Intravital multiphoton microscopy as a tool to investigate disposition of small molecules, nanoparticles and cells

Intravital multiphoton microscopy is a high resolution and deep penetration fluorescence light microscopy technique for studying physiological and pathological processes in live animals. It enables cellular and subcellular structures to be dynamically imaged over time and for the in vivo disposition of fluorescent compounds to be characterised. We applied multiphoton microscopy coupled with fluorescence lifetime imaging technique to quantitatively evaluate the function of hepatic transporters using transporter specific substrate. Calibrated fluorescence intensity time curves measured in sinusoids and hepatocytes separately together with cumulative biliary excretion data were analysed by a three compartment model to give a robust parameter estimation of transporter function. We also modelled uptake and excretion of fluorescein in normal and diseased rat liver. Fluorescein associated fluorescence were found to be increased in steatotic liver, which was attributed to increased hepatic fluorescein metabolism. We directly imaged the disposition of quantum dots (~3.5 nm) in rat liver and kidney after intravenous injection. We found that negatively charged quantum dots were initially retained in the sinusoids in liver or peritubular capillaries or glomerular arterioles in kidney after injection. They were not taken up by hepatocytes or tubular cells, while were selectively taken up by sinusoidal cells or by mesangial cells. Based on these observation, we then build a mechanistic and physiologically based pharmacokinetic model for accurately characterizing and predicting the in vivo fate of this type of nanoparticles. We also extend the application of intravital imaging technique to characterize the physiological kinetics of administered mesenchymal stem cells (MSCs) by direct visualization of cell spatiotemporal disposition of green-fluorescence-protein labelled cells in different organs of mice. We observed significant trapping of cells in the lung and cell arrest, depletion in the liver. A physiologically based kinetic model was also developed for intravenously administered MSCs based on these observations.