Noninvasive imaging of inflammation by ultrasound detection of phagocytosed microbubbles

Background-We have previously shown that microbubbles adhere to leukocytes in regions of inflammation. We hypothesized that these microbubbles are pha gocytosed by neutrophils and monocytes and remain acoustically active, perm itting their detection in inflamed tissue. Methods and Results-In vitro studies were performed in which activated leuk ocytes were incubated with albumin or lipid microbubbles and observed under microscopy. Microbubbles attached to the surface of activated neutrophils and monocytes, were phagocytosed, and remained intact for up to 30 minutes. The rate of destruction of the phagocytosed microbubbles on exposure to ul trasound was less (P less than or equal to 0.05) than that of free microbub bles at all acoustic pressures applied. Intravital microscopy and simultane ous ultrasound imaging of the cremaster muscle was performed in 6 mice to d etermine whether phagocytosed microbubbles could be detected in vive. Fifte en minutes after intravenous injection of fluorescein-labeled microbubbles, when the blood-pool concentration was negligible, the number of phagocytos ed/ attached microbubbles within venules was 7-fold greater in tumor necros is factor-alpha (TNF-alpha)-treated animals than in control animals (P<0.01 ). This increase in retained microbubbles resulted in a 5- to 6-fold-greate r (P<0.01) degree of ultrasound contrast enhancement than in controls. Conclusions-After attaching to activated neutrophils and monocytes, microbu bbles are phagocytosed intact. Despite viscoelastic damping, phagocytosed m icrobubbles remain responsive to ultrasound and can be detected by ultrasou nd in vive after clearance of freely circulating microbubbles from the bloo d pool. Thus, contrast ultrasound has potential for imaging sites of inflam mation.