title: Optimization of transfection parameters for ultrasound/SonoVue microbubble-mediated hAng-1 gene delivery in vitro.
abstract: This study aimed to explore the effects of microbubble concentration, gene dosage, cell-microbubble mixing mode and fetal bovine serum (FBS) on gene delivery. 293T cells were transfected with Sonovue microbubbles carrying the hAng-1 gene via ultrasound irradiation. Various ultrasound exposure parameters and microbubble and DNA concentrations were investigated. In addition, FBS and the cell suspension or adherent mode was explored. Transfection efficiency and cell viability were used to determine the optimal transfection parameters. hAng-1 gene transfection efficiency gradually increased with elongation of ultrasound exposure and increasing microbubble concentration. However, if ultrasound irradiation exceeded 1.5 W/cm² and 30 sec or the microbubble concentration was over 20%, hAng-1 gene expression was significantly decreased, coupled with extensive cell death. Gene transfection levels were low under DNA concentrations less than 15 µg/ml. Furthermore, the gene transfer rate was significantly increased under cell suspension mode; FBS had no effect on hAng-1 gene transfection. The integrity of hAng-1 DNA was not affected by ultrasonic irradiation under optimal conditions. The optimal transfection parameters for the hAng-1 gene and Sonovue microbubble were ultrasound exposure of 1.5 W/cm² and 30 sec, 20% microbubbles, 15 µg/ml of DNA and under cell suspension mode.
authors.affiliation: Department of Ultrasound Imaging, Renmin Hospital of Wuhan University,                     Wuhan 430060, P.R. China.
authors.forename: Qing
authors.initials: Q
authors.lastname: Zhou
authors.forename: Jin-Ling
authors.initials: JL
authors.lastname: Chen
authors.forename: Qian
authors.initials: Q
authors.lastname: Chen
authors.forename: Xiao
authors.initials: X
authors.lastname: Wang
authors.forename: Qing
authors.initials: Q
authors.lastname: Deng
authors.forename: Bo
authors.initials: B
authors.lastname: Hu
authors.forename: Rui-Qiang
authors.initials: RQ
authors.lastname: Guo
mesh_terms: D042683:Angiopoietin-1 / Q000235:genetics* / Q000378:metabolism; D002470:Cell Survival; D004247:DNA / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D045423:Microbubbles; D010957:Plasmids / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection; D014465:Ultrasonics
chemical_list: D042683:Angiopoietin-1; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D004247:DNA