A D-π-A-type ratiometric fluorescent probe to detect polarity changes and inhibition effect during ferroptosis.

To thoroughly understand ferroptosis's biological functions in living cells, it is crucial to investigate the polarity variations that occur during this unique Fe(II)-facilitated oxidative type of cell death. In this work, we report the development of a ratiometric probe (Po-P) to visualize the polarity changes in living cells and the inhibition effect during ferroptosis. The polarity-responsive fluorophore utilized by Po-P has a D-π-A-type structure. Based on theoretical calculations, ICT was proposed as the basis for Po-P's polarity-responsive mechanism. According to cell imaging results, Po-P had a desirable capacity for monitoring polarity fluctuations and erastin-induced ferroptosis. Furthermore, inhibition imaging revealed that dihydrolipoic acid (DHLA) could potentially prevent polarity changes that occur during erastin-induced ferroptosis, just as vitamin E (VE). We anticipate that the probe Po-P could be a valuable tool to quickly monitor polarity fluctuations and inhibition effects during ferroptosis and create new medications for treating disorders related to ferroptosis.

Development of bishydrazide-based fluorescent probes for the imaging of cellular peroxynitrite (ONOO-) during ferroptosis.

Peroxynitrite (ONOO-) is a critical ROS in living systems, and could induce lipid peroxidation which is the driver of ferroptotic cell death. Therefore, precise and rapid detection of cellular ONOO- is critical for the deep study of the biological functions of ONOO- during ferroptosis. Herein, we developed fluorescent probes (Rh-1, Rh-2 and Rh-3) for the rapid detection of intracellular ONOO- during ferroptosis. These probes used bishydrazide groups as the reactive sites for ONOO-. The response of these probes to ONOO- resulted in the production of the emissive xanthene fluorophore, providing a marked enhancement in the fluorescence intensity at 561 nm. The probe Rh-3 exhibited prominent selectivity and sensitivity towards ONOO-. Bioimaging experiments suggested that Rh-3 could be applied to image exogenous and endogenous ONOO- in living cells. By fluorescence imaging, it was demonstrated that erastin-induced ferroptosis caused increased levels of the endogenous ONOO-, and ferrostatin-1 (Fer-1) and vitamin E (VE) could markedly inhibit the excessive production of ONOO- during ferroptosis in living cells.

The development of an endoplasmic reticulum-targeting fluorescent probe for the imaging of 1,4-dithiothreitol (DTT) in living cells.

1,4-Dithiothreitol (DTT) is a robust reducing agent that contributes significantly to the folding process of proteins and maintaining endoplasmic reticulum (ER) homeostasis. Abnormally high levels of DTT can lead to severe endoplasmic reticulum stress (ERS), which induces cell death. In addition, DTT can also hinder cell growth and enhance reactive oxygen species (ROS) production in the ER. Herein, an effective turn-on ER-targeting fluorescent probe, ER-DTT, was designed to image DTT for the first time. The probe ER-DTT was based upon naphthalimide as a fluorophore, p-toluenesulfonamide as an exceptional unit for ER-targeting, and sulfoxide as a response site for imaging DTT based on an intramolecular charge transfer (ICT) mechanism. Optical-response experiments showed that the probe ER-DTT had good selectivity and sensitivity for DTT. Furthermore, confocal microscopy indicated that ER-DTT was suitable for selectively targeting ER in living cells and could be implemented to recognize cellular DTT.

A sensitive and selective fluorescent probe for the detection of endogenous peroxynitrite (ONOO-) in living cells.

Peroxynitrite (ONOO-) is one of reactive oxygen species, and plays a vital role in numeorus physiological and pathological processes. Given that the ONOO- level is closely related with various serious diseases, the in situ and real time detection of endogenous ONOO- is highly important for the in-depth study of its roles in living systems. Herein, we present a new fluorescent probe (RHPN) for the real-time detection of intracellular ONOO-. The probe RHPN consists of a rhodamine analogue and an arylhydrazide group as a response site for ONOO-. In response to ONOO-, the probe RHPN converts to an open-ring form and generates strong fluorescence. Moreover, the probe RHPN was successfully used for the imaging of the endogenous and exogenous ONOO- level changes in living cells.

An ESIPT-based ratiometric fluorescent probe for the discrimination of live and dead cells.

Cell death can destroy homeostasis and is a hallmark of many pathological conditions. Discrimination of live and dead cells is a crucial task for the biological, medical and pharmaceutical studies. Herein, we constructed an ESIPT-based fluorescent probe (BTE) on the basis of the different esterase activity in live and dead cells. Under excitation, the probe BTE showed the blue emission peaked at 465 nm, while it mainly displayed green emission peaked at 543 nm after it was hydrolyzed by esterase. Imaging of the cells treated by H2O2 and ultraviolet (UV) radiation demonstrated that the probe BTE is effective in the detection of the health of cells, could help us to better understand cell death and its effects in a range of diseases and treatments.

An ultrasensitive ratiometric fluorescent probe based on the ICT-PET-FRET mechanism for the quantitative measurement of pH values in the endoplasmic reticulum (ER).

Herein, we present a dual-site ratiometric fluorescent probe based on the ICT-PET-FRET mechanism for the detection of pH in the ER. The probe showed a highly sensitive response to pH in the range of 5.0-7.2, and could be applied for the quantitative measurement of the pH values in the ER during ER stress and dexamethanose-induced stimulation.