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Comparison of nonviral transfection and adeno-associated viral transduction on cardiomyocytes

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Abstract

Cardiomyocytes are terminally differentiated cells that to date have been characterized as poor targets for nonviral gene transfer. This study was therefore designed to determine the optimal nonviral gene transfer parameters in cell cultures of neonatal rat cardiomyocytes and to compare them with the efficiency of gene transfer using adeno-associated viral vectors (AAV). Transfection efficiency was measured by quantitative chloramphenicol acetyltransferase type I (CAT)-enzyme-linked immunosorbent assay and β-galactosidase staining, based on overexpression of reporter genes (CAT and LacZ).

The efficiency of CAT/LacZ overexpression was assessed using the following techniques: (1) liposomal reagents, such as: FuGENE 6, LipofectAMINE 2000, LipofectAMINE PLUS, GenePORTER, Metafectene, and LipoGen; (2) electroporation and nucleofector techniques; and (3) an AAV2 vector harboring a lacZ reporter gene. Toxicity was monitored by total protein measurement and by analyzing cell metabolism.

On average, Lipofectamine 2000 was the most effective nonviral method examined yielding consistently high transfection rates (8.1% β-galactosidase-positive cells) combined with low toxicity. Electroporation also resulted in high transfection values (7.5%); however, cellular toxicity was higher than that of Lipofectamine 2000. Finally, transduction with AAV2 vectors provided the highest levels of transduction (88.1%) with no cellular toxicity.

We conclude that although transduction with AAV is more efficient (88.1%), transfections with nonviral techniques, when optimized, may provide a useful alternative for overexpression of therapeutic genes in neonatal cardiomyocytes.

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Authors and Affiliations

  1. Department/Institute of Medical Genetics, Ullevaal University Hospital, Kirkeveien 166, N-0407, Oslo, Norway

    Srdjan Djurovic & Nina Iversen

  2. Department of Microbiology, Ullevaal University Hospital, Kirkeveien 166, N-0407, Oslo, Norway

    Stig Jeansson

  3. Department of Oncology, Gene Therapy Program, Haukeland University Hospital, Bergen, Norway

    Frank Hoover

  4. Institute for Experimental Medical Research, Ullevaal University Hospital, Oslo, Norway

    Geir Christensen

  5. Center for Heart Failure Research, Ullevaal University Hospital, Oslo, Norway

    Geir Christensen

Authors
  1. Srdjan Djurovic
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  2. Nina Iversen
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  3. Stig Jeansson
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  4. Frank Hoover
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  5. Geir Christensen
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Correspondence to Srdjan Djurovic.

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Djurovic, S., Iversen, N., Jeansson, S. et al. Comparison of nonviral transfection and adeno-associated viral transduction on cardiomyocytes. Mol Biotechnol 28, 21–31 (2004). https://doi.org/10.1385/MB:28:1:21

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