title: Accelerated high-yield generation of limb-innervating motor neurons from human stem cells.
abstract: Human pluripotent stem cells are a promising source of differentiated cells for developmental studies, cell transplantation, disease modeling, and drug testing. However, their widespread use even for intensely studied cell types like spinal motor neurons is hindered by the long duration and low yields of existing protocols for in vitro differentiation and by the molecular heterogeneity of the populations generated. We report a combination of small molecules that within 3 weeks induce motor neurons at up to 50% abundance and with defined subtype identities of relevance to neurodegenerative disease. Despite their accelerated differentiation, motor neurons expressed combinations of HB9, ISL1, and column-specific markers that mirror those observed in vivo in human embryonic spinal cord. They also exhibited spontaneous and induced activity, and projected axons toward muscles when grafted into developing chick spinal cord. Strikingly, this novel protocol preferentially generates motor neurons expressing markers of limb-innervating lateral motor column motor neurons (FOXP1(+)/LHX3(-)). Access to high-yield cultures of human limb-innervating motor neuron subtypes will facilitate in-depth study of motor neuron subtype-specific properties, disease modeling, and development of large-scale cell-based screening assays.
authors.affiliation: Project A.L.S./Jenifer Estess Laboratory for Stem Cell Research, New York, New York 10032, USA.
authors.forename: Mackenzie W
authors.initials: MW
authors.lastname: Amoroso
authors.forename: Gist F
authors.initials: GF
authors.lastname: Croft
authors.forename: Damian J
authors.initials: DJ
authors.lastname: Williams
authors.forename: Sean
authors.initials: S
authors.lastname: O'Keeffe
authors.forename: Monica A
authors.initials: MA
authors.lastname: Carrasco
authors.forename: Anne R
authors.initials: AR
authors.lastname: Davis
authors.forename: Laurent
authors.initials: L
authors.lastname: Roybon
authors.forename: Derek H
authors.initials: DH
authors.lastname: Oakley
authors.forename: Tom
authors.initials: T
authors.lastname: Maniatis
authors.forename: Christopher E
authors.initials: CE
authors.lastname: Henderson
authors.forename: Hynek
authors.initials: H
authors.lastname: Wichterle
mesh_terms: D000818:Animals; D001369:Axons / Q000502:physiology; D002118:Calcium / Q000502:physiology; D020013:Calcium Signaling / Q000502:physiology; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D002642:Chick Embryo; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D005121:Extremities / Q000294:innervation*; D005260:Female; D051858:Forkhead Transcription Factors / Q000096:biosynthesis / Q000235:genetics; D018398:Homeodomain Proteins / Q000235:genetics; D006801:Humans; D007150:Immunohistochemistry; D060850:LIM-Homeodomain Proteins / Q000235:genetics; D008297:Male; D051379:Mice; D009046:Motor Neurons / Q000378:metabolism / Q000502:physiology*; D058953:Neural Stem Cells / Q000378:metabolism / Q000502:physiology*; D018408:Patch-Clamp Techniques; D034743:RNA-Induced Silencing Complex; D012097:Repressor Proteins / Q000096:biosynthesis / Q000235:genetics; D013116:Spinal Cord / Q000166:cytology / Q000196:embryology; D033581:Stem Cell Transplantation / Q000379:methods; D014157:Transcription Factors / Q000235:genetics
chemical_list: D018076:DNA, Complementary; D051858:Forkhead Transcription Factors; C432917:Foxp1 protein, mouse; D018398:Homeodomain Proteins; D060850:LIM-Homeodomain Proteins; C490003:MNX1 protein, human; D034743:RNA-Induced Silencing Complex; D012097:Repressor Proteins; D014157:Transcription Factors; C063843:insulin gene enhancer binding protein Isl-1; D002118:Calcium