Blue-emitting lipid droplet probes based on coumarin dye for multi-color imaging of living cells and fatty livers of mice

Lipid droplets (LDs) are highly dynamic organelles in cells and are closely related not only to intracellular lipid metabolic processes but also to various lipid metabolism disorders such as fatty liver disease, obesity, and diabetes. Since these processes are deeply associated with the dynamic behaviors between LDs and other organelles in cells, their simultaneous imaging provides valuable information on intracellular processes. Herein, we developed blue-emitting LD-specific probes (DBC30, DBC30-Bu, and DBC30-Ph) based on coumarin dye, and performed multi-color imaging of living cells and hepatic tissues of living mice using a confocal fluorescence microscope. DBC30 derivatives exhibited large molar absorption coefficients and high fluorescence quantum yields in non-polar and polar solvents. The confocal fluorescence microscopy measurements of HeLa cells pretreated with oleic acid followed by staining with DBC30 derivatives revealed that these probes selectivity stain LDs and showed blue fluorescence in LDs, allowing multi-color imaging using green and red fluorescence organelle-specific probes. Additionally, DBC30 derivatives exhibited high photostability compared with a commercially available blue-emitting LD-specific probe. We took dual-color images of the surfaces of livers following intravenous administration of DBC30 and FITC-lectin (a green fluorescent vascular endothelial marker) into normal and high-fat-diet-fed mice. Simultaneous imaging of hepatic tissues of fatty liver mice using these probes allowed the visualization of small and large LDs in hepatocytes and of the distorted sinusoidal network. Furthermore, co-staining experiments using DBC30 and the green-emitting nucleus-specific probe enabled visualization of the distribution of LDs and nuclei in hepatic tissue sections of normal and fatty livers. These results demonstrated that DBC30 is advantageous as an imaging probe for LDs in living cells and in vivo and ex vivo hepatic tissues.