Detection of Atherosclerotic Plaques in the Rabbit Aorta Using Ultrasound Microbubbles Conjugated to Interleukin-18 Antibodies

BACKGROUND: The purpose of the study was to investigate the ability of microbubbles (MBs) targeting interleukin-18 (IL-18) to detect plaques in a rabbit atherosclerotic plaque model. MATERIAL/METHODS: A rabbit atherosclerotic plaque model was established. The locations of the atherosclerotic plaques were verified by two-dimensional scanning and color Doppler flow imaging. An IL-18 antibody was conjugated to naked MBs (MB(c)) using the biotin-streptavidin conjugation method, resulting in the formation of MB(IL-18). MB(c) and MB(IL-18) were then used for contrast-enhanced ultrasound (CEUS) studies. The locations of CD34 and IL-18 within the plaques were determined by immunohistochemistry, and IL-18 expression levels in the plaques were determined by Western blot analysis. The relationships between IL-18 expression and the contrast intensity of the 2 MBs were analyzed. RESULTS: MB(c) and MB(IL-18) were both uniformly dispersed. Fluorescence microscopy and flow cytometry revealed that IL-18 was successfully conjugated to MBs. CEUS images showed that the intensity of the MB(IL-18) signal was substantially enhanced and prolonged compared with that of the MB(c) signal. Immunohistochemistry showed that CD34 expression was significantly increased in the plaques and that IL-18 was mainly located in the inner parts and base of the atherosclerotic plaques. Western blot analysis revealed that IL-18 expression was higher in the plaque regions. Correlation analysis showed that IL-18 expression was correlated with the contrast intensity of MB(IL-18) (r=0.903, P<0.05) but not with MB(c) (r=0.540, P>0.05). CONCLUSIONS: MBs targeting IL-18 may be a novel, noninvasive method of diagnosing atherosclerotic plaques.