Ultrasound mediates iRGD targeted liposome-microbubble complex for thrombolytic therapy in vitro / 中华超声影像学杂志

ABSTRACT

Objective To fabricate iRGD targeted liposome-microbubble complex containing uPA ( iRGD-LMC) ,and to improve the thrombolytic efficiency and reduce the risk of thrombolysis by iRGD-LMC combining with ultrasound targeted microbubbles destruction ( UTMD ) to release drug into the thrombus site with the help of microbubble cavitation effect . Methods Biotinylated iRGD-MBs were fabricated by thin-film rehydration method .Biotinylated liposomes containing uPA were fabricated by freeze-thaw method and were conjugated to the biotinylated iRGD-MBs surface through biotin-avidin linkage . The iRGD-LMC was subjected to confocal microscopy to determine the particle morphology . The concentration , average diameter and size distribution were determined by particle sizing instrument . The uPA loading efficiency was measured by BCA Protein Assay Kit . Ultrasound imaging was performed using a Vevo 2100 ultrasound imaging system . The iRGD-LMC was irradiated by different ultrasound time and intensity to release drug . Thrombolytic effect in vitro of iRGD-LMC combined with UTMD was observed on the thrombosis model which was extracted from mouse blood . Results iRGD-LMC was successfully prepared . iRGD-LMC was exhibited a well-defined spherical morphology and homogeneous distribution ,like ordinary microbubbles . The concentration of iRGD-LMC was ( 0 .51 ± 0 .03 ) × 109 / ml and average diameter was ( 2 .62 ± 0 .12) μm . Drugs loading efficiency was ( 3878 .5 ± 97 .8) μg uPA per 108 microbubbles . iRGD-LMC could achieve contrast-enhanced ultrasound imaging in vitro . The thrombolytic effect of iRGD-LMC +US group ( 87 .66 ± 1 .69) ％ was the best in vitro ,and had significant difference with others groups ( P <0 .05) ,followed by iRGD-LMC group ( 53 .32 ± 4 .86) ％ and uPA group ( 51 .09 ± 9 .01) ％ ,Compared with PBS group ,US group ( 23 .56 ± 9 .46) ％ had thrombolytic effect . Conclusions iRGD-LMC is successfully prepared ,which has the advantages of high drug loading of liposomes and good acoustic properties of microbubbles . iRGD-LMC combined with UTMD achieves a significant thrombolytic effect in vitro .