title: Noggin Over-Expressing Mouse Embryonic Fibroblasts and MS5 Stromal Cells Enhance Directed Differentiation of Dopaminergic Neurons from Human Embryonic Stem Cells.
abstract: Directed methods for differentiating human embryonic stem cells (hESCs) into dopaminergic (DA) precursor cells using stromal cells co-culture systems are already well established. However, not all of the hESCs differentiate into DA precursors using these methods. HSF6, H1, H7, and H9 cells differentiate well into DA precursors, but CHA13 and CHA15 cells hardly differentiate. To overcome this problem, we modified the differentiation system to include a co-culturing step that exposes the cells to noggin early in the differentiation process. This was done using γ-irradiated noggin-overexpressing CF1-mouse embryonic fibroblasts (MEF-noggin) and MS5 stromal cells (MS5-noggin and MS5-sonic hedgehog). After directed differentiation, RT-PCR analyses revealed that engrailed-1 (En-1), Lmx1b, and Nurr1, which are midbrain DA markers, were expressed regardless of differentiation stage. Moreover, tyrosine hydroxylase (Th) and an A9 midbrain-specific DA marker (Girk2) were expressed during differentiation, whereas levels of Oct3/4, an undifferentiated marker, decreased. Immunocytochemical analyses revealed that protein levels of the neuronal markers TH and TuJ1 increased during the final differentiation stage. These results demonstrate that early noggin exposure may play a specific role in the directed differentiation of DA cells from human embryonic stem cells.
authors.affiliation: Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul, Korea; Hanyang Biomedical Research Institute, Hanyang University, Seoul, Korea.
authors.forename: Mi-Sun
authors.initials: MS
authors.lastname: Lim
authors.affiliation: Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul, Korea.
authors.forename: Min-Seop
authors.initials: MS
authors.lastname: Shin
authors.affiliation: Department of Microbiology, College of Medicine, Hanyang University, Seoul, Korea.
authors.forename: Soo Young
authors.initials: SY
authors.lastname: Lee
authors.affiliation: Department of Neurology, Hangang Sacred Heart Hospital, Hallym University, Seoul, Korea.
authors.forename: Yang-Ki
authors.initials: YK
authors.lastname: Minn
authors.affiliation: Department of Obstetrics and Gynecology, College of Medicine, Hanyang University, Seoul, Korea.
authors.forename: Jeong-Kyu
authors.initials: JK
authors.lastname: Hoh
authors.affiliation: Department of Medical Genetics, College of Medicine, Hanyang University, Seoul, Korea.
authors.forename: Youl-Hee
authors.initials: YH
authors.lastname: Cho
authors.affiliation: Department of Physiology and Cell Therapy Center, Yonsei University College of Medicine, Seoul, Korea.
authors.forename: Dong-Wook
authors.initials: DW
authors.lastname: Kim
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Medicine, Hanyang University, Seoul, Korea.
authors.forename: Sang-Hun
authors.initials: SH
authors.lastname: Lee
authors.affiliation: Institute of Green Bio Science and Technology, Seoul National University, Pyeongchang, Korea.
authors.forename: Chun-Hyung
authors.initials: CH
authors.lastname: Kim
authors.affiliation: Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul, Korea; Hanyang Biomedical Research Institute, Hanyang University, Seoul, Korea; Department of Microbiology, College of Medicine, Hanyang University, Seoul, Korea.
authors.forename: Chang-Hwan
authors.initials: CH
authors.lastname: Park
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation / Q000235:genetics*; D018920:Coculture Techniques; D059290:Dopaminergic Neurons / Q000166:cytology / Q000378:metabolism*; D051524:Fibroblast Growth Factor 8 / Q000235:genetics / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels / Q000235:genetics / Q000378:metabolism; D000066449:Human Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D060850:LIM-Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2 / Q000235:genetics / Q000378:metabolism; D017154:Stromal Cells / Q000166:cytology / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014446:Tyrosine 3-Monooxygenase / Q000235:genetics / Q000378:metabolism; D000099176:Noggin Protein
chemical_list: D002352:Carrier Proteins; D051524:Fibroblast Growth Factor 8; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D060850:LIM-Homeodomain Proteins; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2; D014157:Transcription Factors; D014446:Tyrosine 3-Monooxygenase; D000099176:Noggin Protein; C094401:KCNJ6 protein, human; C112149:LIM homeobox transcription factor 1 beta