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Analysis and Comparison of Electrical Pulse Parameters for Gene Electrotransfer of Two Different Cell Lines

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Abstract

Knowledge of the parameters which influence the efficiency of gene electrotransfer has importance for practical implementation of electrotransfection for gene therapy as well as for better understanding of the underlying mechanism. The focus of this study was to analyze the differences in gene electrotransfer and membrane electropermeabilization between plated cells and cells in a suspension in two different cell lines (CHO and B16F1). Furthermore, we determined the viability and critical induced transmembrane voltage (ITVc) for both cell lines. In plated cells we obtained relatively little difference in electropermeabilization and gene electrotransfection between CHO and B16F1 cells. However, significant differences between the two cell lines were observed in a suspension. CHO cells exhibited a much higher gene electrotransfection rate compared to B16F1 cells, whereas B16F1 cells reached maximum electropermeabilization at lower electric fields than CHO cells. Both in a suspension and on plated cells, CHO cells had a slightly better survival rate at higher electric fields than B16F1 cells. Calculation of ITVc in a suspension showed that, for both electropermeabilization and gene electrotransfection, CHO cells have lower ITVc than B16F1 cells. In all cases, ITVc for electropermeabilization was lower than ITVc for gene electrotransfer, which is in agreement with other studies. Our results show that there is a marked difference in the efficiency of gene electrotransfer between suspended and plated cells.

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References

  • Cavazzana-Calvo M, Hacein-Bey S, de Saint Basile G, Gross F, Yvon E, Nusbaum P, Selz F, Hue C, Certain S, Casanova JL, Bousso P, Le Deist F, Fischer A (2000) Gene therapy of human severe combined immunodeficiency (SCID)-X1 disease. Science 288:669–672

    Article  CAS  PubMed  Google Scholar 

  • Cavazzana-Calvo M, Thrasher A, Mavilio F (2004) The future of gene therapy. Nature 427:779–781

    Article  CAS  PubMed  Google Scholar 

  • Cegovnik U, Novakovic S (2004) Setting optimal parameters for in vitro electrotransfection of B16F1, SA1, LPB, SCK, L929 and CHO cells using predefined exponentially decaying electric pulses. Bioelectrochemistry 62:73–82

    Article  CAS  PubMed  Google Scholar 

  • Cemazar M, Sersa G (2007) Electrotransfer of therapeutic molecules into tissues. Curr Opin Mol Ther 9:554–562

    CAS  PubMed  Google Scholar 

  • Corovic S, Zupanic A, Miklavcic D (2008) Numerical modeling and optimization of electric field distribution in subcutaneous tumor treated with electrochemotherapy using needle electrodes., 36:1665–1672

    Google Scholar 

  • Daud AI, DeConti RC, Andrews S, Urbas P, Riker AI, Sondak VK, Munster PN, Sullivan DM, Ugen KE, Messina JL, Heller R (2008) Phase I trial of interleukin-12 plasmid electroporation in patients with metastatic melanoma. J Clin Oncol 26:5896–5903

    Article  CAS  PubMed  Google Scholar 

  • Favard C, Dean DS, Rols MP (2007) Electrotransfer as a nonviral method of gene delivery. Curr Gene Ther 7:67–77

    Article  CAS  PubMed  Google Scholar 

  • Ferber D (2001) Gene therapy: safer and virus-free? Science 294:1638–1642

    Article  CAS  PubMed  Google Scholar 

  • Gehl J, Skovsgaard T, Mir LM (1998) Enhancement of cytotoxicity by electropermeabilization: an improved method for screening drugs. Anti Cancer Drugs 9:319–325

    Article  CAS  PubMed  Google Scholar 

  • Golzio M, Teissié J, Rols MP (2001) Control by membrane order of voltage-induced permeabilization, loading and gene transfer in mammalian cells. Bioelectrochemistry 53:25–34

    Article  CAS  PubMed  Google Scholar 

  • Golzio M, Teissié J, Rols MP (2002) Direct visualization at the single cell level of electrically mediated gene delivery. Proc Natl Acad Sci USA 99:1292–1297

    Article  CAS  PubMed  Google Scholar 

  • Hacein-Bey-Abina S, Von Kalle C, Schmidt M, McCormack MP, Wulffraat N, Leboulch P, Lim A, Osborne CS, Pawliuk R, Morillon E, Sorensen R, Forster A, Fraser P, Cohen JI, de Saint Basile G, Alexander I, Wintergerst U, Frebourg T, Aurias A, Stoppa-Lyonnet D, Romana S, Radford-Weiss I, Gross F, Valensi F, Delabesse E, Macintyre E, Sigaux F, Soulier J, Leiva LE, Wissler M, Prinz C, Rabbitts TH, Le Deist F, Fischer A, Cavazzana-Calvo M (2003) LMO2-associated clonal T cell proliferation in two patients after gene therapy for SCID-X1. Science 302:415–419

    Article  CAS  PubMed  Google Scholar 

  • Hojman P, Gissel H, Franck MA, Cournil-Henrionnet C, Eriksen J, Gehl J, Mir ML (2008) Physiological effects of high- and low-voltage pulse combinations for gene electrotransfer in muscle. Hum Gene Ther 19:1249–1260

    Article  CAS  PubMed  Google Scholar 

  • Kanduser M, Miklavcic D, Pavlin M (2009) Mechanisms involved in gene electrotransfer using high- and low-voltage pulses—an in vitro study. Bioelectrochemistry 74:265–271

    Article  CAS  PubMed  Google Scholar 

  • Lehrman S (1999) Virus treatment questioned after gene therapy death. Nature 401:517–518

    Article  CAS  PubMed  Google Scholar 

  • Li S, Huang L (2000) Nonviral gene therapy: promises and challenges. Gene Ther 7:31–34

    Article  CAS  PubMed  Google Scholar 

  • Mitsuyasu RT, Merigan TC, Carr A, Zack JA, Winters MA, Workman C, Bloch M, Lalezari J, Becker S, Thornton L, Akil B, Khanlou H, Finlayson R, McFarlane R, Smith DE, Garsia R, Ma D, Law M, Murray JM, von Kalle C, Ely JA, Patino SM, Knop AE, Wong P, Todd AV, Haughton M, Fuery C, Macpherson JL, Symonds GP, Evans LA, Pond SM, Cooper DA (2009) Phase 2 gene therapy trial of an anti-HIV ribozyme in autologous CD34+ cells. Nat Med 15:285–292

    Article  CAS  PubMed  Google Scholar 

  • Neumann E, Schaefer-Ridder M, Wang Y, Hofschneider PH (1982) Gene transfer into mouse lymoma cells by electroporation in high electric fields. EMBO J 7:841–845

    Google Scholar 

  • Neumann E, Sowers AE, Jordan CA (1989) Electroporation and electrofusion in cell biology. Plenum Press, New York

    Google Scholar 

  • Parker AL, Newman C, Briggs S, Seymour L, Sheridan PJ (2003) Nonviral gene delivery: techniques and implications for molecular medicine. Expert Rev Mol Med 5(22):1–15

    Article  PubMed  Google Scholar 

  • Pavlin M, Kanduser M, Rebersek M, Pucihar G, Hart FX, Magjarevic R, Miklavcic D (2005) Effect of cell electroporation on the conductivity of a cell suspension. Biophys J 88:4378–4390

    Article  CAS  PubMed  Google Scholar 

  • Pavlin M, Flisar K, Kanduser M (2010) The role of electrophoresis in gene electrotransfer. J Memb Biol. doi:10.1007/s00232-010-9276-z

  • Pucihar G, Mir LM, Miklavcic D (2002) The effect of pulse repetition frequency on the uptake into electropermeabilized cells in vitro with possible applications in electrochemotherapy. Bioelectrochemistry 57:167–172

    Article  CAS  PubMed  Google Scholar 

  • Pucihar G, Kotnik T, Valic B, Miklavcic D (2006) Numerical determination of transmembrane voltage induced on irregularly shaped cells. Ann Biomed Eng 34:642–652

    Article  CAS  PubMed  Google Scholar 

  • Rols MP, Teissié J (1992) Experimental evidence for the involvement of cytoskeleton in mammalian cell electropermeabilization. Biochim Biophys Acta 1111:45–50

    Article  CAS  PubMed  Google Scholar 

  • Rols MP, Teissié J (1998) Electropermeabilization of mammalian cells to macromolecules: control by pulse duration. Biophys J 75:1415–1423

    Article  CAS  PubMed  Google Scholar 

  • Rols MP, Delteil C, Golzio M, Teissié J (1998) Control by ATP and ADP of voltage-induced mammalian-cell-membrane permeabilization, gene transfer and resulting expression. Eur J Biochem 254:382–388

    Article  CAS  PubMed  Google Scholar 

  • Schwan HP (1957) Electrical properties of tissue and cell suspensions. Adv Biol Med Phys 5:147–209

    CAS  PubMed  Google Scholar 

  • Teissié J, Golzio M, Rols MP (2005) Mechanisms of cell membrane electropermeabilization: a minireview of our present (lack of?) knowledge. Biochim Biophys Acta 1724:270–280

    PubMed  Google Scholar 

  • Teissié J, Escoffre JM, Rols MP, Golzio M (2008) Time dependence of electric field effects on cell membranes. A review for a critical selection of pulse duration for therapeutical applications. Radiol Oncol 42:196–206

    Article  Google Scholar 

  • Towhidi L, Kotnik T, Pucihar G, Firoozabadi SMP, Mozdarani H, Miklavcic D (2008) Variability of the minimal transmembrane voltage resulting in detectable membrane electroporation. Electromagn Biol Med 27:372–385

    Article  CAS  PubMed  Google Scholar 

  • Valic B, Golzio M, Pavlin M, Schatz A, Faurie C, Gabriel B, Teissié J, Rols MP, Miklavcic D (2003) Effect of electric field induced transmembrane potential on spheroidal cells: theory and experiment. Biophys J 32:519–528

    Google Scholar 

  • Vaughan EE, Dean DA (2006) Intracellular trafficking of plasmids during transfection is mediated by microtubules. Mol Ther 3:422–428

    Article  Google Scholar 

  • Weaver JC, Chizmadzhev YA (1996) Theory of electroporation: a review. Bioelectrochem Bioenerg 41:135–160

    Article  CAS  Google Scholar 

  • Wolf H, Rols MP, Boldt E, Neumann E, Teissié J (1994) Control by pulse parameters of electric field-mediated gene transfer in mammalian cells. Biophys J 66:524–531

    Article  CAS  PubMed  Google Scholar 

  • Zaharoff DA, Barr RC, Li CY, Yuan Y (2002) Electromobility of plasmid DNA in tumor tissues during electric field–mediated gene delivery. Gene Ther 9:1286–1290

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

This research was supported by the Slovenian Research Agency under grants J2-9770 and P2-0249. We thank also Rosana Hudej (Faculty of Chemistry and Chemical Technology, University of Ljubljana) and Marko Ušaj (Faculty of Electrical Engineering, University of Ljubljana) for help in experimental procedures.

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  1. Faculty of Electrical Engineering, University of Ljubljana, Tržaška 25, 1000, Ljubljana, Slovenia

    Igor Marjanovič, Saša Haberl, Damijan Miklavčič, Maša Kandušer & Mojca Pavlin

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  1. Igor Marjanovič
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  2. Saša Haberl
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  3. Damijan Miklavčič
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  4. Maša Kandušer
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  5. Mojca Pavlin
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Correspondence to Mojca Pavlin.

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Marjanovič, I., Haberl, S., Miklavčič, D. et al. Analysis and Comparison of Electrical Pulse Parameters for Gene Electrotransfer of Two Different Cell Lines. J Membrane Biol 236, 97–105 (2010). https://doi.org/10.1007/s00232-010-9282-1

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  • DOI: https://doi.org/10.1007/s00232-010-9282-1

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