Assessment of cancer cell migration using a viscosity-sensitive fluorescent probe.

A novel viscosity probe (NV1) was developed for assessing cancer cell migration. NV1 can respond to changes of viscosity rapidly and exhibits high sensitivity in HepG2 cells treated with starvation, rotenone and nystatin. Importantly, NV1 was used for the first time to evaluate the relationship between intracellular viscosity changes and cancer cell migration and proved that increased intracellular viscosity inhibits cell migration while decreased intracellular viscosity promotes cell migration.

A Mitochondrial-Targeting Near-Infrared Fluorescent Probe for Revealing the Effects of Hydrogen Peroxide And Heavy Metal Ions on Viscosity.

As an important cell organelle, the mitochondrion has special viscosities, while abnormal mitochondrial viscosity is closely related to many diseases. Hydrogen peroxide (H2O2) is an active molecule related to the cell microenvironment, and its influence on mitochondrial viscosity is still not clear, so further investigation is needed. In addition, since excessive accumulation of heavy metal ions would lead to cells' dysfunction, the study of effect of excessive heavy metal ions on mitochondrial viscosity has not been reported. Herein, we designed and synthesized a mitochondrial-targeting near-infrared fluorescent probe (Mito-NV) for real-time in situ imaging and analysis of mitochondrial viscosity. Furthermore, the probe revealed that H2O2 can raise mitochondrial viscosity, while heavy metal ions reduce the viscosity. This work is of great significance for understanding the execution of mitochondrial functions and the occurrence and development of related diseases.

A mitochondria-targeting near-infrared fluorescent probe for imaging hypochlorous acid in cells.

Hypochlorous acid (HOCl), as an important reactive oxygen species (ROS), plays an important role in various pathological and physiological processes. Therefore, it is of great significance to identify and detect HOCl in organisms. Herein, a mitochondria-targeting near-Infrared fluorescent probe (CVS) has been designed and synthesized for sensing HOCl. Under simulated physiological conditions, CVS can be instantly oxidized by HOCl to CVSO and the fluorescence signal is significantly enhanced. Additionally, CVS shows high selectivity toward HOCl over other ROS and owns low detection limit (94.7 nM) for HOCl. Moreover, CVS can be successfully applied to image HOCl in mitochondria of the HepG2 cells. Thus, CVS is a powerful tool for sensing HOCl and provides a perspective method for further study of the physiological and pathological functions related to HOCl.

Fishing out Methionine-Containing Proteins from Complex Biological Systems Based on a Non-Enzymatic Biochemical Reaction.

Nowadays, it still remains a great challenge to develop a simple, fast, and low-toxic method for identification and separation of proteins from complex biological systems. Herein, a nanocomposite (Fe3O4@Au-Se-peptide) was designed and synthesized to fish out methionine-containing proteins based on a non-enzymatic biochemical reaction, which couples amino groups of lysine with the S-methyl group of methionine in the presence of HOBr. Peptides which contain four lysine residues (Lys-Lys-Lys-Lys-{Se-Cys}) linked to the Fe3O4@Au nanocomposites were used to capture methionine residues efficiently via a S═N cross-linking. The methionine-containing protein was obtained by magnetic separation and released from the Fe3O4@Au-Se-peptide nanocomposites with the influence of H2Se. The HRMS and SDS-PAGE results confirmed the methionine-containing protein could be successfully fished out from a mixture solution. This work provides a useful strategy for recognition and separation of a category of proteins from complex biological systems.

Monitoring NAD(P)H by an ultrasensitive fluorescent probe to reveal reductive stress induced by natural antioxidants in HepG2 cells under hypoxia.

Reductive stress, the opposite of oxidative stress, represents a disorder in the redox balance state which is harmful to biological systems. For decades, the role of oxidative stress in tumor therapy has been the focus of attention, while the effects of reductive stress have been rarely studied. Here, we report the anti-cancer effects of reductive stress induced by three natural antioxidants (resveratrol, curcumin and celastrol). Considering the fact that the solid tumor microenvironment suffers from hypoxia, we performed cell experiments under hypoxic conditions. In order to observe the reductive stress, we first developed an ultrasensitive fluorescent probe (TCF-MQ) for specifically imaging NAD(P)H which is a marker of reductive stress. TCF-MQ responded to NAD(P)H rapidly and exhibited high sensitivity with a detection limit of 6 nM. With the help of TCF-MQ, we found that upon the treatment of HepG2 cells with pharmacological doses of three natural antioxidants under hypoxic conditions, high levels of NAD(P)H were produced before cell death. The excess NAD(P)H resulted in reductive stress instead of oxidative stress. In contrast, under normoxic conditions, there was no reductive stress involved in the process of cell death induced by three natural antioxidants. Therefore, we hypothesize that the mechanism of cancer cell death induced by natural antioxidants under hypoxia should be attributed to the reductive stress.

Cyclic Regulation of the Sulfilimine Bond in Peptides and NC1 Hexamers via the HOBr/H2Se Conjugated System.

The sulfilimine bond (-S═N-), found in the collagen IV scaffold, significantly stabilizes the architecture via the formation of sulfilimine cross-links. However, precisely governing the formation and breakup process of the sulfilimine bond in living organisms for better life functions still remains a challenge. Hence, we established a new way to regulate the breaking and formation of the sulfilimine bond through hydrogen selenide (H2Se) and hypobromous acid (HOBr), which can be easily controlled at simulated physiological conditions. This novel strategy provides a circulation regulation system to modulate the sulfilimine bond in peptides and NC1 hexamers, which can offer a substantial system for further study of the physiological function of collagen IV.

High-Quantum-Yield Mitochondria-Targeting Near-Infrared Fluorescent Probe for Imaging Native Hypobromous Acid in Living Cells and in Vivo.

The discovery that hypobromous acid (HOBr) can regulate the activity of collagen IV has attracted great attention. However, HOBr as an important reactive small molecule has hardly ever been studied using a detection method suitable for organisms. Herein, a high-quantum-yield mitochondria-targeting near-infrared (NIR) fluorescent probe for HOBr, RhSN-mito, was designed. RhSN-mito was easily obtained by the Suzuki cross-coupling reaction. The test results show that RhSN-mito can rapidly respond to HOBr with ultrasensitivity and high selectivity. The achievement of ultrasensitivity lies in the high signal-to-noise ratio and the highest fluorescence quantum yield of the reaction product (ΦF = 0.68) in the near-infrared region, as far as we know. RhSN-mito is successfully applied to image native HOBr in mitochondria of HepG2 cells and zebrafish. Thus, RhSN-mito is a powerful tool for detecting native HOBr in vivo and is expected to provide a method to further study the physiological and pathological functions related to HOBr.