EFFECT OF A STABILIZED MICROBUBBLE ECHO CONTRAST AGENT ON HEMOLYSIS OF HUMAN ERYTHROCYTES EXPOSED TO HIGH-INTENSITY PULSED ULTRASOUND

Microbubble contrast agents have been shown to enhance ultrasonic cell lysis in vitro when exposed to continuous-wave ultrasound having spatial peak temporal average (SPTA) intensities of a few W/cm2. The response is strongly dependent upon the hematocrit (HCT) of the cell sample; detectable cell lysis essentially disappears as the HCT approaches 5%-10%. This study was conducted to determine whether high intensity pulsed ultrasound is an effective lytic agent in the presence of preexisting potential cavitation nuclei (Albunex(R) contrast agent). Human erythrocytes were suspended in autologous plasma to HCTs ranging from 1%-40%. Suspensions were exposed or sham exposed for 60 seconds to focused, pulsed ultrasound. The pulse duration was 1 msec, and the pulse repetition frequency was 20 Hz. The pressure amplitudes, spatial peak pulse average (SPPA) intensity, and SPTA intensity were 4.7 MPa peak positive pressure, -2.7 MPa peak negative pressure, 420 W/cm2, and 8.5 W/cm2, respectively. Samples were exposed to ultrasound in a dialysis membrane exposure vessel rotating at 200 rpm. When included in the erythrocyte samples, the Albunex concentration was 35 muL/mL suspension. Significant ultrasound-induced hemolysis in the absence of Albunex was observed only at the lowest HCT value tested (1%). In the presence of Albunex significant cell lysis was observed at all tested HCT values. The relative fraction of cells lysed by the combination of ultrasound exposure and Albunex diminished with increasing HCT, but the number of cells lysed per sample was nearly constant over the range of 5%-40% HCT. The ultrasound exposure parameters used in this study differ substantially from those associated with diagnostic imaging equipment; it is not valid to infer from the present results that the use of Albunex in diagnostic applications will induce or enhance hemolysis in vivo.