Elevated urinary CRELD2 is associated with endoplasmic reticulum stress–mediated kidney disease
Yeawon Kim, … , Anthony J. Bleyer, Ying Maggie Chen
Yeawon Kim, … , Anthony J. Bleyer, Ying Maggie Chen
Published December 7, 2017
Citation Information: JCI Insight. 2017;2(23):e92896. https://doi.org/10.1172/jci.insight.92896.
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Research Article Nephrology

Elevated urinary CRELD2 is associated with endoplasmic reticulum stress–mediated kidney disease

Abstract

ER stress has emerged as a signaling platform underlying the pathogenesis of various kidney diseases. Thus, there is an urgent need to develop ER stress biomarkers in the incipient stages of ER stress–mediated kidney disease, when a kidney biopsy is not yet clinically indicated, for early therapeutic intervention. Cysteine-rich with EGF-like domains 2 (CRELD2) is a newly identified protein that is induced and secreted under ER stress. For the first time to our knowledge, we demonstrate that CRELD2 can serve as a sensitive urinary biomarker for detecting ER stress in podocytes or renal tubular cells in murine models of podocyte ER stress–induced nephrotic syndrome and tunicamycin- or ischemia-reperfusion–induced acute kidney injury (AKI), respectively. Most importantly, urinary CRELD2 elevation occurs in patients with autosomal dominant tubulointerstitial kidney disease caused by UMOD mutations, a prototypical tubular ER stress disease. In addition, in pediatric patients undergoing cardiac surgery, detectable urine levels of CRELD2 within postoperative 6 hours strongly associate with severe AKI after surgery. In conclusion, our study has identified CRELD2 as a potentially novel urinary ER stress biomarker with potential utility in early diagnosis, risk stratification, treatment response monitoring, and directing of ER-targeted therapies in selected patient subgroups in the emerging era of precision nephrology.

Authors

Yeawon Kim, Sun-Ji Park, Scott R. Manson, Carlos A.F. Molina, Kendrah Kidd, Heather Thiessen-Philbrook, Rebecca J. Perry, Helen Liapis, Stanislav Kmoch, Chirag R. Parikh, Anthony J. Bleyer, Ying Maggie Chen

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Figure 1

CRELD2 is a urinary biomarker for detecting podocyte ER stress in the early stages of NS.

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CRELD2 is a urinary biomarker for detecting podocyte ER stress in the ea...
Primary podocytes (P1 or P2) were isolated and cultured from WT mice at P27 (A–C). (A) Mouse primary podocytes were treated with DMSO (control), TM (2 μg/ml), or TG (1 μM) for 24 h. Cell lysates were analyzed by WB with the indicated antibodies. (B) Control and TM- or TG-treated mouse podocytes, as described in A, were stained for CRELD2 (red) and BiP (green). Nuclei were counterstained with Hoechst 33342 (blue). Scale bar: 40 μm. (C) Immunoblot analysis for CRELD2 secretion into the media by cultured primary podocytes described in A (20 μl medium each group). (D) Quantitative PCR analysis of CRELD2 mRNA in primary podocytes from Lamb2+/–, Lamb2–/–;Tg-C321R, and Lamb2–/–;Tg-WT mice at P27. CRELD2 expression was normalized as a ratio to mouse podocyte–specific WT-1 mRNA, and the average CRELD2/WT-1 mRNA ratio in Lamb2+/– mice was set as 1 (n = 5 mice per genotype; mean ± SD). *P < 0.05 by 1-way ANOVA. (E) Primary podocyte cell lysates from age-matched Lamb2+/–, Lamb2–/–;Tg-C321R, and Lamb2–/–;Tg-WT mice at P27 were analyzed by WB with the indicated antibodies. (F) Immunoblot analysis for CRELD2 secretion into the media by primary podocytes (20 μl each group) from the indicated genotypes at P27. (G) Urinary CRELD2 excretion was assessed by WB from Lamb2–/–; Tg-C321R mice and their Lamb2+/– littermates and age-matched Lamb2–/–;Tg-WT mice (n = 8, n = 8, and n = 3, respectively). Shown is 1 representative experiment at the indicated ages. (H) Immunoblot analysis of CRELD2 in crude urine specimens from 1 pair of Lamb2+/– and Lamb2–/–;Tg-C321R littermates following the disease course. Shown is 1 representative experiment. Three more independent experiments were performed with similar results. (I) CRELD2 excretion in unprocessed urine specimens was detected by WB from Lamb2–/–;Tg-C321R mice and their Lamb2+/– littermates and Lamb2–/–;Tg-WT mice at 8–16 weeks. The urinary CRELD2 excretion was normalized to urine Cr excretion such that the urine volume applied to the gel reflected the amount of urine containing 1 μg of Cr for G–I. All WB images are representative of at least 3 independent experiments. CRELD2, cysteine-rich with EGF-like domains 2; TM, tunicamycin; TG, thapsigargin; BiP, Ig binding protein; Tg-C321R, Lamb2–/– mice expressing C321R-LAMB2 in podocytes; Tg-WT, Lamb2–/– mice expressing WT LAMB2 in podocytes.