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Mechanistic studies of gene delivery into mammalian cells by electrical short-circuiting via an aqueous droplet in dielectric oil

We have developed a novel methodology for the delivery of cell-impermeable molecules, based on electrical short-circuiting via a water droplet in dielectric oil. When a cell suspension droplet is placed between a pair of electrodes with an intense DC electric field, droplet bouncing and droplet defo...

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Autores principales: Kurita, Hirofumi, Nihonyanagi, Hirohito, Watanabe, Yuki, Sugano, Kenta, Shinozaki, Ryuto, Kishikawa, Kenta, Numano, Rika, Takashima, Kazunori
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7717561/
https://www.ncbi.nlm.nih.gov/pubmed/33275626
http://dx.doi.org/10.1371/journal.pone.0243361
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author Kurita, Hirofumi
Nihonyanagi, Hirohito
Watanabe, Yuki
Sugano, Kenta
Shinozaki, Ryuto
Kishikawa, Kenta
Numano, Rika
Takashima, Kazunori
author_facet Kurita, Hirofumi
Nihonyanagi, Hirohito
Watanabe, Yuki
Sugano, Kenta
Shinozaki, Ryuto
Kishikawa, Kenta
Numano, Rika
Takashima, Kazunori
author_sort Kurita, Hirofumi
collection PubMed
description We have developed a novel methodology for the delivery of cell-impermeable molecules, based on electrical short-circuiting via a water droplet in dielectric oil. When a cell suspension droplet is placed between a pair of electrodes with an intense DC electric field, droplet bouncing and droplet deformation, which results in an instantaneous short-circuit, can be induced, depending on the electric field strength. We have demonstrated successful transfection of various mammalian cells using the short-circuiting; however, the molecular mechanism remains to be elucidated. In this study, flow cytometric assays were performed with Jurkat cells. An aqueous droplet containing Jurkat cells and plasmids carrying fluorescent proteins was treated with droplet bouncing or short-circuiting. The short-circuiting resulted in sufficient cell viability and fluorescent protein expression after 24 hours’ incubation. In contrast, droplet bouncing did not result in successful gene transfection. Transient membrane pore formation was investigated by uptake of a cell-impermeable fluorescence dye YO-PRO-1 and the influx of calcium ions. As a result, short-circuiting increased YO-PRO-1 fluorescence intensity and intracellular calcium ion concentration, but droplet bouncing did not. We also investigated the contribution of endocytosis to the transfection. The pre-treatment of cells with endocytosis inhibitors decreased the efficiency of gene transfection in a concentration-dependent manner. Besides, the use of pH-sensitive dye conjugates indicated the formation of an acidic environment in the endosomes after the short-circuiting. Endocytosis is a possible mechanism for the intracellular delivery of exogenous DNA.
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spelling pubmed-77175612020-12-09 Mechanistic studies of gene delivery into mammalian cells by electrical short-circuiting via an aqueous droplet in dielectric oil Kurita, Hirofumi Nihonyanagi, Hirohito Watanabe, Yuki Sugano, Kenta Shinozaki, Ryuto Kishikawa, Kenta Numano, Rika Takashima, Kazunori PLoS One Research Article We have developed a novel methodology for the delivery of cell-impermeable molecules, based on electrical short-circuiting via a water droplet in dielectric oil. When a cell suspension droplet is placed between a pair of electrodes with an intense DC electric field, droplet bouncing and droplet deformation, which results in an instantaneous short-circuit, can be induced, depending on the electric field strength. We have demonstrated successful transfection of various mammalian cells using the short-circuiting; however, the molecular mechanism remains to be elucidated. In this study, flow cytometric assays were performed with Jurkat cells. An aqueous droplet containing Jurkat cells and plasmids carrying fluorescent proteins was treated with droplet bouncing or short-circuiting. The short-circuiting resulted in sufficient cell viability and fluorescent protein expression after 24 hours’ incubation. In contrast, droplet bouncing did not result in successful gene transfection. Transient membrane pore formation was investigated by uptake of a cell-impermeable fluorescence dye YO-PRO-1 and the influx of calcium ions. As a result, short-circuiting increased YO-PRO-1 fluorescence intensity and intracellular calcium ion concentration, but droplet bouncing did not. We also investigated the contribution of endocytosis to the transfection. The pre-treatment of cells with endocytosis inhibitors decreased the efficiency of gene transfection in a concentration-dependent manner. Besides, the use of pH-sensitive dye conjugates indicated the formation of an acidic environment in the endosomes after the short-circuiting. Endocytosis is a possible mechanism for the intracellular delivery of exogenous DNA. Public Library of Science 2020-12-04 /pmc/articles/PMC7717561/ /pubmed/33275626 http://dx.doi.org/10.1371/journal.pone.0243361 Text en © 2020 Kurita et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Kurita, Hirofumi
Nihonyanagi, Hirohito
Watanabe, Yuki
Sugano, Kenta
Shinozaki, Ryuto
Kishikawa, Kenta
Numano, Rika
Takashima, Kazunori
Mechanistic studies of gene delivery into mammalian cells by electrical short-circuiting via an aqueous droplet in dielectric oil
title Mechanistic studies of gene delivery into mammalian cells by electrical short-circuiting via an aqueous droplet in dielectric oil
title_full Mechanistic studies of gene delivery into mammalian cells by electrical short-circuiting via an aqueous droplet in dielectric oil
title_fullStr Mechanistic studies of gene delivery into mammalian cells by electrical short-circuiting via an aqueous droplet in dielectric oil
title_full_unstemmed Mechanistic studies of gene delivery into mammalian cells by electrical short-circuiting via an aqueous droplet in dielectric oil
title_short Mechanistic studies of gene delivery into mammalian cells by electrical short-circuiting via an aqueous droplet in dielectric oil
title_sort mechanistic studies of gene delivery into mammalian cells by electrical short-circuiting via an aqueous droplet in dielectric oil
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7717561/
https://www.ncbi.nlm.nih.gov/pubmed/33275626
http://dx.doi.org/10.1371/journal.pone.0243361
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