title: Engineering prokaryotic channels for control of mammalian tissue excitability.
abstract: The ability to directly enhance electrical excitability of human cells is hampered by the lack of methods to efficiently overexpress large mammalian voltage-gated sodium channels (VGSC). Here we describe the use of small prokaryotic sodium channels (BacNav) to create de novo excitable human tissues and augment impaired action potential conduction in vitro. Lentiviral co-expression of specific BacNavorthologues, an inward-rectifying potassium channel, and connexin-43 in primary human fibroblasts from the heart, skin or brain yields actively conducting cells with customizable electrophysiological phenotypes. Engineered fibroblasts ('E-Fibs') retain stable functional properties following extensive subculture or differentiation into myofibroblasts and rescue conduction slowing in an in vitro model of cardiac interstitial fibrosis. Co-expression of engineered BacNavwith endogenous mammalian VGSCs enhances action potential conduction and prevents conduction failure during depolarization by elevated extracellular K+, decoupling or ischaemia. These studies establish the utility of engineered BacNavchannels for induction, control and recovery of mammalian tissue excitability.
authors.affiliation: Department of Biomedical Engineering, Duke University, 101 Science Drive, Room 1427, Fitzpatrick CIEMAS, Durham, North Carolina 27708, USA.
authors.forename: Hung X
authors.initials: HX
authors.lastname: Nguyen
authors.affiliation: Department of Biomedical Engineering, Duke University, 101 Science Drive, Room 1427, Fitzpatrick CIEMAS, Durham, North Carolina 27708, USA.
authors.forename: Robert D
authors.initials: RD
authors.lastname: Kirkton
authors.affiliation: Department of Biomedical Engineering, Duke University, 101 Science Drive, Room 1427, Fitzpatrick CIEMAS, Durham, North Carolina 27708, USA.
authors.forename: Nenad
authors.initials: N
authors.lastname: Bursac
mesh_terms: D000200:Action Potentials / Q000235:genetics / Q000502:physiology*; D000818:Animals; D000831:Animals, Newborn; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D018031:Connexin 43 / Q000235:genetics / Q000378:metabolism; D055724:Electrophysiological Phenomena; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D057809:HEK293 Cells; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D017207:Rats, Sprague-Dawley; D014162:Transfection; D061566:Voltage-Gated Sodium Channels / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D018031:Connexin 43; D061566:Voltage-Gated Sodium Channels