Rational design of an activatable dual-color fluorogenic probe for revealing the interaction of adenosine-5′-triphosphate and peroxynitrite in pyroptosis associated with acute kidney injury

Abstract

ATP and ONOO- play unique roles in various biological events and exhibit notable interactions. To date, there is no available chemical tool for investigating the correlation between ATP and ONOO- concentrations in pyroptosis associated with acute kidney injury (AKI). Herein, we designed a novel dual-color near-infrared fluorescent (NIRF) probe P2 for simultaneous visualization of ATP and ONOO- both in vitro and in vivo. Unlike previously reported single-site fluorescent probes, P2 enabled concurrent imaging of ATP and ONOO- in two distinct fluorescence channels, with emission wavelengths centered at 585 and 690 nm, which greatly reduced spectral cross-talk. Employing a HK-2 pyroptosis model, a significant interaction between ATP and ONOO- was unveiled. Elevated ONOO- production was found to correlate with decreased ATP levels; conversely, an increase in ATP levels was associated with rapid ONOO- scavenging. Remarkably, P2 allowed the assessment of cellular hypoxia by monitoring ATP and ONOO- concentrations. The commercial ONOO--scavenger uric acid showcased a protective effect on HK-2 cells via inhibition of the cellular pyroptosis pathway. Furthermore, P2 was successfully employed for imaging of ATP and ONOO- in the AKI mice model. Our findings confirmed that renal ischemia-reperfusion triggered a rise in ONOO- levels, concurrent with a decline in ATP levels. Surprisingly, the cells exhibited a compensatory recovery of ATP levels as the reperfusion time was prolonged. These results suggested the newly devised P2, as a pivotal chemical tool for the simultaneous monitoring of ATP and ONOO-, might open new avenues for disease diagnosis and treatment.