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A mouse model for inherited renal fibrosis associated with endoplasmic reticulum stress.

ABSTRACT: Renal fibrosis is a common feature of renal failure resulting from multiple etiologies, including diabetic nephropathy, hypertension and inherited renal disorders. However, the mechanisms of renal fibrosis are incompletely understood and we therefore explored these by establishing a mouse model for a renal tubular disorder, referred to as autosomal dominant tubulointerstitial kidney disease (ADTKD) due to missense uromodulin (UMOD) mutations (ADTKD-UMOD). ADTKD-UMOD, which is associated with retention of mutant uromodulin in the endoplasmic reticulum (ER) of renal thick ascending limb cells, is characterized by hyperuricemia, interstitial fibrosis, inflammation and renal failure, and we used targeted homologous recombination to generate a knock-in mouse model with an ADTKD-causing missense cysteine to arginine uromodulin mutation (C125R). Heterozygous and homozygous mutant mice developed reduced uric acid excretion, renal fibrosis, immune cell infiltration and progressive renal failure, with decreased maturation and excretion of uromodulin, due to its retention in the ER. The ER stress marker 78 kDa glucose-regulated protein (GRP78) was elevated in cells expressing mutant uromodulin in heterozygous and homozygous mutant mice, and this was accompanied, both in vivo and ex vivo, by upregulation of two unfolded protein response pathways in primary thick ascending limb cells from homozygous mutant mice. However, this did not lead to an increase in apoptosis in vivo Thus, we have developed a novel mouse model for renal fibrosis, which will be a valuable resource to decipher the mechanisms linking uromodulin mutations with ER stress and renal fibrosis.

SUBMITTER: Piret SE

PROVIDER: S-EPMC5483009 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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A mouse model for inherited renal fibrosis associated with endoplasmic reticulum stress.

Piret Sian E SE Olinger Eric E Reed Anita A C AAC Nesbit M Andrew MA Hough Tertius A TA Bentley Liz L Devuyst Olivier O Cox Roger D RD Thakker Rajesh V RV

Disease models & mechanisms 20170321 6

Renal fibrosis is a common feature of renal failure resulting from multiple etiologies, including diabetic nephropathy, hypertension and inherited renal disorders. However, the mechanisms of renal fibrosis are incompletely understood and we therefore explored these by establishing a mouse model for a renal tubular disorder, referred to as autosomal dominant tubulointerstitial kidney disease (ADTKD) due to missense uromodulin (UMOD) mutations (ADTKD-UMOD). ADTKD-UMOD, which i ...[more]

PMID: 28325753

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Project description:Endoplasmic reticulum (ER) stress-associated apoptosis plays a role in organ remodeling after insult. The effect of ER stress on renal tubular damage and fibrosis remains controversial. This study aims to investigate whether ER stress is involved in tubular destruction and interstitial fibrosis in vivo. Renal cell apoptosis was proven by terminal deoxynucleotidyl transferase dUTP nick end-labeling (TUNEL) stain and poly-ADP ribose polymerase expression in the unilateral ureteral obstruction (UUO) kidney. ER stress was evoked and confirmed by the upregulation of glucose-regulated protein 78 (GRP78) and the common Lys-Asp-Glu-Leu (KDEL) motif of ER retention proteins after UUO. ER stress-associated proapoptotic signals, including B-cell chronic lymphocytic leukemia (CLL)/lymphoma 2-associated × protein (BAX) expression, caspase-12 and c-Jun N-terminal kinase (JNK) phosphorylation, were activated in the UUO kidney. Prolonged ER stress attenuated both unsplicing and splicing X-box binding protein 1 (XBP-1) protein expression, but continued to activate inositol-requiring 1? (IRE1?)-JNK phosphorylation, protein kinase RNA-like endoplasmic reticulum kinase (PERK), eukaryotic translation initiation factor 2? subunit (eIF2?), activating transcription factor (ATF)-4, CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP) and cleavage activating transcription factor 6 (cATF6)-CHOP signals, which induce ER stress-related apoptosis but attenuate adaptive unfolded protein responses in UUO kidneys. However, renal apoptosis and fibrosis were attenuated in candesartan-treated UUO kidney. Candesartan was associated with maintenance of XBP-1 expression and attenuated ATF4, cATF6 and CHOP protein expression. Taken together, results show that overwhelming ER stress leads to renal cell apoptosis and subsequent fibrosis; and candesartan, at least in part, restores renal integrity by blocking ER stress-related apoptosis. Reducing ER stress may present a way to attenuate renal fibrosis.

Project description:The gain-of-function minor allele of the MUC5B (mucin 5B, oligomeric mucus/gel-forming) promoter (rs35705950) is the strongest risk factor for idiopathic pulmonary fibrosis (IPF), a devastating fibrotic lung disease that leads to progressive respiratory failure in adults. We have previously demonstrated that Muc5b overexpression in mice worsens lung fibrosis after bleomycin exposure and have hypothesized that excess Muc5b promotes endoplasmic reticulum (ER) stress and apoptosis, stimulating fibrotic lung injury. Here, we report that ER stress pathway members ATF4 (activating transcription factor 4) and ATF6 coexpress with MUC5B in epithelia of the distal IPF airway and honeycomb cyst and that this is more pronounced in carriers of the gain-of-function MUC5B promoter variant. Similarly, in mice exposed to bleomycin, Muc5b expression is temporally associated with markers of ER stress. Using bulk and single-cell RNA sequencing in bleomycin-exposed mice, we found that pathologic ER stress-associated transcripts Atf4 and Ddit3 (DNA damage inducible transcript 3) were elevated in alveolar epithelia of SFTPC-Muc5b transgenic (SFTPC-Muc5bTg) mice relative to wild-type (WT) mice. Activation of the ER stress response inhibits protein translation for most genes by phosphorylation of Eif2α (eukaryotic translation initiation factor 2 alpha), which prevents guanine exchange by Eif2B and facilitates translation of Atf4. The integrated stress response inhibitor (ISRIB) facilitates interaction of phosphorylated Eif2α with Eif2B, overcoming translation inhibition associated with ER stress and reducing Atf4. We found that a single dose of ISRIB diminished Atf4 translation in SFTPC-Muc5bTg mice after bleomycin injury. Moreover, ISRIB resolved the exaggerated fibrotic response of SFTPC-Muc5bTg mice to bleomycin. In summary, we demonstrate that MUC5B and Muc5b expression is associated with pathologic ER stress and that restoration of normal translation with a single dose of ISRIB promotes lung repair in bleomycin-injured Muc5b-overexpressing mice.

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A mouse model for inherited renal fibrosis associated with endoplasmic reticulum stress.

Publications

A mouse model for inherited renal fibrosis associated with endoplasmic reticulum stress.

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