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Nephronophthisis (NPHP) is the most prevalent monogenic disease leading to end-stage renal failure in childhood. RhoA activation is involved in NPHP pathogenesis. This study explored the role of the RhoA activator guanine nucleotide exchange factor (GEF)-H1 in NPHP pathogenesis. We analyzed the expression and distribution of GEF-H1 in NPHP1 knockout ( NPHP1 KO ) mice using Western blotting and immunofluorescence, followed by GEF-H1 knockdown. Immunofluorescence and renal histology were used to examine the cysts, inflammation, and fibrosis. A RhoA GTPase activation assay and Western blotting were used to detect the expression of downstream GTP-RhoA and p-MLC2, respectively. In NPHP1 knockdown ( NPHP1 KD ) human kidney proximal tubular cells (HK2 cells), we detected the expressions of E-cadherin and α-smooth muscle actin (α-SMA). In vivo, increased expression and redistribution of GEF-H1, and higher levels of GTP-RhoA and p-MLC2 in renal tissue of NPHP1 KO mice were observed, together with renal cysts, fibrosis, and inflammation. These changes were alleviated by GEF-H1 knockdown. In vitro, the expression of GEF-H1 and activation of RhoA were also increased, with increased expression of α-SMA and decreased E-cadherin. GEF-H1 knockdown reversed these changes in NPHP1 KD HK2 cells. Thus, the GEF-H1/RhoA/MLC2 axis is activated in NPHP1 defects and may play a pivotal role in NPHP pathogenesis.

Nephronophthisis (NPH) is the most prevalent monogenetic disorder leading to end-stage renal failure (ESRD) in childhood. Mutations in Nphp1, encoding a cilia-localized protein, account for the majority of NPH cases. Despite its identification many years ago, Nphp1 deletions targeting exon 4 or exon 20 have not reproduced the histological features of human NPH in murine models. In this study, we deleted exon 2-20 of Nphp1 by CRISPR/Cas9 gene editing to create a near-total knockout (KO) mouse model (Nphp1del2-20/del2-20). Nphp1del2-20/del2-20 mice faithfully reproduced the renal and extrarenal phenotypes associated with human NPH, including renal cyst development, tubular basement membrane thickening, retinal degeneration and abnormal spermatogenesis. Importantly, Nphp1 re-expression using an adenoviral-associated-virus-9 vector could partially rescue both renal and retinal phenotypes in Nphp1del2-20/del2-20 mice. Our results reported the first relevant Nphp1 mouse model with renal phenotypes for human disease. It will be a valuable model for future studies of Nphp1 function and to develop novel treatments for this common childhood disease., (© The Author(s) 2021. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background: Nephronophthisis (NPHP) is a kind of ciliopathy. Interstitial fibrosis occurs at the early stage of the disease. TGF-β/Smad is a key signaling pathway in regulating interstitial fibrosis and epithelial-mesenchymal transition (EMT). In this study, we explored the activation of the TGF-β/Smad signaling pathway and EMT in NPHP1-defective MDCK cells to further understand the pathogenesis of NPHP., Methods: NPHP1-knockdown (NPHP1 KD ) MDCK cells were constructed by recombinant lentiviral short hairpin RNA, and NPHP1-knockout (NPHP1 KO ) MDCK cells were constructed by using the CRISPR/Cas9 technique. The morphology and migration ability were observed under a microscope. Western blotting was used to detect the expression of E-cadherin, β-catenin, α-smooth muscle actin (α-SMA), fibroblast-specific protein-1(FSP1), TGF-β1, Smad2, Smad3, p-Smad3, Smad4 and Smad7. The localization of Smad3 was determined by immunofluorescence assay., Results: NPHP1 KD and NPHP1 KO MDCK cells were spindle-shaped and presented EMT-like changes. E-cadherin and β-catenin expression decreased, while α-SMA and FSP1 expression increased; the TGF-β/Smad signaling pathway was activated, Smad2, Smad3, p-Smad3 and Smad4 expression increased, Smad3 translocated to nuclear and Smad7 expression decreased compared with those in wild type MDCK cells. Overexpression of Smad7 reversed these changes to different degrees., Conclusions: Our results indicate that NPHP1 defects induce the activation of the TGF-β/Smad signaling pathway and EMT in MDCK cells. These factors may be implicated in the pathogenesis of interstitial fibrosis in NPHP., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Background: Nephronophthisis (NPHP) is the most common genetic cause of end-stage renal disease (ESRD) in children. This study was performed to explore the pathogenic gene mutations and clinical and pathological features of Chinese patients with NPHP., Methods: Patients for whom causative mutations were not identified in our previous study, as well as those recruited later, were subjected to whole-exome next-generation sequencing (NGS) or the exome of 63 primary cilia disease genes., Results: We recruited 55 patients (27 boys and 28 girls) from 48 families, mainly from South China. We subjected 35 patients to NGS. Disease-causing mutations were revealed in seven more families (nine patients) by NGS. In total, disease-causing mutations were identified in 25 patients from 19 families, accounting for 39.6% (19/48) of all families, and novel mutation rate was 77.8% (35/45). NPHP1 and NPHP3 mutations were identified in 14.6% (7/48) and 12.5% (6/48) of all families, respectively. The patient with CEP83 mutations presented with prominent glomerular cysts and glomeruli dysplasia without extrarenal involvement., Conclusion: A high novel mutation rate was identified, and disease-causing mutations of NPHP3 prevailed in this group of Chinese NPHP patients. This is the second report of a patient with CEP83 mutations., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Aim: The present study was designed to explore mutations of NPHP2 and NPHP3 and clinical features in 18 Chinese infantile nephronophthisis (NPHP) patients., Methods: Patients were subjected to screen for mutations in both NPHP2 and NPHP3, and clinical data were collected., Results: Eighteen patients from 17 families were included in this study. Eight of 17 (47.1%) patients detected were identified to have mutations in NPHP3, but none had a mutation in NPHP2. Of the patients with NPHP3 mutations, four had compound heterozygous mutations, and the other four harboured single heterozygous mutations. Ten of the NPHP3 mutations were novel. Low molecular weight proteinuria was observed in all 16 detected patients. Renal histology were available in seven children, five patients showed infantile type NPHP features, and the other two patients from the same family showed juvenile type NPHP features. Liver involvement was observed in all patients with NPHP3 mutations and congenital heart disease in two patients harbouring NPHP3 mutation of c.2369 T > C (p.L790P)., Conclusions: In this group of infantile NPHP patients, mutations of NPHP3 were prevalent, whereas mutation of NPHP2 was absent. Genotype to phenotype correlations were observed in patients with NPHP3 mutations and all patients with NPHP3 mutations showed renal-hepatic phenotype., (© 2015 Asian Pacific Society of Nephrology.)