Targeted Lifetime Probe Enables Quantitative FLIM Mapping of Lipid Droplet Polarity Remodeling under Copper-Induced Oxidative Stress.

Lipid droplet (LD) microenvironmental remodeling accompanies oxidative stress, mitochondrial dysfunction, and metal-induced metabolic perturbation, yet remains difficult to quantify in living systems. Here, we report LDP-Pol, a BODIPY-derived, LD-targeted fluorescence lifetime imaging microscopy (FLIM) probe for mapping LD polarity heterogeneity. LDP-Pol combines an ICT-active triphenylamine-BODIPY scaffold with a C12 methyl ester-containing chain inspired by endogenous lipid components, improving compatibility with neutral lipid-rich environments and promoting selective LD partitioning for polarity-dependent lifetime readout. Solvent and lipid-mimetic studies established a calibrated lifetime response to polarity. In living cells, LDP-Pol resolved LD polarity-associated microenvironmental changes during H2O2-induced oxidative stress and CuCl2/elesclomol-induced copper stress, while dual-color super-resolution imaging confirmed its compatibility with live-cell organelle tracking and LD-mitochondria contact analysis. In tumor models, LDP-Pol enabled tissue-level FLIM visualization of heterogeneous lipid polarity landscapes under copper-induced stress. These results establish a lifetime-based strategy for probing LD microenvironmental remodeling associated with oxidative and metal-induced cellular perturbation.