title: Direct conversion of mouse fibroblasts into induced neural stem cells.
abstract: Terminally differentiated cells can be directly converted into different types of somatic cells by using defined factors, thus circumventing the pluripotent state. However, low reprogramming efficiency, along with the absence of proliferation of some somatic cell types, makes it difficult to generate large numbers of cells with this method. Here we describe a protocol to directly convert mouse fibroblasts into self-renewing induced neural stem cells (iNSCs) that can be expanded in vitro, thereby overcoming the limitations associated with low reprogramming efficiency. The four transcription factors required for direct conversion into iNSCs (Sox2, Klf4, Myc (also known as c-Myc) and Pou3f4 (also known as Brn4)) do not generate a pluripotent cell state, and thus the risk for tumor formation after transplantation is reduced. By following the current protocol, iNSCs are observed 4-5 weeks after transduction. Two additional months are required to establish clonal iNSC cell lines that exhibit retroviral transgene silencing and that differentiate into neurons, astrocytes and oligodendrocytes.
authors.affiliation: 1] Department of Stem Cell Biology, School of Medicine, Konkuk University, Seoul, Republic of Korea. [2] Department of Animal Biotechnology, Konkuk University, Seoul, Republic of Korea. [3].
authors.forename: Sung Min
authors.initials: SM
authors.lastname: Kim
authors.affiliation: 1] Department of Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany. [2].
authors.forename: Hannah
authors.initials: H
authors.lastname: Flaßkamp
authors.affiliation: 1] Division for Neurodegenerative Diseases, Department of Neurology, Dresden University of Technology, Dresden, Germany. [2] Center for Regenerative Therapies Dresden, Dresden University of Technology, Dresden, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Hermann
authors.affiliation: 1] Department of Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany. [2] Group of Computational Biology and Bioinformatics, Biodonostia Health Research Institute, San Sebastián, Spain. [3] IKERBASQUE, Basque Foundation for Science, Bilbao, Spain.
authors.forename: Marcos Jesús
authors.identifier: 000000023264464X
authors.initials: MJ
authors.lastname: Araúzo-Bravo
authors.affiliation: Department of Stem Cell Biology, School of Medicine, Konkuk University, Seoul, Republic of Korea.
authors.forename: Seung Chan
authors.initials: SC
authors.lastname: Lee
authors.affiliation: Department of Animal Biotechnology, Konkuk University, Seoul, Republic of Korea.
authors.forename: Sung Ho
authors.initials: SH
authors.lastname: Lee
authors.affiliation: Department of Internal Medicine, School of Medicine, Konkuk University, Seoul, Republic of Korea.
authors.forename: Eun Hye
authors.initials: EH
authors.lastname: Seo
authors.affiliation: Department of Microbiology, School of Medicine, Konkuk University, Seoul, Republic of Korea.
authors.forename: Seung Hyun
authors.initials: SH
authors.lastname: Lee
authors.affiliation: 1] Division for Neurodegenerative Diseases, Department of Neurology, Dresden University of Technology, Dresden, Germany. [2] Center for Regenerative Therapies Dresden, Dresden University of Technology, Dresden, Germany. [3] Center for Regenerative Therapies Dresden, Dresden University of Technology, Dresden, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Storch
authors.affiliation: Department of Animal Biotechnology, Konkuk University, Seoul, Republic of Korea.
authors.forename: Hoon Taek
authors.initials: HT
authors.lastname: Lee
authors.affiliation: 1] Department of Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany. [2] Medical Faculty, University of Münster, Münster, Germany.
authors.forename: Hans R
authors.initials: HR
authors.lastname: Schöler
authors.affiliation: Department of Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany.
authors.forename: Natalia
authors.initials: N
authors.lastname: Tapia
authors.affiliation: Department of Stem Cell Biology, School of Medicine, Konkuk University, Seoul, Republic of Korea.
authors.forename: Dong Wook
authors.initials: DW
authors.lastname: Han
mesh_terms: D000818:Animals; D015415:Biomarkers; D018929:Cell Culture Techniques / Q000379:methods*; D065150:Cellular Reprogramming; D005347:Fibroblasts / Q000166:cytology*; D016259:Genes, myc; D057026:Induced Pluripotent Stem Cells / Q000502:physiology*; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000235:genetics; D051379:Mice; D008809:Mice, Inbred C3H; D009419:Nerve Tissue Proteins / Q000235:genetics; D058953:Neural Stem Cells; D050809:POU Domain Factors / Q000235:genetics; D055748:SOXB1 Transcription Factors / Q000235:genetics; D014161:Transduction, Genetic
chemical_list: D015415:Biomarkers; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D009419:Nerve Tissue Proteins; D050809:POU Domain Factors; D055748:SOXB1 Transcription Factors; C527783:Sox2 protein, mouse; C074067:Pou3f4 protein, mouse