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Ultrasound-targeted microbubble destruction enhances AAV-mediated gene transfection in human RPE cells in vitro and rat retina in vivo

Gene Therapy volume 16, pages 1146–1153 (2009)Cite this article

Abstract

This study was conducted to investigate the efficacy and safety of ultrasound (US)-targeted microbubble (MB) destruction (UTMD)-mediated rAAV2-CMV-EGFP transfection to cultured human retinal pigment epithelium (RPE) cells in vitro and to the rat retina in vivo. In the in vitro study, cultured human RPE cells were exposed to US under different conditions with or without MBs. Furthermore, the effect of UTMD on rAAV2-CMV-EGFP itself and on cells was evaluated. In the in vivo study, gene transfer was examined by injecting rAAV2-CMV-EGFP into the subretinal space of rats with or without MBs and then exposed to US. We investigated enhanced green fluorescent protein (EGFP) expression in vivo by stereomicroscopy and performed quantitative analysis using Axiovision 3.1 software. Hematoxylin and eosin staining and frozen sections were used to observe tissue damage and location of the EGFP gene expression. In the in vitro study, the transfection efficiency of rAAV2-CMV-EGFP under optimal UTMD was significantly higher than that of the control group (P=0.000). Furthermore, there was almost no cytotoxicity to the cells and to rAAV2-CMV-EGFP itself. In the in vivo study, UTMD could be used safely to enhance and accelerate the transgene expression of the retina. Fluorescence expression was mainly located in the retinal layer. UTMD is a promising method for gene delivery to the retina.

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Acknowledgements

This project was supported by the National Natural Science Foundation of China (30772369, 30672440) and Shanghai Leading Academic Discipline Project (S30203). We thank professors Xiaodong Sun, Qing Gu and Jin Qian of Experimental Center and Experimental Research Laboratory of Ophthalmology of Shanghai Jiaotong University Affiliated First People's Hospital for their contribution to this study. Technical assistance from Fang Wei, Feng Wang, Huiming Li, Xiafang Chen and the entire staff at the Experimental Research Center of Shanghai Jiaotong University Affiliated First People's Hospital is gratefully acknowledged.

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  1. H L Li and X Z Zheng: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Ultrasound, Shanghai Jiaotong University Affiliated First People's Hospital, Shanghai, China

    H L Li, X Z Zheng, F Li, Y Wu & L F Du

  2. Experimental Research Center of Shanghai Jiaotong University Affiliated First People's Hospital, Shanghai, China

    H P Wang

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  1. H L Li
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Correspondence to L F Du.

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Li, H., Zheng, X., Wang, H. et al. Ultrasound-targeted microbubble destruction enhances AAV-mediated gene transfection in human RPE cells in vitro and rat retina in vivo. Gene Ther 16, 1146–1153 (2009). https://doi.org/10.1038/gt.2009.84

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