title: Involvement of a Rac1-Dependent Macropinocytosis Pathway in Plasmid DNA Delivery by Electrotransfection.
abstract: Electrotransfection is a widely used method for delivering genes into cells with electric pulses. Although different hypotheses have been proposed, the mechanism of electrotransfection remains controversial. Previous studies have indicated that uptake and intracellular trafficking of plasmid DNA (pDNA) are mediated by endocytic pathways, but it is still unclear which pathways are directly involved in the delivery. To this end, the present study investigated the dependence of electrotransfection on macropinocytosis. Data from the study demonstrated that electric pulses induced cell membrane ruffling and actin cytoskeleton remodeling. Using fluorescently labeled pDNA and a macropinocytosis marker (i.e., dextran), the study showed that electrotransfected pDNA co-localized with dextran in intracellular vesicles. Furthermore, electrotransfection efficiency could be decreased significantly by reducing temperature or treatment of cells with a pharmacological inhibitor of Rac1 and could be altered by changing Rac1 activity. Taken together, the findings suggested that electrotransfection of pDNA involved Rac1-dependent macropinocytosis.
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA.
authors.forename: Mao
authors.initials: M
authors.lastname: Mao
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA.
authors.forename: Liangli
authors.initials: L
authors.lastname: Wang
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA.
authors.forename: Chun-Chi
authors.initials: CC
authors.lastname: Chang
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA.
authors.forename: Katheryn E
authors.initials: KE
authors.lastname: Rothenberg
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA.
authors.forename: Jianyong
authors.initials: J
authors.lastname: Huang
authors.affiliation: Department of Bioengineering, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: Yingxiao
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA.
authors.forename: Brenton D
authors.initials: BD
authors.lastname: Hoffman
authors.affiliation: Department of Ophthalmology, Duke University, Durham, NC 27708, USA.
authors.forename: Paloma B
authors.initials: PB
authors.lastname: Liton
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA; Department of Ophthalmology, Duke University, Durham, NC 27708, USA. Electronic address: fyuan@duke.edu.
authors.forename: Fan
authors.initials: F
authors.lastname: Yuan
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D018274:Electroporation; D004705:Endocytosis; D015870:Gene Expression; D018014:Gene Transfer Techniques; D006801:Humans; D051379:Mice; D008856:Microscopy, Fluorescence; D010873:Pinocytosis; D010957:Plasmids / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D020830:rac1 GTP-Binding Protein / Q000235:genetics* / Q000378:metabolism*
chemical_list: D000199:Actins; D020830:rac1 GTP-Binding Protein
keywords: electrogene transfer; electroporation; endocytosis