Determining Ultrasound Parameters for Bursting Polymer Microbubbles for Future Use in Spinal Cord Injury

Abstract

Objective: We believe our poly(lactic acid) (PLA) microbubbles are well suited for therapeutic delivery to spinal cord injury (SCI) using ultrasound-triggered bursting. We investigated the feasibility of clinical ultrasound bursting in situ, the optimal bursting parameters in vitro and the loading and release of a model bio-active DNA.

Methods: Microbubbles were tested using clinical ultrasound in a rat cadaver SCI model. Burst pressure thresholds were determined using the change in enhancement after ultrasound exposure. Resonance frequency, acoustic enhancement, sizing and morphology were evaluated by comparing two microbubble porogens, ammonium carbonate and ammonium carbamate. Oligonucleotides were loaded into the shell and released using the found optimized ultrasound bursting parameters.

Results: In situ imaging and bursting were successful. In vitro bursting thresholds using frequencies 1, 2.25 and 5 MHz were identified between peak negative pressures 0.2 and 0.5 MPa, believed to be safe for spinal cord. The pressure threshold decreased with decreasing frequencies. PLA bursting was optimized near the resonance frequency of 2.5 to 3.0 MHz using 2.25 MHz and not at lower frequencies. PLA microbubbles, initially with a mean size of approximately 2 µm, remained in one piece, collapsed to between 0.5 and 1 µm and did not fragment. Significantly more oligonucleotide was released after ultrasound bursting of loaded microbubbles. Microbubble-sized debris was detected when using ammonium carbamate, leading to inaccurate microbubble concentration measurements.

Conclusion: PLA microbubbles made with ammonium carbonate and burst at appropriate parameters have the potential to safely improve intrathecal therapeutic delivery to SCI using targeted ultrasound.

Keywords: Bursting; Drug delivery; Microbubble; Spinal cord injury; Ultrasound.