Theranostic USPIO-Loaded Microbubbles for Mediating and Monitoring Blood-Brain Barrier Permeation

Abstract

Efficient and safe drug delivery across the blood-brain barrier (BBB) remains to be one of the major challenges of biomedical and (nano-) pharmaceutical research. Here, we show that poly(butyl cyanoacrylate)-based microbubbles (MB), carrying ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles within their shell, can be used to mediate and monitor BBB permeation. Upon exposure to transcranial ultrasound pulses, USPIO-MB are destroyed, resulting in acoustic forces inducing vessel permeability. At the same time, USPIO are released from the MB shell, they extravasate across the permeabilized BBB and they accumulate in extravascular brain tissue, thereby providing non-invasive R₂*-based magnetic resonance imaging information on the extent of BBB opening. Quantitative changes in R₂* relaxometry were in good agreement with 2D and 3D microscopy results on the extravascular deposition of the macromolecular model drug FITC-dextran into the brain. Such theranostic materials and methods are considered to be useful for mediating and monitoring drug delivery across the BBB, and for enabling safe and efficient treatment of CNS disorders.

Keywords: Drug Delivery; Hybrid Materials; Magnetic Nanoparticles; Medical Applications; Stimuli-Responsive Materials.

Figure 1

Schematic experimental setup for mediating and monitoring drug delivery across the BBB using USPIO-loaded MB. After an MRI pre-scan (A), USPIO-MB and 70 kDa FITC-dextran were co-injected (B). Then, the USPIO-MB were destroyed using US, resulting in the generation of acoustic forces, USPIO release from the MB, permeation of the BBB, and USPIO extravasation (C). After an MRI post scan, to visualize and quantify USPIO deposition in the brain (D), rhodamine-lectin was injected and FITC-dextran extravasation and penetration was assessed using 2D and 3D microscopy (E).

Figure 2

Synthesis and characterization of USPIO-loaded MB. A: Schematic synthetic protocol. B: Size and size distribution of USPIO-loaded and regular MB. C-H: SEM and TEM images of intact (C,E,G) and destroyed (D,F,H) USPIO-MB, exemplifying an average diameter of ~2.5 μm, a shell thickness of ~50 nm, efficient USPIO incorporation into the shell, and (partial) USPIO release upon US-mediated MB destruction. See Table S1 for more details.

Figure 3

Mediating and monitoring BBB permeation using USPIO-MB. A: Power Doppler US images before and directly after the i.v. infusion of USPIO-MB. B: MR imaging of USPIO deposition across the BBB upon US-induced USPIO-MB destruction. Color-coded R₂*-maps are overlaid on morphological T₂*-weighted images, which were recorded before and after exposure to US. C: Quantification of the increase in R₂* values, presented as percentage ± SEM. * indicates p<0.0001 vs. USPIO-MB alone and US alone. D: Prussian Blue staining of brain tissue, confirming the deposition of USPIO nanoparticles (arrows) across the BBB and within the CNS.

Figure 4

FITC-dextran extravasation and penetration. A: 2D fluorescence (2D-FM) and 3D two-photon microscopy (3D-2PM) of FITC-dextran (green) extravasation across rhodamin lectin-stained (red) blood vessels, showing efficient macromolecular (model) drug delivery across the BBB upon the combination of USPIO-MB with 5 and 30 min of US. B: Extravasation and penetration were quantified using a procedure in which 1: vessels were segmented by rhodamine-lectin-thresholding, 2: a distance map was calculated, 3: concentric rings were drawn around the vessels, and 4: the signal intensity of FITC-dextran was measured within each ring. C: Signal intensity of extravasated FITC-dextran as a function of distance to vessel surface. Values represent the mean signal intensity within each concentric rings ± SEM. * and # indicate p<0.005 vs. UPSIO-MB alone and US alone.