title: A microdevice to locally electroporate embryos with high efficiency and reduced cell damage.
abstract: The ability to follow and modify cell behaviour with accurate spatiotemporal resolution is a prerequisite to study morphogenesis in developing organisms. Electroporation, the delivery of exogenous molecules into targeted cell populations through electric permeation of the plasma membrane, has been used with this aim in different model systems. However, current localised electroporation strategies suffer from insufficient reproducibility and mediocre survival when applied to small and delicate organisms such as early post-implantation mouse embryos. We introduce here a microdevice to achieve localised electroporation with high efficiency and reduced cell damage. In silico simulations using a simple electrical model of mouse embryos indicated that a dielectric guide-based design would improve on existing alternatives. Such a device was microfabricated and its capacities tested by targeting the distal visceral endoderm (DVE), a migrating cell population essential for anterior-posterior axis establishment. Transfection was efficiently and reproducibly restricted to fewer than four visceral endoderm cells without compromising cell behaviour and embryo survival. Combining targeted mosaic expression of fluorescent markers with live imaging in transgenic embryos revealed that, like leading DVE cells, non-leading ones send long basal projections and intercalate during their migration. Finally, we show that the use of our microsystem can be extended to a variety of embryological contexts, from preimplantation stages to organ explants. Hence, we have experimentally validated an approach delivering a tailor-made tool for the study of morphogenesis in the mouse embryo. Furthermore, we have delineated a comprehensive strategy for the development of ad hoc electroporation devices.
authors.affiliation: Laboratoire de Photonique et de Nanostructures, LPN-CNRS, route de Nozay, 91460 Marcoussis, France.
authors.forename: Elsa
authors.initials: E
authors.lastname: Mazari
authors.affiliation: Laboratoire de Photonique et de Nanostructures, LPN-CNRS, route de Nozay, 91460 Marcoussis, France.
authors.forename: Xuan
authors.initials: X
authors.lastname: Zhao
authors.affiliation: Institut de Recherche Interdisciplinaire en Biologie Humaine et Moléculaire, Université Libre de Bruxelles, Brussels 1070, Belgium.
authors.forename: Isabelle
authors.initials: I
authors.lastname: Migeotte
authors.affiliation: Institut Jacques Monod, CNRS, UMR7592, Univ Paris Diderot, Sorbonne Paris Cité, F-75205 Paris, France.
authors.forename: Jérôme
authors.initials: J
authors.lastname: Collignon
authors.affiliation: Laboratoire de Photonique et de Nanostructures, LPN-CNRS, route de Nozay, 91460 Marcoussis, France charlie.gosse@lpn.cnrs.fr pereagomez.aitana@ijm.univ-paris-diderot.fr.
authors.forename: Charlie
authors.initials: C
authors.lastname: Gosse
authors.affiliation: Institut Jacques Monod, CNRS, UMR7592, Univ Paris Diderot, Sorbonne Paris Cité, F-75205 Paris, France charlie.gosse@lpn.cnrs.fr pereagomez.aitana@ijm.univ-paris-diderot.fr.
authors.forename: Aitana
authors.initials: A
authors.lastname: Perea-Gomez
mesh_terms: D000818:Animals; D002465:Cell Movement; D003198:Computer Simulation; D018274:Electroporation / Q000295:instrumentation* / Q000379:methods; D004622:Embryo, Mammalian / Q000378:metabolism; D004707:Endoderm / Q000378:metabolism; D004867:Equipment Design; D005260:Female; D020342:Finite Element Analysis; D005456:Fluorescent Dyes / Q000737:chemistry; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D008904:Miniaturization; D008962:Models, Theoretical
chemical_list: D005456:Fluorescent Dyes
keywords: Finite element model simulations; Localised electroporation; Microsystem; Migration; Mouse embryo; Organ explant; Visceral endoderm