Large-Scale Production of Mature Neurons from Human Pluripotent Stem Cells in a Three-Dimensional Suspension Culture System.
Human pluripotent stem cells (hPSCs) offer a renewable source of cells that can be expanded indefinitely and differentiated into virtually any type of cell in the human body, including neurons. This opens up unprecedented possibilities to study neuronal cell and developmental biology and cellular pathology of the nervous system, provides a platform for the screening of chemical libraries that affect these processes, and offers a potential source of transplantable cells for regenerative approaches to neurological disease. However, defining protocols that permit a large number and high yield of neurons has proved difficult. We present differentiation protocols for the generation of distinct subtypes of neurons in a highly reproducible manner, with minimal experiment-to-experiment variation. These neurons form synapses with neighboring cells, exhibit spontaneous electrical activity, and respond appropriately to depolarization. hPSC-derived neurons exhibit a high degree of maturation and survive in culture for up to 4-5 months, even without astrocyte feeder layers.
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA.
Alessandra
A
Rigamonti
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA.
Giuliana G
GG
Repetti
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA.
Chicheng
C
Sun
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA.
Feodor D
FD
Price
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA.
Danielle C
DC
Reny
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA.
Francesca
F
Rapino
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA.
Karen
K
Weisinger
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA.
Chen
C
Benkler
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA.
Quinn P
QP
Peterson
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA.
Lance S
LS
Davidow
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA; Department of Medicine, KI-AZ Integrated Cardio Metabolic Centre, Karolinska Institutet, NOVUM, Hälsovägen 7, 141 57 Huddinge, Sweden.
Emil M
EM
Hansson
Department of Stem Cell and Regenerative Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA. Electronic address: lee_rubin@harvard.edu.
Lee L
LL
Rubin
D015415:Biomarkers / Q000378:metabolism; D019208:Brain-Derived Neurotrophic Factor / Q000494:pharmacology; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000187:drug effects; D020934:Ciliary Neurotrophic Factor / Q000494:pharmacology; D051100:Glial Cell Line-Derived Neurotrophic Factor / Q000494:pharmacology; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D009415:Nerve Net / Q000166:cytology* / Q000502:physiology; D055495:Neurogenesis / Q000187:drug effects* / Q000235:genetics; D009474:Neurons / Q000145:classification / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D015588:Observer Variation; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D015203:Reproducibility of Results; D051785:Smad Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D018874:Spheroids, Cellular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D020825:T-Box Domain Proteins / Q000235:genetics / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism
C479740:BCL11B protein, human; D015415:Biomarkers; D019208:Brain-Derived Neurotrophic Factor; D020934:Ciliary Neurotrophic Factor; D051100:Glial Cell Line-Derived Neurotrophic Factor; C489788:MAP2 protein, human; D008869:Microtubule-Associated Proteins; D012097:Repressor Proteins; D051785:Smad Proteins; D020825:T-Box Domain Proteins; C574705:TBR1 protein, human; D025521:Tumor Suppressor Proteins; C579455:BDNF protein, human