NIR-excited imaging and in vivo visualization of ß-galactosidase activity using a pyranonitrile-modified upconversion nanoprobe.

ß-galactosidase (ß-gal) is a diagnostic biomarker of primary ovarian cancers. The development of effective fluorescent probes for investigating the activity of ß-gal will be beneficial to cancer diagnosis. Herein, a near-infrared (NIR) excited ratiometric nanoprobe (DCM-ß-gal-UCNPs) by assembling pyranonitrile dye (DCM-ß-gal) on the surface of upconversion nanophosphors (UCNPs) was designed for the evaluation of ß-gal activity in vivo. Upon the interaction with ß-gal, a marked decrease of upconversion luminescence (UCL) signal in the green channel was observed owing to the luminescence resonance energy transfer from the UCNPs to pyranonitrile chromophore, whereas the NIR UCL emission at 800 nm was almost no influence. Thus, the ß-gal activity could be quantitatively detected by the UCL intensity ratio of UCL543 nm/UCL800 nm with the limit of detection of 3.1 × 10-4 U/mL. Moreover, DCM-ß-gal-UCNPs was effectively applied for monitoring ß-gal fluctuation in living cells and zebrafish by a ratiometric UCL signal excited by 980 nm laser. We envision that nanoprobe DCM-ß-gal-UCNPs might be used as a potential bioimaging tool to disclose more biological information of ß-gal in ß-gal-associated diseases in the future.

A novel near-infrared ratiometric fluorescent probe targeting lysosomes for imaging HOCl in vitro and in vivo.

Hypochlorous acid (HOCl), as an important biological reactive oxygen species (ROS), plays an important role in microbial immune defense and inflammatory response. Abnormal levels of HOCl in lysosomes can cause lysosomal membrane rupture and release of various hydrolases, leading to a variety of diseases, including cancer. In order to better monitor the level of HOCl in lysosomes, phenothiazine was chosen as fluorophore to construct a NIR fluorescent probe PMM with intramolecular change transfer process (ICT). PMM is a colorimetric and ratiometric fluorescent probe, which has high sensitivity with a low detection limit (20 nM), high selectivity and anti-interference. PMM has good stability in the weakly acidic environment of pH 4.0-5.5. PPM has good localization ability for lysosomes and has been successfully used for fluorescence imaging of exogenous and endogenous HOCl in HepG2 cells. Moreover, nude mouse imaging also demonstrated that PMM could be used to detect HOCl in vivo.

Deep-Depth Imaging of Hepatic Ischemia/Reperfusion Injury Using a Carbon Monoxide-Activated Upconversion Luminescence Nanosystem.

Exploring a chemical imaging tool for visualizing the endogenous CO biosignaling molecule is of great importance in understanding the pathophysiological functions of CO in complex biological systems. Most of the existing CO fluorescent probes show excitation and emission in the region of ultraviolet and visible light, which are not suitable for application in in vivo deep-depth imaging of CO. Herein, a new near-infrared (NIR) to NIR upconversion luminescence (UCL) nanosystem for in vivo visualization of CO was developed, which possesses the merits of high selectivity and sensitivity, a deep tissue penetration depth, and a high signal-to-noise ratio. In this design, upon interaction with CO, the maxima absorption peak of the nanosystem showed a significant blue shift from 795 nm to 621 nm and triggered a remarkable turn-on NIR UCL signal due to the luminescence resonance energy transfer process. Leveraging this nanosystem, we achieved an NIR UCL visualization of the generation of CO biosignals caused by hypoxic, acute inflammation, or ischemic injury in living cells, zebrafish, and mice. Moreover, the protective effect of CO in zebrafish models of oxygen and glucose deprivation/reperfusion (OGD/R) and mice models of lipopolysaccharide-induced oxidative stress (LOS) and hepatic ischemia/reperfusion (HI/R) was also further verified. Therefore, this work discloses that the nanosystem not only serves as a promising nanoplatform to study biological signaling pathways of CO in pathophysiological events, but may also provide a powerful tool for HI/R injury diagnosis.

A Hemicyanine-Assembled Upconversion Nanosystem for NIR-Excited Visualization of Carbon Monoxide Bio-Signaling In Vivo.

Carbon monoxide (CO) is considered as the second gasotransmitter involved in a series of physiological and pathological processes. Although a number of organic fluorescent probes have been developed for imaging CO, these probes display excitation within the ultraviolet or visible range, which restrict their applications in the complex biosystems. In the present work, a strategy is developed to construct an upconversion nanoparticles-based nanosystem for upconversion luminescent (UCL) sensing CO. This nanosystem exhibits a fast response to CO with high sensitivity and selectivity in aqueous solution by a near-infrared-excited ratiometric UCL detection method. Meanwhile, laser scanning upconversion luminescence microscope experiments demonstrate that this nanosystem can visualize the endogenous CO bio-signaling in living cells, deep tissues, zebrafish, and living mice by ratiometric UCL imaging. In particular, this nanosystem has been successfully employed in visualization of the endogenous CO bio-signaling through up-regulation of heme oxygenase-1 (HO-1) in the progression of hypoxia, acute inflammation, or ischemic injury. This work demonstrates that the outstanding performance of the nanosystem not only can provide an effective tool for further understanding the role of CO in the physiological and pathological environment, but also may have great potential ability for tracking the expression of HO-1 in living systems.