title: Efficient conversion of astrocytes to functional midbrain dopaminergic neurons using a single polycistronic vector.
abstract: Direct cellular reprogramming is a powerful new tool for regenerative medicine. In efforts to understand and treat Parkinson's Disease (PD), which is marked by the degeneration of dopaminergic neurons in the midbrain, direct reprogramming provides a valuable new source of these cells. Astrocytes, the most plentiful cells in the central nervous system, are an ideal starting population for the direct generation of dopaminergic neurons. In addition to their potential utility in cell replacement therapies for PD or in modeling the disease in vitro, astrocyte-derived dopaminergic neurons offer the prospect of direct in vivo reprogramming within the brain. As a first step toward this goal, we report the reprogramming of astrocytes to dopaminergic neurons using three transcription factors - ASCL1, LMX1B, and NURR1 - delivered in a single polycistronic lentiviral vector. The process is efficient, with 18.2±1.5% of cells expressing markers of dopaminergic neurons after two weeks. The neurons exhibit expression profiles and electrophysiological characteristics consistent with midbrain dopaminergic neurons, notably including spontaneous pacemaking activity, stimulated release of dopamine, and calcium oscillations. The present study is the first demonstration that a single vector can mediate reprogramming to dopaminergic neurons, and indicates that astrocytes are an ideal starting population for the direct generation of dopaminergic neurons.
authors.affiliation: Department of Cell and Developmental Biology, Institute for Regenerative Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, United States of America. raddis@mail.med.upenn.edu
authors.forename: Russell C
authors.initials: RC
authors.lastname: Addis
authors.forename: Fu-Chun
authors.initials: FC
authors.lastname: Hsu
authors.forename: Rebecca L
authors.initials: RL
authors.lastname: Wright
authors.forename: Marc A
authors.initials: MA
authors.lastname: Dichter
authors.forename: Douglas A
authors.initials: DA
authors.lastname: Coulter
authors.forename: John D
authors.initials: JD
authors.lastname: Gearhart
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000166:cytology* / Q000378:metabolism*; D018076:DNA, Complementary / Q000235:genetics; D059290:Dopaminergic Neurons / Q000166:cytology* / Q000378:metabolism*; D055724:Electrophysiological Phenomena; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D005796:Genes / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D007150:Immunohistochemistry; D008636:Mesencephalon / Q000166:cytology* / Q000378:metabolism; D051379:Mice; D016366:Open Reading Frames / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D018076:DNA, Complementary; D014157:Transcription Factors