitelman syndrome (GS), an autosomal recessive kidney disorder, is characterized by hypokalemia, hypomagnesemia, hypocalciuria, and metabolic alkalosis. Generally, diagnosis is made in school-aged children but multiple cases have been diagnosed in adulthood. This study examines the phenotypic differences between genetically confirmed cases and mutation-negative cases in adults. A comprehensive screening of 168 genes, including GS-related genes, was performed for 84 independent individuals who were referred to our institute with a clinical diagnosis of GS. The cases of pseudo-Bartter syndrome (BS)/GS because of diuretic abuse or other causes, which was determined based on patients' medical records, were excluded during registration. Of these 70 eligible cases for analysis, 27 (38.6%) had genetic confirmation of GS, while 37 (52.8%) had no known variants associated with GS and were considered to be unsolved cases. Note that unsolved cases comprised older, mostly female, individuals with decreased kidney function and multiple basic features of GS. The phenotype of unsolved cases is similar to that of pseudo BS/GS cases, although these cases were excluded in advance. However, the genetic and autoimmune profiles of these unsolved cases have not yet been investigated to date. Therefore, these cases may be categorized into new disease groups.

Deletion of Alox15 improves kidney dysfunction and inhibits fibrosis by increased PGD2 in the kidney.

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-ß1-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.

Renal TNFα activates the WNK phosphorylation cascade and contributes to salt-sensitive hypertension in chronic kidney disease.

The inappropriate over-activation of the with-no-lysine kinase (WNK)-STE20/SPS1-related proline/alanine-rich kinase (SPAK)-sodium chloride cotransporter (NCC) phosphorylation cascade increases sodium reabsorption in distal kidney nephrons, resulting in salt-sensitive hypertension. Although chronic kidney disease (CKD) is a common cause of salt-sensitive hypertension, the involvement of the WNK phosphorylation cascade is unknown. Moreover, the effect of immune systems on WNK kinases has not been investigated despite the fact that immune systems are important for salt sensitivity. Here we demonstrate that the protein abundance of WNK1, but not of WNK4, was increased at the distal convoluted tubules in the aristolochic acid nephropathy mouse model of CKD. Accordingly, the phosphorylation of both SPAK and NCC was also increased. Moreover, a high-salt diet did not adequately suppress activation of the WNK1-SPAK-NCC phosphorylation cascade in this model, leading to salt-sensitive hypertension. WNK1 also was increased in adenine nephropathy, but not in subtotal nephrectomy, models of CKD. By comparing the transcripts of these three models focusing on immune systems, we hypothesized that tumor necrosis factor (TNF)-α regulates WNK1 protein expression. In fact, TNF-α increased WNK1 protein expression in cultured renal tubular cells by reducing the transcription and protein levels of NEDD4-2 E3-ligase, which degrades WNK1 protein. Furthermore, the TNF-α inhibitor etanercept reversed the reduction of NEDD4-2 expression and upregulation of the WNK1-SPAK-NCC phosphorylation cascade in distal convoluted tubules in vivo in the aristolochic acid nephropathy model. Thus, salt-sensitive hypertension is induced in CKD via activation of the renal WNK1- SPAK-NCC phosphorylation cascade by TNF-α, reflecting a link with the immune system.

WNK1-TAK1 signaling suppresses lipopolysaccharide-induced cytokine production and classical activation in macrophages.

With-no-lysine kinase (WNK) plays important roles in regulating electrolyte homeostasis, cell signaling, survival, and proliferation. It has been recently demonstrated that WNK1, a member of the WNK family, modifies the function of immune cells. Here we report that in macrophages, WNK1 has suppressive effects on lipopolysaccharide (LPS)-induced inflammatory responses via TGFß-activated kinase 1 (TAK1)-mediated activation of nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathway. We found that WNK1 heterozygous (WNK1+/-) mice produced excessive proinflammatory cytokines in an experimental LPS-induced sepsis model, and peritoneal macrophages isolated from WNK1+/- mice produced higher levels of LPS-induced cytokines and NOS2 expression as canonical proinflammatory M1 macrophage markers. We confirmed that small hairpin RNA (shRNA)-mediated knockdown of WNK1 activated LPS-induced cytokine production and NOS2 expression in RAW 264.7 macrophages. Moreover, we demonstrated that WNK1 knockdown increased the nuclear translocation of NF-κB and activated the p38 and Jun N-terminal kinase (JNK) MAPK signaling pathway and that a TAK1 inhibitor diminished these effects of WNK1 knockdown. These results suggest that WNK1 acts as a physiologic immune modulator via interactions with TAK1. WNK1 may be a therapeutic target against the cytokine storm caused by sepsis.

Genetics May Predict Effectiveness of Tolvaptan in Autosomal Dominant Polycystic Kidney Disease.

BACKGROUND: Tolvaptan is the only therapeutic drug for autosomal dominant polycystic kidney disease (ADPKD). The influence of mutations in polycystic kidney disease 1 and 2 genes (PKD1 and PKD2) on the treatment effects of tolvaptan is not well documented in the literature. METHODS: We retrospectively evaluated the relationship between genotype and the efficacy of tolvaptan in 18 patients with ADPKD who had been treated at Toranomon Hospital and undergone genetic testing between April 2016 and February 2020. RESULTS: The annual change in estimated glomerular filtration rate (ΔeGFR/y) from before to after tolvaptan was from a median of -5.5 to -2.5 mL/min/1.73 m2 in the PKD1 truncating group, -3.3 to -2.4 mL/min/1.73 m2 in the PKD1 non-truncating group, -3.1 to -1.6 mL/min/1.73 m2 in the PKD2 group, and -1.9 to -2.6 mL/min/1.73 m2 in the group with no PKD1/2 mutation. The median degrees of improvement of ΔeGFR/y were 2.5 (45%), 0.4 (10%), 0.6 (28%), and -0.7 (-37%) mL/min/1.73 m2, respectively. Compared with the group of patients with any PKD1/2 mutation, the group with no PKD1/2 mutation showed significantly less improvement in ΔeGFR/y with tolvaptan (0.6 vs. -0.7 mL/min/1.73 m2, respectively; p = 0.01) and significantly less improvement in the annual rate of increase in total kidney volume (TKV) with tolvaptan (-6.7 vs. -1.1%, respectively; p = 0.02). CONCLUSION: Patients with ADPKD and no PKD1/2 mutation showed less improvement in ΔeGFR/y and the annual rate of increase in TKV with tolvaptan. Detecting PKD1/2 mutations may be useful for predicting the effectiveness of tolvaptan.

Failure to sense energy depletion may be a novel therapeutic target in chronic kidney disease.

The kidneys consume a large amount of energy to regulate volume status and blood pressure and to excrete uremic toxins. The identification of factors that cause energy mismatch in the setting of chronic kidney disease (CKD) and the development of interventions aimed at improving this mismatch are key research imperatives. Although the critical cellular energy sensor 5'-adenosine monophosphate (AMP)-activated protein kinase (AMPK) is known to be inactivated in CKD, the mechanism of AMPK dysregulation is unknown. In a mouse model of CKD, metabolome analysis confirmed a decrease in AMPK activation in the kidneys despite a high AMP: ATP ratio, suggesting that AMPK did not sense energy depletion. Similar AMPK inactivation was found in heart and skeletal muscle in CKD mice. Several uremic factors were shown to inactivate AMPK in vitro and in ex vivo preparations of kidney tissue. The specific AMPK activator A-769662, which bypasses the AMP sensing mechanism, ameliorated fibrosis and improved energy status in the kidneys of CKD mice, whereas an AMP analog did not. We further demonstrated that a low-protein diet activated AMPK independent of the AMP sensing mechanism, leading to improvement in energy metabolism and kidney fibrosis. These results suggest that a failure to sense AMP is the key mechanism underlying the vicious cycle of energy depletion and CKD progression and direct AMPK activation may be a novel therapeutic approach in CKD.

Genotype-Clinical Correlations in Polycystic Kidney Disease with No Apparent Family History.

BACKGROUND: Genetic characteristics of polycystic kidney disease (PKD) patients without apparent family history were reported to be different from those with a positive family history. However, the clinical course of PKD patients with no apparent family history is not well documented in the literature. METHODS: We evaluated the relationship between genotype and the clinical course of 62 PKD patients with no apparent family history. RESULTS: The annual decline of renal function was faster in the patients with PKD1/PKD2 mutation (PKD1 truncating [-3.08; 95% CI -5.30 to -0.87, p = 0.007], PKD1 nontruncating [-2.10; -3.82 to -0.38, p = 0.02], and PKD2 [-2.31; -4.40 to -0.23, p = 0.03]) than in the other patients without PKD1/PKD2 mutation. Similar results were obtained after adjustment for gender, age, estimated glomerular filtration rate (eGFR), height-adjusted total kidney volume (TKV), and mean arterial pressure (MAP). There was no significant difference in the annual decline of renal function among the different PKD1/PKD2 groups, but Kaplan-Meier analysis showed that progression to eGFR < 15 mL/min/1.73 m2 was significantly faster in PKD1 truncating group (p = 0.05). The annual rate of TKV increase was larger in the patients with PKD1/PKD2 mutation (PKD1 truncating [4.63; 95% CI 0.62-8.64, p = 0.03], PKD1 nontruncating [3.79; 0.55-7.03, p = 0.02], and PKD2 [2.11; -1.90 to 6.12, p = 0.29]) than in the other patients without PKD1/PKD2 mutation. Similar results were obtained after adjustment for gender, age, eGFR, and MAP. CONCLUSION: Detection of PKD1/PKD2 mutation, especially PKD1 truncating, is useful for predicting the renal outcome and rate of TKV increase in PKD patients with no apparent family history.

Renal histology and MRI findings in a 37-year-old Japanese patient with autosomal recessive polycystic kidney disease
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A 37-year-old Japanese man with a serum creatinine level of 2.5 mg/dL and hepatomegaly was admitted to our hospital for investigation of renal failure. Magnetic resonance imaging (MRI) showed hepatomegaly with small cystic lesions that had high signal intensity on T2-weighted images. There was no splenomegaly, and the kidneys were nearly normal in size with a few small cystic lesions. Renal biopsy revealed that interstitial fibrosis and tubular atrophy affected 60% of the cortex. There was cystic tubular dilation, mainly affecting the distal loop of Henle and distal tubules, since immunohistochemical staining of the dilated tubules was positive for cytokeratin 7 and Tamm-Horsfall protein but was negative for aquaporin 3 and CD10. Immunofluorescence microscopy and electron microscopy did not demonstrate any immune deposits. Genetic analysis identified two different heterozygous missense variants of PKHD1, while the patient's asymptomatic parents were each heterozygous for a single PKHD1 mutation. Accordingly, autosomal recessive polycystic kidney disease (-ARPKD) due to compound heterozygous PKHD1 mutation was diagnosed. The renal biopsy findings of this patient may be nonspecific, but they were different from the typical renal histology of infantile ARPKD. In conclusion, the renal features of adult-onset ARPKD may differ from those of infantile disease.
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Comprehensive genetic testing approach for major inherited kidney diseases, using next-generation sequencing with a custom panel.

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.

Association of serum chloride level with mortality and cardiovascular events in chronic kidney disease: the CKD-ROUTE study.

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.

Combination of low body mass index and serum albumin level is associated with chronic kidney disease progression: the chronic kidney disease-research of outcomes in treatment and epidemiology (CKD-ROUTE) study.

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/m2); group 2, high serum albumin level (≥4 g/dL) and low BMI; group 3, low serum albumin level and high BMI (≥23.5 kg/m2); 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 m2/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 ß (standard error) -2.62 (0.75), p = 0.001]. CONCLUSION: This study suggests that combined effects of low BMI (<23.5 kg/m2) and serum albumin level (<4 g/dL) are associated with CKD progression.

Responsiveness to erythropoiesis-stimulating agents and renal survival in patients with chronic kidney disease.

BACKGROUND: Renal anemia of chronic kidney disease (CKD) is generally treated by erythropoiesis-stimulating agents (ESAs). However, there are individual differences in patients' responsiveness to ESA, which may affect the prognosis of CKD. METHODS: The effect of ESAs on hemoglobin was followed in 297 CKD patients with renal anemia. Three types of ESA, epoetin alfa or beta, darbepoetin alfa, and epoetin beta pegol, were used in this study and dose of ESA was converted to that of epoetin using a dose conversion ratio (epoetin:darbepoetin alfa:epoetin beta pegol = 200:1:0.93). After initial 12-week administration of ESAs, the patients were divided into three groups: poor, intermediate, and good responders based on ΔHb/week/epoetin dose as an index. Hemoglobin values were followed for 144 weeks. RESULTS: Initial patient characteristics--including age, body mass index, hemoglobin, estimated glomerular filtration rate, transferrin saturation, ferritin, albumin, calcium, parathyroid hormone, C-reactive protein, and urine protein--were similar in the three responder groups, except phosphate in the poor responder group was significantly higher than in the other two groups. The period from ESA use to renal death (RD) was significantly shortest in the poor responder group, and the number of RD patients was fewer in the good responder group. Multivariate Cox regression revealed that low final ΔHb(ΔHb from ESA use to just before dialysis)/week/epoetin dose, and low Hb after 12-week ESA use were significant factors related to responsiveness to ESA, suggesting that hyporesponsiveness to ESA was a risk factor for RD. Cox regression also found that hyperphosphatemia and diabetic nephropathy were risks for RD as well. CONCLUSIONS: The study results suggest that hyporesponsiveness to ESA after the first 12-week administration as well as after 12 weeks is a risk for RD in pre-dialysis CKD patients. Furthermore, hyperphosphatemia and diabetic nephropathy are risk factors for RD.