Molecular Biotechnology

, Volume 28, Issue 1, pp 21–31 | Cite as

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

  • Srdjan Djurovic
  • Nina Iversen
  • Stig Jeansson
  • Frank Hoover
  • Geir Christensen
Research

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.

Index Entries

Liposomes lipofection electroporation gene therapy nucleofection adeno-associated virus 

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Copyright information

© Humana Press Inc 2004

Authors and Affiliations

  • Srdjan Djurovic
    • 1
  • Nina Iversen
    • 1
  • Stig Jeansson
    • 2
  • Frank Hoover
    • 3
  • Geir Christensen
    • 4
    • 5
  1. 1.Department/Institute of Medical GeneticsUllevaal University HospitalOsloNorway
  2. 2.Department of MicrobiologyUllevaal University HospitalOsloNorway
  3. 3.Department of Oncology, Gene Therapy ProgramHaukeland University HospitalBergenNorway
  4. 4.Institute for Experimental Medical ResearchUllevaal University HospitalOsloNorway
  5. 5.Center for Heart Failure ResearchUllevaal University HospitalOsloNorway

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