Detecting labile heme and ferroptosis through 'turn-on' fluorescence and lipid droplet localization post Fe2+ sensing.

Iron, a crucial biologically active ion essential for metabolic processes in living organisms, plays a vital role in biological functions, and imbalances in iron levels can lead to various diseases. In this study, we have developed two simple "turn-on" fluorescent probes, NOPy and NOCN, for the quick and selective detection of Fe2+ at nanomolar levels (LOD of 35 nM), accompanied by significant absorption and emission shifts, along with colorimetric demarcation. Both fluorophores exhibit an excellent "turn-on" emission response upon encountering Fe2+ in the cells. Flow cytometry and confocal fluorescence imaging studies demonstrate enhanced fluorescence signals in response to labile iron, efficiently detecting heme during erastin-induced ferroptosis. Interestingly, we also observed that the product formed after Fe2+ sensing localizes within the lipid droplets. These water-soluble and highly sensitive reactive probes, NOPy and NOCN, enable investigations of iron-dependent physiological and pathological conditions. The development of these probes represents an advancement in the field, offering a rapid and selective means for detecting Fe2+ with minimal cytotoxicity.

A multipurpose mitochondrial NIR probe for imaging ferroptosis and mitophagy.

This paper explores the use of a di-cationic fluorophore for visualizing mitochondria in live cells independent of membrane potential. Through the synthesized di-cationic fluorophore, we investigate the monitoring of viscosity, ferroptosis, stress-induced mitophagy, and lysosomal uptake of damaged mitochondria. The designed fluorophore is based on DQAsomes, cationic vesicles responsible for transporting drugs and DNA to mitochondria. The symmetric fluorophores possess two charge centres separated by an alkyl chain and are distinguished by a pyridinium group for mitochondrial selectivity, the C-12 alkyl substitution for membrane affinity, and an electron donor-π-acceptor fluorescent scaffold for intramolecular charge transfer. The synthesized fluorophores, PP and NP, emit wavelengths exceeding 600 nm, with a significant Stokes shift (130-211 nm), and NP demonstrates near-infrared emission (∼690 nm). Our study underscores the potential of these fluorophores for live-cell imaging, examining physiological responses such as viscosity and ferroptosis, and highlights their utility in investigating mitophagy damage and lysosomal uptake.

A two-in-one probe: imaging lipid droplets and endoplasmic reticulum in tandem.

The endoplasmic reticulum (ER) and lipid droplets (LDs) intricately interact in cellular processes, with the ER serving as a hub for lipid synthesis and LDs acting as storage organelles for lipids. Developing fluorescent probes that can simultaneously visualise the ER and LDs provides a means for real-time and specific visualisation of these subcellular organelles and elucidating their interaction. Herein, we present synthetically simple and novel donor-π-acceptor styryl fluorophores (PFC, PFN and PFB) incorporating pentafluorophenyl (PFP) to demonstrate exquisite discriminative imaging of ER and LD with a single excitation wavelength. The PFP moiety aids the ER selectivity, while the overall hydrophobicity of the molecule aids in the LD targeting. Furthermore, the fluorophores are utilised in studying the changes in size, distribution, and biogenesis of LDs within ER regions after treatment with oleic acid. Strong emission, lower concentrations ∼100 nM requirement, minimal cytotoxicity, and photostability make these fluorophores excellent tools for probing sub-cellular dynamics.

Fluorescent sterol probes for intracellular transport, imaging, and therapeutics.

Sterols play a significant role in many physiological processes affecting membrane organization, transport, permeability, and signal transduction. The development of fluorescent sterol analogs that have immediate functional relevance to the natural biomolecules is one approach to understanding the sterol-driven physiological processes. Visualizing cellular compartments with tailor-made fluorescent molecules through specific labeling methods enables organelle targeting and reveals dynamic information. In this review, we focus on the recent literature published between 2020 and 2022, with particular emphasis on extrinsic fluorophores and their investigations of sterol-driven biological processes involving sterol transport, biomolecular interactions, and biological imaging.