Xiuqi Kong

Development of a Two‐Photon Fluorescent Probe for Imaging of Endogenous Formaldehyde in Living Tissues

Investigation of the physiological and pathological functions of formaldehyde (FA) are largely restricted by a lack of useful FA imaging agents, in particular, those that allow detection of FA in the context of living tissues. Herein, we present the rational design, synthesis, and photophysical property studies of the first two-photon fluorescent FA probe, Na-FA. Importantly, the highly desirable attributes of the probe Na-FA (such as a very large turn-on signal (up to 900-fold), a low detection limit, and a very fast onset imparted by the unique design aspects of the probe), make it possible to monitor endogenous FA in living tissues for the first time. Furthermore, sodium bisulfite was identified as a simple and convenient inhibitor of FA within biological environments.

Simultaneous Near-Infrared and Two-Photon In Vivo Imaging of H2O2 Using a Ratiometric Fluorescent Probe based on the Unique Oxidative Rearrangement of Oxonium

A new ratiometric fluorescent H2 O2 probe, benzopyrylium-coumarin (BC), is designed by using an oxonium moiety as the unique H2 O2 response site. The BC probe exhibits an extremely large emission shift of 221 nm in response to H2 O2 , and is successfully applied for the simultaneous near-infrared and two-photon imaging of H2 O2 in living cells, mouse-liver tissues, and zebrafish.

A lysosome-targeted and ratiometric fluorescent probe for imaging exogenous and endogenous hypochlorous acid in living cells

Hypochorous acid plays important roles in numerous physiological and pathological processes. At the cell organelle level, an abnormal concentration of hypochorous acid in the lysosomes causes redox imbalance and the loss of function of the lysosomes. Herein, the first small molecule based, lysosomal-targeted ratiometric fluorescent HOCl probe (Lyso-HA) was synthesized through a rational design. The new probe was highly selective toward HOCl over other reactive oxygen species and exhibits a large variation (up to 97-fold) in its fluorescence ratio (I585/I450), with good signal resolution. The probe Lyso-HA is membrane-permeable and is suitable for ratiometric visualization of exogenous and endogenous HOCl at lysosomes in living cells.

Single Fluorescent Probe for Dual-Imaging Viscosity and H2O2 in Mitochondria with Different Fluorescence Signals in Living Cells

Mitochondria, as essential and interesting organelles within the eukaryotic cells, play key roles in a variety of pathologies, and its abnormalities are closely associated with Alzheimers disease (AD) and other diseases. Studies have shown that the abnormal of viscosity and concentration of hydrogen peroxide in mitochondria were all associated with AD. Accordingly, the detection of viscosity and hydrogen peroxide in mitochondria has attracted great attention. However, it remains a great challenge to explore a single probe, which can dual-detect the viscosity and H2O2 in mitochondria. Herein, in two ways to prevent the twisted internal charge transfer (TICT) process, we designed and sythesized the first dual-detection fluorescent probe Mito-VH that can visualize viscosity and H2O2 in mitochondria with different fluorescence signals in living cells.

Rational Design of a Robust Fluorescent Probe for the Detection of Endogenous Carbon Monoxide in Living Zebrafish Embryos and Mouse Tissue

Carbon monoxide (CO) is one of the most important gaseous signal molecules in biological systems. However, the investigation of the functions of CO in living organisms is restricted by the lack of functional molecular tools. To address this critical challenge, we present herein the rational design, synthesis, and in vivo imaging studies of a powerful two-photon excited near-infrared fluorescent probe (1-Ac) for endogenous CO monitoring. The advantageous features of the new probe include high stability, low background fluorescence, large fluorescence enhancement, high sensitivity, and two-photon excitation with emission in the near-infrared region. Significantly, these merits of the probe enable the tracking of endogenous CO in zebrafish embryos and mouse tissues for the first time.

A novel near-infrared fluorescent probe with a large Stokes shift for biothiol detection and application in in vitro and in vivo fluorescence imaging

Biothiols such as cysteine (Cys) and glutathione (GSH) are important reductive species that are widely spread in the liver, heart, brain and so on, and are essential for maintaining the fundamental roles of these organs. Fluorescence detection of biothiols in living organisms is of great importance for exploring the metabolic pathways and physiological function in living organisms. Most of the near-infrared probes for biothiols have small Stokes shifts, usually less than 50 nm, which greatly hamper their applications in biological imaging; herein, a novel fluorescent CS-thiol probe based on a Changsha (CS) near-infrared dye analogue with a large Stokes shift was developed and applied for live animal imaging. In the presence of biothiols, such as Cys, CS-thiols show a broad fluorescence emission at 660 nm and exhibit high selectivity toward biothiols among other amino acids and reactive species. CS-Thiols exhibited little toxicity to HeLa cells and were successfully applied to image biothiols in living cells and living animals.

A fast-response two-photon fluorescent probe for the detection of Cys over GSH/Hcy with a large turn-on signal and its application in living tissues

Cysteine (Cys), a small-molecule aminothiol, plays important roles in various physiological processes in connection with various diseases, such as skin lesions, edema, hair depigmentation and liver damage. We developed a novel two-photon fluorescent probe for sensing Cys in presence of GSH and Hcy in vivo. The two-photon fluorescent probe exhibited favorable properties, including fast response (about 20 min), good selectivity, and low cytotoxicity. Furthermore, it was successfully applied for imaging Cys in living cells and tissues.

A tumor-targeting and lysosome-specific two-photon fluorescent probe for imaging pH changes in living cells

Lysosomal pH is closely related to the metastasis and apoptosis of cancer cells. Detecting lysosomal pH changes in cancer cells could be helpful for analyzing tumor progressions and in-depth study of the roles of lysosomes in tumor invasion and metastasis. Herein, we describe a novel tumor-targeting and lysosome-specific two-photon fluorescent probe (BN-lys) for imaging pH changes for the first time. Biotin was employed as the tumor-targeting module, and morpholine was selected as the lysosome-specific group and the pH site to control the fluorescence by photoinduced electron transfer (PET) mechanism. With a pKa value of 5.36, BN-lys showed a fast and reversible fluorescence response to pH. Under the guidance of the biotin group, BN-lys displayed strong one-photon and two-photon fluorescence responses to lysosomal pH in cancer cells, while it displayed weak fluorescence in normal cells. Furthermore, BN-lys could be applied for the imaging of chloroquine-stimulated lysosomal pH changes in living cells. These features demonstrate that this probe could have practical applications in biological research.

Hydrogen Sulfide Triggered Charge-Reversal Micelles for Cancer-Targeted Drug Delivery and Imaging

Currently, the development of polymeric micelles combining diagnosis and targeted therapy is theoretically and practically significant in cancer treatment. In addition, it has been reported that cancer cells can produce large amounts of hydrogen sulfide (H2S) and their survival depends on the content of H2S. In this study, a series of N-(2-hydroxyethyl)-4-azide-1,8-naphthalimide ended amphiphilic diblock copolymer poly(2-hydroxyethyl methacrylate)-block-poly(methyl methacrylate) (N3-Nap-PHEMA-b-PMMA-N3) micelles were prepared. Around cancer tissues, the N3-Nap-PHEMA45-b-PMMA42-N3 micelles exhibited dual characteristics of monitoring H2S and H2S triggered charge reversal with the reduction of the azido group. The surface charge of N3-Nap-PHEMA45-b-PMMA42-N3 micelles reversed from negative to positive after monitoring H2S. With H2S triggered charge reversal, the cellular uptake of DOX-loaded N3-Nap-PHEMA45-b-PMMA42-N3 micelles was effectively enhanced through electrostatic attraction mediated targeting, and a fast doxorubicin (DOX) release rate was observed. The MTT assay demonstrated that N3-Nap-PHEMA45-b-PMMA42-N3 micelles were biocompatible to HeLa cells, and DOX-loaded N3-Nap-PHEMA45-b-PMMA42-N3 micelles showed enhanced cytotoxicity in HeLa cells in the presence of H2S. Furthermore, in vivo fluorescence imaging and biodistribution experiments revealed that DOX-loaded N3-Nap-PHEMA45-b-PMMA42-N3 micelles could provide good tumor imaging and accumulate in tumor tissue. Therefore, N3-Nap-PHEMA45-b-PMMA42-N3 micelles can be used as a promising platform for tumor diagnosis and therapy.

Two-photon red-emissive fluorescent probe for imaging nitroxyl (HNO) in living cells and tissues

Nitroxyl (HNO) plays important roles in the regulation of many physiological and pathological processes, and can serve as a potential therapeutic agent for cardiovascular disease. The development of HNO detection in living systems is greatly important for in-depth studies of its biosynthesis and activities. Herein, we describe a novel two-photon red-emissive fluorescence probe (RP) for imaging HNO in living cells and tissues. RP was based on a red-emissive dye, Rho, and showed no fluorescence. When responding to HNO, RP can emit red fluorescence with the emission wavelength at 638 nm. RP exhibited a sensitive and selective response to HNO. Theoretical calculations demonstrated that the overlaps between the HOMO and LUMO were large for Rho and tiny for RP, consistent with the absorption and fluorescence properties of Rho and RP. Assisted by three-dimensional (3D) imaging, the two-photon imaging of HNO with red emission color in living tissues was successfully performed.