Optimization of low-frequency low-intensity ultrasound-mediated microvessel disruption on prostate cancer xenografts in nude mice using an orthogonal experimental design.

The present study aimed to provide a complete exploration of the effect of sound intensity, frequency, duty cycle, microbubble volume and irradiation time on low-frequency low-intensity ultrasound (US)-mediated microvessel disruption, and to identify an optimal combination of the five factors that maximize the blockage effect. An orthogonal experimental design approach was used. Enhanced US imaging and acoustic quantification were performed to assess tumor blood perfusion. In the confirmatory test, in addition to acoustic quantification, the specimens of the tumor were stained with hematoxylin and eosin and observed using light microscopy. The results revealed that sound intensity, frequency, duty cycle, microbubble volume and irradiation time had a significant effect on the average peak intensity (API). The extent of the impact of the variables on the API was in the following order: Sound intensity; frequency; duty cycle; microbubble volume; and irradiation time. The optimum conditions were found to be as follows: Sound intensity, 1.00 W/cm2; frequency, 20 Hz; duty cycle, 40%; microbubble volume, 0.20 ml; and irradiation time, 3 min. In the confirmatory test, the API was 19.97±2.66 immediately subsequent to treatment, and histological examination revealed signs of tumor blood vessel injury in the optimum parameter combination group. In conclusion, the Taguchi L18 (3)6 orthogonal array design was successfully applied for determining the optimal parameter combination of API following treatment. Under the optimum orthogonal design condition, a minimum API of 19.97±2.66 subsequent to low-frequency and low-intensity mediated blood perfusion blockage was obtained.