Cavitation bubble nuclei in healthy versus cancerous 3-D cell cultures

The safety of biomedical ultrasound depends on controlling cavitation bubbles in vivo, yet the spatial location of bubble nuclei in tissues (intracellular versus extracellular) and their behavior in healthy versus cancerous cellular environments remain unexplored. Here we evaluated the spatial location of bubble nuclei and cavitation thresholds in healthy (L6) and cancerous (L8) rat musculoskeletal myoblasts (n = 5 each), cultured in 3-D using PureCol® EZ-Gel scaffolds. A 3.68-MHz focused ultrasound transducer (f# = 1) induced low-density cavitation with pulse durations (PD) of 10–100 μs at 1 Hz PRF and pressures up to p- = 13 MPa. Brightfield microscopy and high-speed photography (20 000 fps) monitored bubble location. Cavitation bubbles (r ≈ 5 μm) appeared stochastically and exclusively in the extracellular space of the healthy and cancerous 3-D cultures despite significant radiation force. In cancerous L8 cells, no cavitation occurred at PD = 50 μs with pressures up to p- = 13 MPa over 15 pulses; however, PD = 100 μs induced extracellular cavitation at p ≈ 12.3 MPa. In contrast, extracellular cavitation was detected in healthy L6 cultures at PD = 50 μs at p ≈ 11.8 MPa, suggesting the potential influence of cell stiffness on cavitation thresholds. Future work will computationally investigate the effects of scaffold and tissue stiffness on bubble formation. [Work supported by NSF CAREER 1943937.]