RAAV2-EGFP transfer to rat retina mediated by ultrasound and SonoVue after intravitreal and subretinal injection

Abstract

Objective To investigate the practical efficacy and safety of ultrasound with microbubbles mediated rAAV2-EGFP to retina of rat after intravitreal and subretinal injection. Methods Gene transfer was examined by rAAV2-EGFP intravitreal and subretinal injection into the Wistar rats with or without microbubbles. The eyes were exposed to US (1 MHz,2 W/cm2, duration 5 minutes,duty cycle 50%,pulse recurrent frequency 100 Hz). The onset of EGFP gene expression, lightness of fluorescence, area of fluorescence and its distribution in the fundus in vivo via fluorescence stereosocope were investigated on the 4th,7th, 35th,49th and 120th day respectively. The value of gene transfer was quantified through the EGFP fluorescence quantitative methods by Axiovision 3. 1 software. HE staining was used to observe tissue damage. Results There was no fluorescence observed by fluorescence stereosocope after intravitreal injection after two-month study. After subretinal injection, ultrasound-targeted microbubbles destruction (UTMD) strongly increased gene transfer efficiency. UTMD used in the experiment did no harm to the rat retina structure. Conclusions UTMD could not enhance rAAV2-EGFP transfecion efficiency to rat retina after intravitreal injection but the transduction could be enhanced significantly after subretinal injection. Key words: Ultrasonic therapy; Retina; Transfection; Adenoviridae