Impact of microbubble-to-cell parameters on heterogeneous sonoporation at the single-cell level

The aim of this investigation is to understand how microbubble-to-cell relative parameters lead to heterogensous sonoporation. Using a customized platform, we observed sonoporation dynamics at the single-cell level and quantified exogenous molecular uptake levels to evaluate the degree of sonoporation. Sonovue microbubbles at different concentrations were introduced to passively regulate the bubble-to-cell parameters, i.e. bubble-to-cell distance and numbers. 1 MHz ultrasound with 10-cycle duration and 0.6 MPa peak negative pressure were applied to trigger the inertial collapse of microbubbles. Our data revealed the impact of bubble-to-cell parameters on the heterogeneity of sonoporation when bubble diameter (D) was less than 5.5µm. First, sonoporation was influenced by the distance (d) between the bubble and the cell; no sonoporation (type I) was observed when d/D > 1, while reversible sonoporation (type II) occurred by the localized collapse of the microbubble when d/D < 1. Second, the number of bubbles is positively correlated with the degree of sonoporation when d/D < 1. Localized collapse of two to three bubbles resulted in type II sonoporation, while irreversible sonoporation (type III) was found after the collapse of four or more bubbles. Our study suggests that bubble-to-cell parameters is essential for predictable sonoporation, and provides a possible method to optimize the use of sonoporation in drug delivery.