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Mechanistic analysis of electroporation-induced cellular uptake of macromolecules.

Publication ,  Journal Article
Zaharoff, DA; Henshaw, JW; Mossop, B; Yuan, F
Published in: Experimental Biology and Medicine (Maywood, N.J.)
January 2008

Pulsed electric field has been widely used as a nonviral gene delivery platform. The delivery efficiency can be improved through quantitative analysis of pore dynamics and intracellular transport of plasmid DNA. To this end, we investigated mechanisms of cellular uptake of macromolecules during electroporation. In the study, fluorescein isothiocyanate-labeled dextran (FD) with molecular weight of 4,000 (FD-4) or 2,000,000 (FD-2000) was added into suspensions of a murine mammary carcinoma cell (4T1) either before or at different time points (ie, 1, 2, or 10 sec) after the application of different pulsed electric fields (in high-voltage mode: 1.2-2.0 kV in amplitude, 99 microsec in duration, and 1-5 pulses; in low-voltage mode: 100-300 V in amplitude, 5-20 msec in duration, and 1-5 pulses). The intracellular concentrations of FD were quantified using a confocal microscopy technique. To understand transport mechanisms, a mathematical model was developed for numerical simulation of cellular uptake. We observed that the maximum intracellular concentration of FD-2000 was less than 3% of that in the pulsing medium. The intracellular concentrations increased linearly with pulse number and amplitude. In addition, the intracellular concentration of FD-2000 was approximately 40% lower than that of FD-4 under identical pulsing conditions. The numerical simulations predicted that the pores larger than FD-4 lasted <10 msec after the application of pulsed fields if the simulated concentrations were on the same order of magnitude as the experimental data. In addition, the simulation results indicated that diffusion was negligible for cellular uptake of FD molecules. Taken together, the data suggested that large pores induced in the membrane by pulsed electric fields disappeared rapidly after pulse application and convection was likely to be the dominant mode of transport for cellular uptake of uncharged macromolecules.

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Published In

Experimental Biology and Medicine (Maywood, N.J.)

DOI

EISSN

1535-3699

ISSN

1535-3702

Publication Date

January 2008

Volume

233

Issue

1

Start / End Page

94 / 105

Related Subject Headings

  • Permeability
  • Models, Biological
  • Mice
  • Fluorescein-5-isothiocyanate
  • Electroporation
  • Dextrans
  • Cell Membrane
  • Cell Line, Tumor
  • Biological Transport
  • Biochemistry & Molecular Biology
 

Citation

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Zaharoff, D. A., Henshaw, J. W., Mossop, B., & Yuan, F. (2008). Mechanistic analysis of electroporation-induced cellular uptake of macromolecules. Experimental Biology and Medicine (Maywood, N.J.), 233(1), 94–105. https://doi.org/10.3181/0704-rm-113
Zaharoff, David A., Joshua W. Henshaw, Brian Mossop, and Fan Yuan. “Mechanistic analysis of electroporation-induced cellular uptake of macromolecules.Experimental Biology and Medicine (Maywood, N.J.) 233, no. 1 (January 2008): 94–105. https://doi.org/10.3181/0704-rm-113.
Zaharoff DA, Henshaw JW, Mossop B, Yuan F. Mechanistic analysis of electroporation-induced cellular uptake of macromolecules. Experimental Biology and Medicine (Maywood, NJ). 2008 Jan;233(1):94–105.
Zaharoff, David A., et al. “Mechanistic analysis of electroporation-induced cellular uptake of macromolecules.Experimental Biology and Medicine (Maywood, N.J.), vol. 233, no. 1, Jan. 2008, pp. 94–105. Epmc, doi:10.3181/0704-rm-113.
Zaharoff DA, Henshaw JW, Mossop B, Yuan F. Mechanistic analysis of electroporation-induced cellular uptake of macromolecules. Experimental Biology and Medicine (Maywood, NJ). 2008 Jan;233(1):94–105.
Journal cover image

Published In

Experimental Biology and Medicine (Maywood, N.J.)

DOI

EISSN

1535-3699

ISSN

1535-3702

Publication Date

January 2008

Volume

233

Issue

1

Start / End Page

94 / 105

Related Subject Headings

  • Permeability
  • Models, Biological
  • Mice
  • Fluorescein-5-isothiocyanate
  • Electroporation
  • Dextrans
  • Cell Membrane
  • Cell Line, Tumor
  • Biological Transport
  • Biochemistry & Molecular Biology