Dual-Channel Fluorescent Probe for the Simultaneous Monitoring of Peroxynitrite and Adenosine-5′-triphosphate in Cellular Applications

Abstract

Changes in adenosine triphosphate (ATP) and peroxynitrite (ONOO − ) concentrations have been correlated in a number of diseases including ischemia-reperfusion injury and drug-induced liver injury. Herein, we report the development of a fluorescent probe ATP-LW, which enables the simultaneous detection of ONOO − and ATP. ONOO − selectively oxidizes the boronate pinacol ester of ATP-LW to afford the fluorescent 4-hydroxy-1,8-naphthalimide product NA-OH (λ ex = 450 nm, λ em = 562 nm or λ ex = 488 nm, λ em = 568 nm). In contrast, the binding of ATP to ATP-LW induces the spirolactam ring opening of rhodamine to afford a highly emissive product (λ ex = 520 nm, λ em = 587 nm). Due to the differences in emission between the ONOO − and ATP products, ATP-LW allows ONOO − levels to be monitored in the green channel (λ ex = 488 nm, λ em = 500−575 nm) and ATP concentrations in the red channel (λ ex = 514 nm, λ em = 575−650 nm). The use of ATP-LW as a combined ONOO − and ATP probe was demonstrated using hepatocytes (HL-7702 cells) in cellular imaging experiments. Treatment of HL-7702 cells with oligomycin A (an inhibitor of ATP synthase) resulted in a reduction of signal intensity in the red channel and an increase in that of the green channel as expected for a reduction in ATP concentrations. Similar fluorescence changes were seen in the presence of SIN-1 (an exogenous ONOO − donor).

Progress and perspectives: fluorescent to long-lived emissive multifunctional probes for intracellular sensing and imaging

“…Apart from the development of multi-organelle targeting molecular probes, multi-analyte responsive probes have been employed to monitor the intracellular changes due to the different analyte levels under diverse pathophysiological conditions. 90,91 For example, the overexpression of HClO is known to cause oxidative stress in cells and is related to several neurodegenerative diseases. 90,92 However, H 2 S acts as an HClO inhibitor and reduces the chances of oxidative stress.…”

Genetically Encoded ATP Biosensors for Direct Monitoring of Cellular ATP Dynamics

“…Furthermore, simultaneous or parallel analyses of ATP signals with other crucial biological parameters, such as calcium, pH, and ROS, should open new windows to previously unknown biological interactions during energy metabolism with different compartments of cells. For example, Wu et al recently developed a dual-channel fluorescent probe for the simultaneous monitoring of peroxynitrite and ATP [ 169 ]. These innovative technologies enable spatiotemporal analyses of ATP dynamics along with other key metabolites in living cells under physiological and pathological contexts.…”

An efficient PeT based fluorescent probe for mapping mitochondrial oxidative stress produced via the Nox2 pathway

“…Small molecular fluorescent probes play a pivotal role in offering deep insight into living systems to elucidate mechanistic pathways because of their simplicity, non-invasive nature, and high sensitivity with good resolution at organelles. [17][18][19][20][21][22][23][24][25][26][27][28][29][30][31][32][33] To date, several HOCl responsive fluorescent probes have been reported based on well-known fluorophore small molecules as well as using DNA-based technologies. [34][35][36][37][38][39] Several reported activity-based fluorescent probes (ABFP) are limited by their synthetic difficulties, modulation of the optical properties, aqueous solubility, and especially organelle targeting ability.…”