Utilizing Resazurin fluorescence to monitor kidney function and organic anion transporter activity - ABSTRACT A comprehensive and accurate assessment of kidney function is essential for managing renal diseases, but current diagnostic methods often provide an incomplete evaluation. Standard tests, such as glomerular filtration rate (GFR) measurements and proteinuria, primarily assess glomerular function and renal perfusion, offering limited insight into tubular health. Furthermore, compensatory increases in single-nephron filtration can sustain GFR even in the presence of nephron loss, often masking disease progression. These limitations are especially apparent in acute kidney injury (AKI), where GFR may return to normal even as tubular dysfunction persists, obscuring the transition to chronic kidney disease (CKD). Direct assessment of tubular function can provide valuable prognostic information by complementing existing diagnostic methods. To this end, we recently identified resazurin dye as a novel, non-invasive sensor for evaluating tubular function and nephron mass. Following intravenous injection, resazurin is selectively taken up by tubular cells, reduced to fluorescent metabolites, and excreted in urine, offering a GFR-independent measure of tubular secretion. In murine models of severe AKI and subclinical nephron loss (unilateral nephrectomy and genetic renal hypoplasia), urinary excretion of resazurin metabolites sensitively reflected both tubular dysfunction and nephron mass loss. Based on these findings, we propose that resazurin provides a rapid, sensitive, and GFR-independent method for quantifying tubular function, nephron mass, and renal reserve following kidney injury. This hypothesis is based on the premise that while nephron loss can lead to compensatory increases in single-nephron filtration to maintain GFR, tubular secretion is less adaptable, making it a more reliable indicator of tubular function and nephron mass. This proposal aims to investigate the tubular transport and metabolism of resazurin (Aim 1) as well as its effectiveness and sensitivity (Aim 2) in quantifying nephron mass loss in murine and porcine models of AKI. The results will provide proof-of-principle data to support clinical translation, facilitating earlier detection of nephron loss, enabling the identification of high-risk AKI patients, and improving monitoring of recovery from AKI. Ultimately, this work has the potential to transform the way we assess kidney function in both preclinical and clinical settings by providing a non-invasive and precise tool for detecting early tubular dysfunction and nephron loss.
