title: Novel assays for detection of urinary KIM-1 in mouse models of kidney injury.
abstract: Kidney injury molecule-1 (KIM-1) has been qualified by the Food and Drug Administration and European Medicines Agency as a urinary biomarker to monitor preclinical nephrotoxicity in rats and on a case-by-case basis for the translation of potentially nephrotoxic drugs into first-in human studies. Although mouse models are widely employed in preclinical studies, few urinary biomarker studies have been performed in mice due to limited urine availability and lack of sensitive assays. Here, we report the development and validation of two different assays for quantitative assessment of mouse urinary KIM-1 (uKIM-1) and compare the sensitivity of KIM-1 relative to other standard markers in ischemia reperfusion and aristolochic acid (AA)-induced kidney injury in mice. A sensitive, reproducible, and quantitative microbead-based KIM-1 ELISA was established, which requires only 10 μl urine for triplicate determination with an assay range of 12.21 pg/ml to 50 ng/ml. The second assay is a laminar flow dipstick assay, which has an assay range of 195 pg/ml to 50 ng/ml and provides quantitative assessment of KIM-1 in 15 min. uKIM-1 levels increased with increasing time of ischemia or time after AA administration. After only 10-min ischemia followed by 24-h reperfusion, uKIM-1 was significantly elevated by 13-fold, whereas serum creatinine (sCr), blood urea nitrogen, N-acetyl-β-glucosaminidase (NAG), and proteinuria levels did not change. After AA administration, uKIM-1 levels were significantly upregulated by greater than threefold within 12 h, whereas sCr and NAG levels were unchanged. Mouse KIM-1 was stable for multiple freeze-thaw cycles, for up to 5 days at room temperature and up to at least an year when stored at -80°C.
authors.affiliation: Renal Division, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA. vsabbisetti@partners.org
authors.forename: Venkata S
authors.initials: VS
authors.lastname: Sabbisetti
authors.forename: Kazumi
authors.initials: K
authors.lastname: Ito
authors.forename: Chang
authors.initials: C
authors.lastname: Wang
authors.forename: Li
authors.initials: L
authors.lastname: Yang
authors.forename: Stephen C
authors.initials: SC
authors.lastname: Mefferd
authors.forename: Joseph V
authors.initials: JV
authors.lastname: Bonventre
mesh_terms: D000818:Animals; D034341:Aristolochic Acids / Q000633:toxicity; D001681:Biological Assay; D015415:Biomarkers / Q000097:blood / Q000652:urine; D004195:Disease Models, Animal; D000072596:Hepatitis A Virus Cellular Receptor 1; D007668:Kidney / Q000098:blood supply / Q000187:drug effects / Q000473:pathology; D007677:Kidney Function Tests; D057230:Limit of Detection; D008297:Male; D008565:Membrane Proteins / Q000652:urine*; D051379:Mice; D051437:Renal Insufficiency / Q000139:chemically induced / Q000209:etiology / Q000473:pathology / Q000652:urine*; D015427:Reperfusion Injury / Q000150:complications / Q000652:urine; D015203:Reproducibility of Results
chemical_list: D034341:Aristolochic Acids; D015415:Biomarkers; C442240:Havcr1 protein, mouse; D000072596:Hepatitis A Virus Cellular Receptor 1; D008565:Membrane Proteins; C000228:aristolochic acid I