Development of an In Vitro Model to Study Mechanisms of Ultrasound-Targeted Microbubble Cavitation-Mediated Blood-Brain Barrier Opening.

OBJECTIVE: Ultrasound-targeted microbubble cavitation (UTMC)-mediated blood-brain barrier (BBB) opening is being explored as a method to increase drug delivery to the brain. This strategy has progressed to clinical trials for various neurological disorders, but the underlying cellular mechanisms are incompletely understood. In the study described here, a contact co-culture transwell model of the BBB was developed that can be used to determine the signaling cascade leading to increased BBB permeability. METHODS: This BBB model consists of bEnd.3 cells and C8-D1A astrocytes seeded on opposite sides of a transwell membrane. Pulsed ultrasound (US) is applied to lipid microbubbles (MBs), and the change in barrier permeability is measured via transendothelial electrical resistance and dextran flux. Live cell calcium imaging (Fluo-4 AM) is performed during UTMC treatment. RESULTS: This model exhibits important features of the BBB, including endothelial tight junctions, and is more restrictive than the endothelial cell (EC) monolayer alone. When US is applied to MBs in contact with the ECs, BBB permeability increases in this model by two mechanisms: UTMC induces pore formation in the EC membrane (sonoporation), leading to increased transcellular permeability, and UTMC causes formation of reversible inter-endothelial gaps, which increases paracellular permeability. Additionally, this study determines that calcium influx into ECs mediates the increase in BBB permeability after UTMC in this model. CONCLUSION: Both transcellular and paracellular permeability can be used to increase drug delivery to the brain. Future studies can use this model to determine how UTMC-induced calcium-mediated signaling increases BBB permeability.

Adaptable Xerogel-Layered Amperometric Biosensor Platforms on Wire Electrodes for Clinically Relevant Measurements.

Biosensing strategies that employ readily adaptable materials for different analytes, can be miniaturized into needle electrode form, and function in bodily fluids represent a significant step toward the development of clinically relevant in vitro and in vivo sensors. In this work, a general scheme for 1st generation amperometric biosensors involving layer-by-layer electrode modification with enzyme-doped xerogels, electrochemically-deposited polymer, and polyurethane semi-permeable membranes is shown to achieve these goals. With minor modifications to these materials, sensors representing potential point-of-care medical tools are demonstrated to be sensitive and selective for a number of conditions. The potential for bedside measurements or continuous monitoring of analytes may offer faster and more accurate clinical diagnoses for diseases such as diabetes (glucose), preeclampsia (uric acid), galactosemia (galactose), xanthinuria (xanthine), and sepsis (lactate). For the specific diagnostic application, the sensing schemes have been miniaturized to wire electrodes and/or demonstrated as functional in synthetic urine or blood serum. Signal enhancement through the incorporation of platinum nanoparticle film in the scheme offers additional design control within the sensing scheme. The presented sensing strategy has the potential to be applied to any disease that has a related biomolecule and corresponding oxidase enzyme and represents rare, adaptable, sensing capabilities.