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Dendritic cells from mice with an uromodulin mutation demonstrate changes in gene expression resulting in increased migration and efferocytosis capability

ABSTRACT: Introduction: The effects of Uromodulin on the innate immune system remain uncertain. Mice with a cysteine to phenylalanine mutation (C105F) in the UMOD gene homologous to a pathogenic human mutation demonstrate high levels of circulating uromodulin following ischemia reperfusion kidney injury, increased markers of immune tolerance and improved kidney repair. We hypothesized that this may occur through interaction of circulating uromodulin with dendritic cells. Materials and Methods: Dendritic cells were isolated from WT and C105F mutant mice and used to compare gene expression using RNA sequencing. In vitro motility and efferocytosis assays were used to verify phenotypic changes predicted from affected pathways. Results: RNA sequencing demonstrated increased expression of scavenger receptor and motility associated genes. In vitro assays demonstrated that mutant uromodulin increased activation of the NF-kB pathway compared with WT protein and that mutant dendritic cells had increased migration and efferocytosis compared with WT cells. Discussion/Conclusion: Dendritic cells from mutant C105F mice display gene expression and phenotypic changes associated with increased scavenger receptor expression, motility and efferocytosis. These results suggest a possible mechanism of DC regulation by circulating uromodulin that may affect tissue repair following injury.

ORGANISM(S): Mus musculus

PROVIDER: GSE210500 | GEO | 2022/08/06

REPOSITORIES: GEO

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Project description:Background: Uromodulin (Tamm-Horsfall) protein is secreted by the thick ascending limb into urine and crosses the basolateral membrane into the kidney interstitium and blood. Uromodulin may engage the kidney’s innate immune system with pro and anti-inflammatory effects described. Mutations in the UMOD gene are a cause of autosomal dominant tubulointerstitial kidney disease (ADTKD) due to protein misfolding and endoplasmic reticulum stress. We identified a family with a UMOD mutation (C106F) that leads to glomerular and interstitial inflammation atypical for ADTKD. Methods: To determine if the observed phenotype is due to mutant uromodulin induced changes in the innate or adaptive immune system, we developed a transgenic (tg) mouse with a homologous cysteine to phenylalanine mutation (C105F) in the UMOD gene using CRISPR-Cas9 gene editing. Results: LLC-PK1 cells expressing wt and mutant protein had increased basolateral secretion of protein compared with cells expressing wt or mutant protein alone. EM examination of mutant medium showed protein aggregates in contrast to filaments in wt medium. Immunoprecipitation of plasma uromodulin from mutant mice demonstrated aggregate formation not seen in wt mice. Tg/+ mice displayed increased F4/80+ mononuclear phagocyte (MP) and podocyte inflammasome activity at baseline. Tg/+ mice developed glomerular and interstitial matrix deposition, myofibroblast proliferation and increased Creatinine with aging. Following ischemia-reperfusion injury, tg/+ mice had no increase in inflammasome activation over baseline in contrast to wt littermates and displayed improved tubular repair and renal function at 7 days. Differential gene expression analysis by RNA sequencing of kidney MP revealed activation of the ATF4 mediated stress response pathway and genes associated with the development of tissue fibrosis. Conclusions: The C106F mutation results in glomerular and interstitial inflammatory kidney disease due to interaction of aggregated misfolded protein with mononuclear phagocytic cells resulting in ER stress response. The resulting alteration in the immune system leads to improved kidney repair following acute injury but fibrosis and chronic renal failure with aging.

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Dendritic cells from mice with an uromodulin mutation demonstrate changes in gene expression resulting in increased migration and efferocytosis capability

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