418 Urinary cell iron processing as a noninvasive indicator of iron toxicity and treatment response in CKD

Objectives/Goals: Define how CKD and iron therapy influence tubular epithelial cell (TEC) iron handling through parallel kidney and urine studies in mice and patients, uncovering cellular signatures that predict injury or recovery and may guide individualized iron therapy in CKD. Methods/Study Population: We used two mouse models of CKD and fresh urine samples from pediatric CKD patients and healthy controls to assess tubular iron handling. Kidney tubular epithelial cells (TECs; CD45 − AQP1 + ) were analyzed by flow cytometry for labile iron pool (LIP) using FerroOrange and sorted into LIP^hi and LIP^low subsets for RNA-seq. Expression of injury markers such as KIM1 and NGAL was used to evaluate tubular stress, linking iron accumulation to epithelial injury and adaptive responses. By combining mouse CKD models with patient urine analysis, we traced how kidney tubular epithelial cells (TECs) manage iron across species. Results/Anticipated Results: In CKD mouse models, tubular epithelial cells (TECs) showed reduced labile iron pool (LIP) and increased ferroptosis, with a smaller population of iron-overloaded cells exhibiting elevated KIM-1 and EMT-related transcriptional changes. Iron chelation lowered LIP in tubular cells and iron therapy exasperated KIM-1 expression. Urinary TECs mirrored tissue findings, with LIP and KIM-1 levels reflecting systemic iron status and CKD severity. In human CKD, urinary LIPHI TECs expressed higher KIM-1 and injury-associated signatures, validating urinary TEC profiling as a noninvasive readout of tubular health. In both species, iron handling and toxicity proceeded differently depending on the nature of the damage (tubular or glomerular), identifying urinary TECs as a noninvasive window into kidney iron toxicity. Discussion/Significance of Impact: Chronic kidney disease affects 10–15% of the world population, and anemia is a common complication. Iron therapy is toxic to kidney cells, contributing to tubular injury. Our studies identify urinary cells as a noninvasive test of renal iron status and link iron handling to injury, enabling safer, personalized therapy to slow kidney damage in patients.