Klotho Ameliorates Renal Fibrosis in a Murine Model of ADTKD‐UMOD by Enhancing UMOD secretion and Downregulation of TGFBI

Heterozygous Uromodulin (UMOD) mutations cause autosomal dominant tubulo‐interstitial kidney disease (ADTKD). ADTKD‐ UMOD is a model for renal fibrosis. Currently, no therapies are available for ADTKD‐ UMOD . The antiaging hormone Klotho improves glomerular disease, hypertension, uremic cardiomyopathy, and acute kidney injury (AKI). We tested if Klotho ameliorates renal fibrosis and tubular disease in a murine model of ADTKD‐ UMOD . To generate a more pronounced phenotype we used homozygous Umod C93F/C93F mice. Mice were crossed with mice overexpressing Klotho (TgKl). We compared the phenotype with wild‐type (WT), TgKl, and Umod C93F/C93F mice (8 animals per group). We studied blood pressure, urine and blood samples at ages of 6 and 13 months. In different animals of the four groups, we harvested kidneys at 3 months to perform quantitative proteome analysis via LC‐MS/MS on renal outer medulla. Compared to Umod C93F/C93F mice, TgKl/ Umod C93F/C93F had better renal function as indicated by significantly lower BUN, creatinine, and cystatin C values, and less renal fibrosis. PTH and FGF23 improved significantly in TgKl/ Umod C93F/C93F vs. Umod C93F/C93F mice. Mutant UMOD is retained in the endoplasmic reticulum. We found that relative urinary UMOD secretion was higher in TgKl/ Umod C93F/C93F vs. Umod C93F/C93F . No major serum electrolyte differences were found among the four groups. Urinary studies in TgKl/ Umod C93F/C93F mice showed less urinary wasting of Na + , Ca 2+ , Mg 2+ and Phosphorus than in Umod C93F/C93F mice. Compared to Umod C93F/C93F , Klotho‐overexpressing animals had significantly lower systolic and diastolic blood pressures. Markers of cardiac hypertrophy such as ANP, BNP, and MYH7 mRNA were significantly lower in TgKl/ Umod C93F/C93F than in Umod C93F/C93F animals. We confirmed impaired unfolded protein response (UPR) and autophagy in Umod C93F/C93F compared to WT mice. However, improved renal function and fibrosis in Klotho‐overexpressing mice were not due to changes in UPR or autophagy. Therefore, we performed an unbiased proteomics approach. We identified 262 downregulated and 179 upregulated proteins in kidneys. Analyzing proteins by signaling pathways, we identified in TgKl/ Umod C93F/C93F vs. Umod C93F/C93F animals downregulation of Transforming growth factor‐beta‐induced protein (TGFBI) together with collagens Col1a1, Col4a3, Col4a4, Col4a5, Col6a1, Col6a2, Col6a3, Col12a1, Col14a1, prolargin, biglycan, and osteoglycin/mimecan. We confirmed lower Col1a1 mRNA expression with qPCR. We show that Klotho improves renal outcome in ADTKD‐ UMOD by increasing urinary UMOD secretion and ameliorating renal fibrosis by downregulation of TGFBI and collagen. TGFBI is secreted by mesenchymal stromal cells and infiltrating macrophages and stimulates collagen synthesis. Infiltrating macrophages are an important part of AKI and ADTKD‐ UMOD . Our data reveal a new specific target for treatment of renal fibrosis. Support or Funding Information NIH R03 DK111776, NIH P30 DK079328, and CCRAC Senior Investigator Bridge Award, Children’s Health Dallas At 6 and 13 months TgKl/Umod C93F/C93F show significantly lower BUN, creatinine, and cystatin C values. Figure 1 STRING analysis showing networks formed by downregulated proteins in renal outer medulla of TgKl/UmodC93F/C93F compared to UmodC93F/C93F mice. Red circle outlines downregulated TGFBI and collagens. Figure 2