Hironobu Mukaiyama

Epithelial-to-mesenchymal transition in cyst lining epithelial cells in an orthologous PCK rat model of autosomal-recessive polycystic kidney disease

In polycystic kidney disease (PKD), cyst lining cells show polarity abnormalities. Recent studies have demonstrated loss of cell contact in cyst cells, suggesting induction of epithelial-to-mesenchymal transition (EMT). Recently, EMT has been implicated in the pathogenesis of PKD. To explore further evidence of EMT in PKD, we examined age- and segment-specific expression of adhesion molecules and mesenchymal markers in PCK rats, an orthologous model of human autosomal-recessive PKD. Kidneys from 5 male PCK and 5 control rats each at 0 days, 1, 3, 10, and 14 wk, and 4 mo of age were serially sectioned and stained with segment-specific markers and antibodies against E-cadherin, Snail1, β-catenin, and N-cadherin. mRNAs for E-cadherin and Snail1 were quantified by real-time PCR. Vimentin, fibronectin, and α-smooth muscle actin (α-SMA) expressions were assessed as mesenchymal markers. E-cadherin expression pattern was correlated with the disease pathology in that tubule segments showing the highest expression in control had much severer cyst formation in PCK rats. In PCK rats, E-cadherin and β-catenin in cystic tubules was attenuated and localized to lateral areas of cell-cell contact, whereas nuclear expression of Snail1 increased in parallel with cyst enlargement. Some epithelial cells in large cysts derived from these segments, especially in adjacent fibrotic areas, showed positive immunoreactivity for vimentin and fibronectin. In conclusion, these findings suggest that epithelial cells in cysts acquire mesenchymal features in response to cyst enlargement and participate in progressive renal fibrosis. Our study clarified the nephron segment-specific cyst profile related to EMT in PCK rats. EMT may play a key role in polycystic kidney disease.

Renal biopsy criterion in children with asymptomatic constant isolated proteinuria

BACKGROUND The criterion of a renal biopsy in children with asymptomatic persistent isolated proteinuria is controversial. METHODS To determine an adequate renal biopsy criterion in children with asymptomatic constant isolated proteinuria, the optimal cutoff maximum urinary protein/creatinine ratio (uP/Cr) to separate minor glomerular abnormalities (MGA) and other significant glomerular changes was obtained by receiver operating characteristic analysis in 44 children with asymptomatic constant isolated proteinuria (uP/Cr ≥ 0.2 g/g) screened from 1167 patients who underwent a renal biopsy between September 2000 and April 2010. Patients were divided into two groups according to the cutoff value to verify its validity. RESULTS The optimal uP/Cr was 0.5 g/g. In Group 1 (uP/Cr <0.5 g/g, n = 15), only one patient (6.7%) showed focal segmental glomerulosclerosis (FSGS) and the other 14 patients (93.3%) had MGA. In Group 2 (uP/Cr ≥ 0.5 g/g at least once before biopsy, n = 29), 5 patients showed FSGS and 7 patients had nephritis such as IgA nephropathy (41.4%, n = 12) and the other 17 patients (58.6%) showed MGA. These findings indicated that the ratio of non-MGA/MGA was significantly higher in Group 2 than that in Group 1 (P = 0.016) and that if renal biopsies were performed with a criterion of a maximum uP/Cr ≥ 0.5 g/g (criterion for Group 2), renal biopsies could be avoided in 45.2% of patients with MGA. One patient with FSGS in Group 1 showed proteinuria with uP/Cr ≥ 0.5 g/g in the clinical course. CONCLUSIONS An adequate renal biopsy criterion in children with asymptomatic constant isolated proteinuria is uP/Cr ≥ 0.5 g/g.

Renal biopsy criterion in idiopathic nephrotic syndrome with microscopic hematuria at onset.

BackgroundThe criterion for performing a renal biopsy in children with idiopathic nephrotic syndrome (NS) showing microscopic hematuria at onset remains controversial.MethodsTo determine an adequate renal biopsy criterion in children with NS showing hematuria, the optimal cutoff for the maximum red blood cell (RBC) range in urine sediment to separate minimal change disease (MCD) from other glomerular changes was obtained by receiver operating characteristic analysis. We studied 29 children with NS showing hematuria who were screened from 1,320 patients who underwent renal biopsies between January 2001 and September 2011. Patients were divided into two groups according to the cutoff value to verify its validity.ResultsThe optimal maximum RBC range was 30–49/high-power field (HPF). In group 1 (RBC ≤29/HPF, n = 14), 3 patients showed nephritis and the other 11 patients showed MCD. In group 2 (RBC ≥30/HPF, n = 15), 1 patient showed focal segmental glomerulosclerosis, 12 showed nephritis, and the other 2 showed MCD. These findings indicated that the ratio of non-MCD/MCD was significantly higher in group 2 than in group 1 (P < 0.01).ConclusionsThe use of maximum RBC range (30–49/HPF) for a criterion of renal biopsy in patients with NS showing hematuria may be reasonable for clinical practice.

First Japanese case of Pierson syndrome with mutations in LAMB2.

Pierson syndrome (OMIM 609049) is typically characterized by congenital nephritic syndrome and peculiar ocular anomalies with microcoria. It is caused by mutations in LAMB2, which encodes laminin β2. Approximately 50 mutations of LAMB2 from approximately 40 unrelated families have been identified; however, most of them were from Western countries. Although three patients in Asia with mutations of LAMB2 have been reported, they were not typical cases. We report the first Japanese case of Pierson syndrome with proven causative LAMB2 mutations. She presented with congenital nephrotic syndrome and bilateral microcoria at birth, and developed end‐stage renal disease at 2 months of age. This is the first report of a typical case from Asia. LAMB2 analysis by direct sequencing revealed the compound heterozygous mutations c.3974_3975insA (p.N1325KfsX1331, maternal, novel) in exon 25 and c.4519C>T (p.Q1507X, paternal) in exon 27. The phenotype due to LAMB2 mutations appears to be similar between different ethnic groups.

Aberrant Smad3 phosphoisoforms in cyst-lining epithelial cells in the cpk mouse, a model of autosomal recessive polycystic kidney disease

Cystic epithelia acquire mesenchymal-like features in polycystic kidney disease (PKD). In this phenotypic alteration, it is well known that transforming growth factor (TGF)-β/Smad3 signaling is involved; however, there is emerging new data on Smad3 phosphoisoforms: Smad3 phosphorylated at linker regions (pSmad3L), COOH-terminal regions (pSmad3C), and both (pSmad3L/C). pSmad3L/C has a pathological role in colorectal cancer. Mesenchymal phenotype-specific cell responses in the TGF-β/Smad3 pathway are implicated in carcinomas. In this study, we confirmed mesenchymal features and examined Smad3 phosphoisoforms in the cpk mouse, a model of autosomal recessive PKD. Kidney sections were stained with antibodies against mesenchymal markers and domain-specific phospho-Smad3. TGF-β, pSmad3L, pSmad3C, JNK, cyclin-dependent kinase (CDK) 4, and c-Myc were evaluated by Western blotting. Cophosphorylation of pSmad3L/C was assessed by immunoprecipitation. α-Smooth muscle actin, which indicates mesenchymal features, was expressed higher in cpk mice. pSmad3L expression was increased in cpk mice and was predominantly localized in the nuclei of tubular epithelial cells in cysts; however, pSmad3C was equally expressed in both cpk and control mice. Levels of pSmad3L, JNK, CDK4, and c-Myc protein in nuclei were significantly higher in cpk mice than in controls. Immunoprecipitation showed that Smad3 was cophosphorylated (pSmad3L/C) in cpk mice. Smad3 knockout/cpk double-mutant mice revealed amelioration of cpk abnormalities. These findings suggest that upregulating c-Myc through the JNK/CDK4-dependent pSmad3L pathway may be key to the pathophysiology in cpk mice. In conclusion, a qualitative rather than a quantitative abnormality of the TGF-β/Smad3 pathway is involved in PKD and may be a target for disease-specific intervention.