HNO In Cells Research Articles

A near-infrared fluorescent probe with ultra-large Stokes shift for the detection of HNO in cells and mice

As one of the significant reactive nitrogen species in living systems, nitroxyl (HNO) plays a paramount role in a series of physiological and pathological processes. However, completely elucidating the role of endogenous HNO in living systems remains a huge challenge due to the lack of effective detection tools for the real-time monitoring of HNO. Recently, fluorescent probe has been widely accepted as a powerful molecular tool for the non-invasive detection of active species in living systems. Hence, in this work, we developed a new NIR fluorescent probe (N-HBT-HNO) with ultra-large Stokes shift for the effective detection and bioimaging of HNO in living cells and mouse. In the design blueprint of probe, the dicyanoisophorone moiety was integrated into the benzothiazole unit to expand the emission wavelength to NIR region of 728 nm and remarkably increase the Stokes shift to ultra large of 303 nm, which could endow the probe with superiority in bioimaging. Additionally, the 2-(diphenylphosphino) benzoate served as the efficient recognition group and fluorescence quenching group. Results demonstrated that the newly developed probe possessed appreciable sensing performance, including good selectivity, large signal to noise ratio, high sensitivity and fast response toward HNO. Subsequently, this probe was successfully applied for the bioimaging of exogenous HNO in living cells and mouse, reflecting the application potential of exogenous HNO (AS).

An isophorone-fused near-infrared fluorescent probe with a large Stokes shift for imaging endogenous nitroxyl in living cells and zebrafish.

Nitroxyl (HNO) plays an important role in multiple physiological and pathological processes, but the detailed generation mechanism of the endogenous HNO still remained to explore and perfect further. There is an urgent need to develop an excellent fluorescent probe for selective recognition and sensitive detection of HNO in biological systems. Near-infrared (NIR) fluorescent probes with a large Stokes shift are an ideal tool for bioimaging applications. Here, we have developed a NIR fluorescent probe with a large Stokes shift, namely, NIR-HNO, to monitor HNO in cells and zebrafish. NIR-HNO consists of an isophorone-fused NIR fluorescence reporter and a diphenylphosphinobenzoyl HNO-responsive unit. Based on an aza-ylide intramolecular ester aminolysis reaction, NIR-HNO showed a rapid selective NIR fluorescent turn-on response for HNO, high sensitivity (detection limit was 39.6 nM), and large Stokes shift (265 nm). The biological imaging results indicate that NIR-HNO is a good candidate for imaging of endogenous HNO in living systems.

A Near-Infrared Fluorescent Probe for Detection of Nitroxyl in Living Cells

Nitroxyl (HNO), the one-electron reduced and protonated congener of nitric oxide (NO), has been demonstrated with excellent bio-pharmacological effects in cardiovascular disorder treatment, which is distinctive from that of NO. Despite its high reactivity, the accurate detection of HNO is a challenging issue. To resolve this problem, in this work, a near-infrared (NIR) metal-free fluorescent probe, ER-JN, was designed and synthesized for the detection of intracellular HNO concentration in simulated physiological conditions and living cells. The probe was consisted of two moieties, the BF2-chelated tetraarylazadipyrromethane fluorophore (aza-BODIPY) and the HNO recognition unit, diphenylphosphinobenzoyl group. The probe was purified by silica column chromatography eluting with CH2Cl2 to obtain the green solid product with a yield of 28%. The as-prepared probe exhibited high sensitivity, good selectivity and low cytotoxicity, and was applied to fluorescent bio-imaging of HNO in simulated physiological conditions. When used in detection of HNO, quantum yield of the probe increased from 0.01 to 0.35. The linear range was 0-50 mu M, with the detection limit of 0.03 mu M (S/N = 3). With confocal laser scanning microscope imaging analysis, the probe could be used to detect HNO concentration in living cells. Furthermore, the results of flow cytometry confirmed that the probe could be employed for the qualitative and quantitative detection of intracellular HNO level. In this work, we found that probe ER-JN could not only detect HNO in aqueous solution and in living cells, but also target endoplasmic reticulum. We anticipate that ER-JN will provide experimental bases in studying physiological and pathological functions of HNO in cells, in vitro and in vivo.