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Ultrasound-targeted microbubble destruction enhances polyethylenimine-mediated gene transfection in vitro in human retinal pigment epithelial cells and in vivo in rat retina

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The aim of the present study was to investigate the efficacy and safety of ultrasound-targeted microbubble destruction (UTMD)-mediated polyethylenimine (PEI) transfection in cultured human retinal pigment epithelial (RPE) cells in vitro and rat retinas in vivo. An enhanced green fluorescent protein plasmid (pEGFP) was incubated with PEI to prepare a cationic complex (PEI/pEGFP), which was confirmed using a gel retardation assay. In the in vitro study, cultured human RPE cells were subjected to US waves under different conditions with or without microbubbles. The effect of UTMD on the viability of the cells was evaluated. In the in vivo study, gene transfer was examined by injecting PEI/pEGFP into the subretinal space of the rats. The rats treated with PEI/pEGFP and UTMD served as the experimental group, while rats treated with PEI/pEGFP alone served as the control group. The transfected tissue was visualized using an inverted fluorescence microscope. The expression of EGFP was classified into three groups, negative, weak positive and strong positive. Hematoxylin and eosin staining of frozen sections was used to observe tissue damage and the location of the EGFP gene expression. The electrophoresis experiment revealed that PEI treatment was able to condense DNA efficiently. In the in vitro study, the gene transfer efficiency under the optimal UTMD condition was enhanced and significantly higher than control groups. In the in vivo study, UTMD was able to enhance transgene expression in the retina without marked tissue damage. Frozen sections of the optic cups exhibited pEGFPpositive cells, predominantly distributed in the retina. This noninvasive novel combination of UTMD with PEI was able to enhance targeted gene delivery and gene expression in the rat retina without causing any apparent tissue damage, and may be a safe method to transfer genes and drug treatments directly to the retina, therefore being of potential therapeutic value.
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Document Type: Research Article

Affiliations: 1: Department of Ultrasound, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200127, P.R. China 2: Department of Ophthalmology, The Baoshan Branch Institute of Shanghai Shuguang Hospital, Shanghai 201900, P.R. China

Publication date: August 1, 2015

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  • Molecular Medicine Reports is a monthly, peer-reviewed journal available in print and online, that includes studies devoted to molecular medicine, underscoring aspects including pharmacology, pathology, genetics, neurosciences, infectious diseases, molecular cardiology and molecular surgery. In vitro and in vivo studies of experimental model systems pertaining to the mechanisms of a variety of diseases offer researchers the necessary tools and knowledge with which to aid the diagnosis and treatment of human diseases.
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