Three-Dimensional Quantitative Co-Mapping of Pulmonary Morphology and Nanoparticle Distribution with Cellular Resolution in Nondissected Murine Lungs

Deciphering
biodistribution, biokinetics, and biological effects
of nanoparticles (NPs) in entire organs with cellular resolution remains
largely elusive due to the lack of effective imaging tools. Here,
light sheet fluorescence microscopy in combination with optical tissue
clearing was validated for concomitant three-dimensional mapping of
lung morphology and NP biodistribution with cellular resolution in
nondissected ex vivo murine lungs. Tissue autofluorescence
allowed for label-free, quantitative morphometry of the entire bronchial
tree, acinar structure, and blood vessels. Co-registration of fluorescent
NPs with lung morphology revealed significant differences in pulmonary
NP distribution depending on the means of application (intratracheal
instillation and ventilator-assisted aerosol inhalation under anesthetized
conditions). Inhalation exhibited a more homogeneous NP distribution
in conducting airways and acini indicated by a central-to-peripheral
(C/P) NP deposition ratio of unity (0.98 ± 0.13) as compared
to a 2-fold enhanced central deposition (C/P = 1.98 ± 0.37) for
instillation. After inhalation most NPs were observed in the proximal
part of the acini as predicted by computational fluid dynamics simulations.
At cellular resolution patchy NP deposition was visualized in bronchioles
and acini, but more pronounced for instillation. Excellent linearity
of the fluorescence intensity–dose response curve allowed for
accurate NP dosimetry and revealed ca. 5% of the
inhaled aerosol was deposited in the lungs. This single-modality imaging
technique allows for quantitative co-registration of tissue architecture
and NP biodistribution, which could accelerate elucidation of NP biokinetics
and bioactivity within intact tissues, facilitating both nanotoxicology
studies and the development of nanomedicines.