title: Stable Somatic Gene Expression in Mouse Lungs Following Electroporation-mediated Tol2 Transposon Delivery.
abstract: Gene delivery to the lung has rapidly progressed as an important method for studying various chronic lung diseases. Viral vectors, albeit highly efficient, are limited by the host immune response. Electroporation, a well-known non-viral method, can efficiently deliver genes to the lung, but is unable to induce stable gene expression. The Tol2 transposon is another non-viral method that can induce stable gene expression by reinserting its genes into the host genome. In this study, we combined electroporation and Tol2 transposons to obtain stable, high-level gene expression in the mouse lung. Tol2 transposon plasmids (pT2A-EGFP; Tol2, pCAGGS-TP; transposase) were optimized in vitro, and the electroporation procedure (pCAG-EGFP) was optimized in mouse lungs. After optimization, a combination of electroporation plus the Tol2 transposon was used in a comparative analysis with electroporation plus pCAG-EGFP. GFP expression levels were quantified and visualized on days 4 and 7 post-electroporation. We successfully reproduced the Tol2 transposon system in vitro and the electroporation procedure in vivo. We observed sustainable GFP expression using electroporation plus the Tol2 transposon on days 4 and 7, while electroporation plus pCAG-EGFP resulted in decreased GFP expression on day 7. We were able to induce high-level, stable gene expression in mouse lungs using a combination of electroporation and the Tol2 transposon. This represents a safer method for lung gene delivery that can be used as an alternative to viral vectors.
authors.affiliation: Division of Cardiovascular Medicine, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Japan.
authors.forename: Hary Sakti
authors.initials: HS
authors.lastname: Muliawan
authors.affiliation: Division of Cardiovascular Medicine, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Japan.|Department of Clinical Pharmacy, Kobe Pharmaceutical University, Kobe, Japan.
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Nakayama
authors.affiliation: Department of Clinical Pharmacy, Kobe Pharmaceutical University, Kobe, Japan.
authors.forename: Keiko
authors.initials: K
authors.lastname: Yagi
authors.affiliation: Department of Clinical Pharmacy, Kobe Pharmaceutical University, Kobe, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Ikeda
authors.affiliation: Department of Physiology and Systems Bioscience, Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Yagita
authors.affiliation: Division of Cardiovascular Medicine, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Japan.
authors.forename: Ken-ichi
authors.initials: K
authors.lastname: Hirata
authors.affiliation: Division of Cardiovascular Medicine, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Japan.|Department of Clinical Pharmacy, Kobe Pharmaceutical University, Kobe, Japan.
authors.forename: Noriaki
authors.initials: N
authors.lastname: Emoto
mesh_terms: D000818:Animals; D016466:CHO Cells; D003412:Cricetulus; D004251:DNA Transposable Elements / Q000235:genetics*; D018274:Electroporation / Q000379:methods; D015870:Gene Expression; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008168:Lung / Q000378:metabolism*; D008297:Male; D051379:Mice; D057507:Mice, 129 Strain; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D004251:DNA Transposable Elements; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins
keywords: Lung gene delivery; Stable gene expression; Tol2 transposon; electroporation