Visualizing liver anatomy, physiology and pharmacology using multiphoton microscopy

Multiphoton microscopy (MPM) has become increasingly popular and widely used in both basic and clinical liver studies over the past few years. This technology provides insights into deep live tissues with less photobleaching and phototoxicity, which helps us to better understand the cellular morphology, microenvironment, immune responses and spatiotemporal dynamics of drugs and therapeutic cells in the healthy and diseased liver. This review summarizes the principles, opportunities, applications and limitations of MPM in hepatology. A key emphasis is on the use of fluorescence lifetime imaging (FLIM) to add additional quantification and specificity to the detection of endogenous fluorescent species in the liver as well as exogenous molecules and nanoparticles that are applied to the liver in vivo. We anticipate that in the near future MPM‐FLIM will advance our understanding of the cellular and molecular mechanisms of liver diseases, and will be evaluated from bench to bedside, leading to real‐time histology of human liver diseases. Multiphoton microscopy is a new tool which provides insights into deep live tissues to understand the cellular morphology, microenvironment, immune responses and spatiotemporal dynamics of drugs and therapeutic cells in the liver. This review summarizes its principles, opportunities, applications and limitations in hepatology. A key emphasis is on the use of fluorescence lifetime imaging to add additional quantification and specificity to the detection of fluorescent species in the liver.