Fluorescence lifetime imaging: A novel approach to simultaneous visualizing lipid droplets and endoplasmic reticulum polarity in live cells and fatty liver

Cellular polarity plays an indispensable role in both physiological functions and the progression of diseases, with organelles demonstrating specific polarities that are integral to their roles. The advent of a single fluorescent probe capable of concurrently and selectively visualizing two distinct organelles through distinct fluorescence emission spectra has emerged as a powerful approach for investigating their interactions within cellular mechanisms. However, the use of changes in dual fluorescence lifetimes to monitor polarity changes in two organelles has rarely been employed, despite its potential benefits for more accurate detection of subtle changes in organelle polarity. Herein, two polarity-sensitive probes (ER-LDs-1 and ER-LDs-2) were developed. ER-LDs-2 exhibited high sensitivity to polarity alterations in both lipid droplets (LDs) and endoplasmic reticulum (ER) under various physiological and pathological conditions, including ferroptosis and endoplasmic reticulum stress, as demonstrated by distinct changes in fluorescence lifetime. ER-LDs-2 enables the monitoring of polarity changes in mouse and human normal and fatty liver tissues by fluorescence lifetime imaging. Collectively, this study presents innovative tools that hold great potential for advancing our understanding of cellular organelle polarity dynamics and the progression of associated diseases. •Two polarity-sensitive fluorescent probes are developed.•ER-LDs-2 enables simultaneous fluorescence lifetime imaging polarity changes in lipid droplets and endoplasmic reticulum.•ER-LDs-2 can image two-organelles polarity changes during ferroptosis and endoplasmic reticulum stress.•ER-LDs-2 enables fluorescence lifetime imaging polarity changes in mouse and human normal and fatty liver tissues.