Visualizing treatment delivery and deposition in mouse lungs using in vivo x-ray imaging

Abstract

The complexity of lung diseases makes pre-clinical in vivo respiratory research in mouse lungs of great importance for a better understanding of physiology and therapeutic effects. Synchrotron-based imaging has been successfully applied to lung research studies, however longitudinal studies can be difficult to perform due to limited facility access. Laboratory-based x-ray sources, such as inverse Compton x-ray sources, remove this access limitation and open up new possibilities for pre-clinical small-animal lung research at high spatial and temporal resolution. The in vivo visualization of drug deposition in mouse lungs is of interest, particularly in longitudinal research, because the therapeutic outcome is not only dependent on the delivered dose of the drug, but also on the spatial distribution of the drug. An additional advantage of this approach, when compared to other imaging techniques, is that anatomic and dynamic information is collected simultaneously. Here we report the use of dynamic x-ray phase-contrast imaging to observe pulmonary drug delivery via liquid instillation, and by inhalation of micro-droplets. Different liquid volumes (4 l  , 20 l  , 50 l ) were tested and a range of localized and global distributions were observed with a temporal resolution of up to 1.5 fps. The in vivo imaging results were confirmed by ex vivo x-ray and fluorescence imaging. This ability to visualize pulmonary substance deposition in live small animals has provided a better understanding of the two key methods of delivery; instillation and nebulization.

Multimodal Precision Imaging of Pulmonary Nanoparticle Delivery in Mice: Dynamics of Application, Spatial Distribution, and Dosimetry

“…PB‐PCXI utilizes differences in X‐ray refractive index and associated phase shifts at air–tissue interfaces for enhancing soft tissue contrast (e.g., in the lung) as compared to X‐ray absorption . Building on our previously published in vivo PB‐PCXI studies in murine lungs, the present study demonstrates that combined in vivo PB‐PCXI and ex vivo fluorescence imaging can provide complementary information, which enhances the understanding of complex processes such as NM/NP delivery to the lung in a quantitative way on various scales (whole lung to cellular). The strengths and weaknesses of all five imaging modalities used here are summarized with respect to 2D/3D imaging capability, resolution, fidelity (conservation of original anatomical 3D structure), anatomical information, and technical complexity as summarized in Table .…”

“…It is not intuitively clear how a bulk liquid injected into the trachea via intratracheal instillation can reach the deeper parts of the lung. In vivo PB‐PCXI revealed that the instilled bulk liquid is not primarily flowing down the bronchial tree, but is distributed quite uniformly throughout the lung by secondary aerosol formation due to occasional blockage of the airways with liquid and subsequent bursting of this blockage during breathing activity (Videos S1–S3, Supporting Information) . As mentioned above, the slow‐instillation process (24 s for 100 µL; 4.2 µL s −1 ), which was used here for the sake of in vivo PB‐PCXI imaging, is not identical to the typically used, rapid manual instillation (<1 s, >50 µL s −1 ).…”

“…The intratracheal instillation and X‐ray imaging setup is depicted in Figure a. As described by Gradl et al, mice were fixed in an upright position, mechanically ventilated, and an iodine‐polystyrene NP suspension was instilled slowly into the lungs using a remotely controlled syringe pump. This permits delivery of a known amount of liquid at a controllable and constant rate (here 4.2 µL s −1 , i.e., 100 µL within 24 s).…”

“…This permits delivery of a known amount of liquid at a controllable and constant rate (here 4.2 µL s −1 , i.e., 100 µL within 24 s). Triggered by the ventilator, one image per breath was captured during an end‐inspiratory breath‐hold phase, which minimizes inflation/deflation‐induced blurring effects . This allowed for time‐resolved visualization of the pulmonary NP delivery process.…”

“…For weakly absorbing materials such as soft tissue and air, the difference in the phase shift is significant, which leads to contrast enhancement. Propagation‐based phase‐contrast X‐ray imaging (PB‐PCXI) was chosen here since this technique requires only a single exposure (compared to, e.g., grating‐based phase‐contrast imaging), so it is ideal for imaging dynamic processes . Furthermore, it is a simple technique, because the only change with respect to conventional absorption imaging is an increased sample‐to‐detector distance and increased source coherence.…”