Development of a novel near-infrared fluorescent probe for nitroxyl imaging in acute kidney injury.

Abstract

Nitroxyl (HNO), a short-lived reactive nitrogen species, participates in regulating cellular redox and nitrosative homeostasis, and its level fluctuations may reflect biochemical alterations associated with renal tubular injury during acute kidney injury (AKI). Therefore, the development of HNO fluorescent probes for AKI models may help visualize HNO-related changes under kidney injury conditions. In this study, we developed a near-infrared fluorescent probe DCM-HNO based on a dicyanoisophorone fluorophore and a 2-(diphenylphosphino)benzoate HNO-responsive unit. Upon triggering of the phosphine-mediated uncaging process by HNO, the fluorophore is released and the intramolecular charge transfer is restored, resulting in enhanced fluorescence at 700 nm. DCM-HNO exhibited a large Stokes shift of 155 nm, a low detection limit of 76 nM, and good resistance to interference. The low-cytotoxic probe enabled imaging of exogenous HNO and chemically generated intracellular HNO in HK-2 cells. In cisplatin-injured HK-2 cells, the fluorescence signal increased with rising cisplatin concentration and decreased upon N-acetylcysteine treatment, confirming the capability of DCM-HNO to visualize HNO-related fluorescence changes during cellular AKI. Ex vivo kidney imaging further showed enhanced fluorescence in kidneys from cisplatin-induced AKI mice. Moreover, significantly stronger fluorescence was exhibited by ischemia-reperfusion kidneys compared with untreated contralateral kidneys. Collectively, DCM-HNO may be considered a near-infrared imaging tool for monitoring HNO-associated alterations in cellular and mouse models of AKI.