Improving ultrasound reflectivity and stability of echogenic liposomal dispersions for use as targeted ultrasound contrast agents.

Targeted echogenic liposome dispersions for ultrasonic enhancement of vasoactive and pathological components of endothelium and atherosclerosis have recently been developed. The component lipids required for acoustic and targeting properties include phosphatidylcholine, phosphatidylethanolamine phosphatidylglycerol (PG), and cholesterol (CH), initially in a 60:8:2:30 mol % ratio. Component lipids, lyophilization, sugars, and freezing conditions were varied to optimize acoustic ultrasound reflectivity and acoustic stability. Echogenic liposome dispersions were made by using the dehydration-rehydration process. The lipid concentrations were varied (CH in the range 1 to 40 mol % and PG from 1 to 16 mol %). Variations in type and concentration of sugars were examined. The effect of freezing conditions and re-lyophilization was examined. Ultrasound reflectivity was assessed by using a 20-MHz intravascular ultrasound catheter and computer-assisted videodensitometry. Ultrasound reflectivity was optimized at a CH concentration of 10 mol %; PG concentration variation had essentially no effect on initial values of echogenicity. Optimal acoustic stability was observed with concentrations of 10-15 mol % CH and with a PG concentration greater than 4 mol %. Preparations made with 0.2 M mannitol were more ultrasound reflective than those made with lactose, trehalose, and sucrose. Re-lyophilization and freezing temperatures below -20 degrees C increased ultrasound reflectivity. We optimized the ultrasound properties of echogenic liposomal dispersions, the conditions of which provide some insight into the underlying lipid structures responsible. The preparations developed are now more stable and acoustically reflective than our previous preparations. This advances the development of echogenic lipid dispersions as targeted ultrasound contrast agents for use in general ultrasound as well as cardiovascular imaging.

Development of echogenic, plasmid-incorporated, tissue-targeted cationic liposomes that can be used for directed gene delivery.

RATIONALE AND OBJECTIVES: Echogenic antibody-conjugated anionic liposomes have been developed that allow directed tissue targeting and acoustic enhancement. These are not efficient for gene delivery. A cationic formulation that allows directed gene delivery while retaining acoustic properties may provide more efficient transfection. METHODS: Cationic liposomes were prepared and acoustic reflectivity was determined. Anti-fibrinogen-conjugated liposomes were laid on fibrin-coated slides and adherence was quantified using fluorescence techniques. Liposomes were combined with a reporter gene and plated on cell cultures. Human umbilical vein endothelial cells were stimulated to upregulate intercellular adhesion molecule-1 (ICAM-1) and were treated with anti-ICAM-1-conjugated liposomes, and gene expression was quantified. RESULTS: Cationic liposomes retained their acoustic reflectivity and demonstrated specific adherence to fibrin under flow conditions. Significant transfection of human umbilical vein endothelial cells was demonstrated, with higher gene expression seen with specific antibody-conjugated liposomes. CONCLUSIONS: Novel acoustic cationic liposomes have been developed that can be antibody conjugated for site-specific adherence and directed cell modification. This presents exciting potential for a vector that allows tissue enhancement and targeted gene delivery.

Loss of contrast intensity during systole in the left ventricular cavity with the use of the contrast agent Albunex. An analysis of its correlation with pressure and velocity.

RATIONALE AND OBJECTIVES: Several investigators have observed a decrease in video intensity in the left ventricular cavity during systole when using contrast echocardiography. It has been suggested that this phenomenon is related to microbubble instability. The authors propose that this phenomenon is, in part, related to the effects of pressure and velocity on the acoustic reflectance of ultrasound contrast agents. METHODS: Using an in vitro flow tube model and varying concentrations of Albunex contrast agent, the effects of pressure and velocity on microbubble video intensity were investigated. Velocity and pressure were varied independently and the imaging tube was scanned using three transducer frequencies at different concentrations of Albunex. Contrast video intensity was analyzed using high and low velocities (at constant pressure) and high and low pressures (at constant velocity). In addition, the fluid from the system was collected and imaged in a nonflowing reservoir tank to investigate the video intensity of the microbubbles when exposed to variable velocity and pressure. RESULTS: The video-intensity measurements were inversely and irreversibly related to ambient pressure changes (independent of velocity) in a tube model. However, video intensity varied inversely but reversibly with velocity (independent of pressure). This observation could not be explained simply by the "laminar flow" theory, by a change in transducer angulation, nor by a change in ultrasound imaging frame rate. This phenomenon was limited to Albunex microbubbles and was not observed with a contrast medium (corn starch) devoid of the acoustic properties of Albunex.