Ultrastructural Analysis of Volumetric Histotripsy Bio-effects in Large Human Hematomas

Large-volume soft tissue hematomas are a serious clinical problem, which, if untreated, can have severe consequences. Current treatments are associated with significant pain and discomfort. It has been reported that in an in vitro bovine hematoma model, pulsed high-intensity focused ultrasound (HIFU) ablation, termed histotripsy, can be used to rapidly and non-invasively liquefy the hematoma through localized bubble activity, enabling fine-needle aspiration. The goals of this study were to evaluate the efficiency and speed of volumetric histotripsy liquefaction using a large in vitro human hematoma model. Large human hematoma phantoms (85 cc) were formed by recalcifying blood anticoagulated with citrate phosphate dextrose/saline-adenine-glucose-mannitol solution. Typical boiling histotripsy pulses (10 or 2 ms) or hybrid histotripsy pulses using higher-amplitude and shorter pulses (0.4 ms) were delivered at 1% duty cycle while continuously translating the HIFU focus location. Histotripsy exposures were performed under ultrasound guidance with a 1.5-MHz transducer (8-cm aperture, F# = 0.75). The volume of liquefied lesions was determined by ultrasound imaging and gross inspection. Untreated hematoma samples and samples of the liquefied lesions aspirated using a fine needle were analyzed cytologically and ultrastructurally with scanning electron microscopy. All exposures resulted in uniform liquid-filled voids with sharp edges; liquefaction speed was higher for exposures with shorter pulses and higher shock amplitudes at the focus (up to 0.32, 0.68 and 2.62 mL/min for 10-, 2- and 0.4-ms pulses, respectively). Cytological and ultrastructural observations revealed completely homogenized blood cells and fibrin fragments in the lysate. Most of the fibrin fragments were less than 20 μm in length, but a number of fragments were up to 150 μm. The lysate with residual debris of that size would potentially be amenable to fine-needle aspiration without risk for needle clogging in clinical implementation.