Martinez Shirely Acosta, Karel Isaac Z, Silvaroli Josie A, Ahmed Eman, Kim Ji Young, Stayton Amanda, Patel Prisha S, Afjal Mohammad Amir, Horton Thomas, Bohmer Margaret, Vanichapol Thitinee, Sander Veronika, Andrade Gabriel Mayoral, Allison Corynne Vermillion, Mondal Milon, Thorson Victoria C, Partey Alexandra, Nimkar Kartik, Williams Victoria, Quimby Jessica, Ganesan Latha, Madhavan Sethu M, Davidson Alan J, Peterson Blake R, Adebiyi Adebowale, Rao Reena, Sweet Douglas H, Singh Prabhleen, Bennett Kevin M, Zepeda-Orozco Diana, Mallipattu Sandeep K, Eisenmann Eric D, Sparreboom Alex, Rovin Brad H, Bajwa Amandeep, Pabla Navjot S
Division of Pharmaceutics and Pharmacology, College of Pharmacy and Comprehensive Cancer Center, The Ohio State University, Columbus, Ohio, USA.
Transplant Research Institute, Department of Surgery, College of Medicine, The University of Tennessee Health Science Center, Memphis, Tennessee, USA.
Kidney Int. 2025 Mar;107(3):508-520. doi: 10.1016/j.kint.2024.12.008. Epub 2024 Dec 27.
Glomerular filtration rate (GFR) is the main functional index of kidney health and disease. Currently, no methods are available to directly measure tubular mass and function. Here, we report a serendipitous finding that the in vitro cell viability dye resazurin can be used in mice as an exogenous sensor of tubular function. Intravenously injected resazurin exhibited significant plasma protein binding and was found to mainly undergo tubular secretion. Mechanistic studies showed that the blue-colored, weakly fluorescent resazurin is taken up by tubular cells through organic anion transporters, followed by conversion to a highly fluorescent, pink-colored resorufin by mitochondrial and cytosolic reductases, converted to an orange-colored β-d-glucuronide with subsequent efflux into the urine. Here we report a simple method in which the intravenous injection of resazurin is followed by the measurement of fluorescent metabolites in the urine, providing a sensitive readout of tubular function. Three mouse models of acute kidney injury (rhabdomyolysis, bilateral ischemia-reperfusion injury, and cisplatin nephrotoxicity) were tested and the resazurin-based method was able to sensitively detect the loss of tubular function much earlier than the increase in serum creatinine levels. Strikingly, in mice with unilateral ischemia-reperfusion injury and genetic mutation-linked kidney hypoplasia (oligosyndactylism, a genetic model for congenital kidney hypoplasia), the resazurin-based method was able to detect loss of tubular mass and function despite normal GFR levels. Collectively, our findings establish the preclinical utility of resazurin as a sensitive exogenous marker of tubular function and support future examination in larger animals for potential clinical translation.
肾小球滤过率（GFR）是肾脏健康与疾病的主要功能指标。目前，尚无直接测量肾小管质量和功能的方法。在此，我们报告一项意外发现：体外细胞活力染料刃天青可在小鼠中用作肾小管功能的外源性传感器。静脉注射的刃天青表现出显著的血浆蛋白结合，并被发现主要通过肾小管分泌。机制研究表明，蓝色、弱荧光的刃天青被肾小管细胞通过有机阴离子转运体摄取，随后被线粒体和胞质还原酶转化为高荧光的粉红色试卤灵，再转化为橙色的β-D-葡萄糖醛酸苷，随后排入尿液。在此我们报告一种简单方法，即静脉注射刃天青后测量尿液中的荧光代谢产物，可提供肾小管功能的灵敏读数。对三种急性肾损伤小鼠模型（横纹肌溶解、双侧缺血-再灌注损伤和顺铂肾毒性）进行了测试，基于刃天青的方法能够比血清肌酐水平升高更早地灵敏检测到肾小管功能丧失。令人惊讶的是，在单侧缺血-再灌注损伤和基因突变相关的肾发育不全（多指畸形，一种先天性肾发育不全的遗传模型）小鼠中，尽管GFR水平正常，基于刃天青的方法仍能够检测到肾小管质量和功能的丧失。总体而言，我们的研究结果确立了刃天青作为肾小管功能灵敏外源性标志物的临床前实用性，并支持未来在大型动物中进行潜在临床转化的研究。
