title: Myelination in coculture of established neuronal and Schwann cell lines.
abstract: Establishing stable coculture systems with neuronal and Schwann cell lines has been considered difficult, presumably because of their high proliferative activity and phenotypic differences from primary cultured cells. The present study is aimed at developing methods for myelin formation under coculture of the neural crest-derived pheochromocytoma cell line PC12 and the immortalized adult rat Schwann cell line IFRS1. Prior to coculture, PC12 cells were seeded at low density (3 × 10(2)/cm(2)) and maintained in serum-free medium with N2 supplement, ascorbic acid (50 μg/ml), and nerve growth factor (NGF) (50 ng/ml) for a week. Exposure to such a NGF-rich environment with minimum nutrients accelerated differentiation and neurite extension, but not proliferation, of PC12 cells. When IFRS1 cells were added to NGF-primed PC12 cells, the cell density ratio of PC12 cells to IFRS1 cells was adjusted from 1:50 to 1:100. The cocultured cells were then maintained in serum-free medium with B27 supplement, ascorbic acid (50 μg/ml), NGF (10 ng/ml), and recombinant soluble neuregulin-1 type III (25 ng/ml). Myelin formation was illustrated by light and electron microscopy performed at day 28 of coculture. The stable PC12-IFRS1 coculture system is free of technical and ethical problems arising from the primary culture and can be a valuable tool to study peripheral nerve degeneration and regeneration.
authors.affiliation: Department of Sensory and Motor Systems, ALS/Neuropathy Project (Laboratory of Peripheral Nerve Pathophysiology), Tokyo Metropolitan Institute of Medical Science, 2-1-6 Kamikitazawa, Setagaya-ku, Tokyo, 156-8506, Japan. sango-kz@igakuken.or.jp
authors.forename: Kazunori
authors.initials: K
authors.lastname: Sango
authors.forename: Emiko
authors.initials: E
authors.lastname: Kawakami
authors.forename: Hiroko
authors.initials: H
authors.lastname: Yanagisawa
authors.forename: Shizuka
authors.initials: S
authors.lastname: Takaku
authors.forename: Masami
authors.initials: M
authors.lastname: Tsukamoto
authors.forename: Kazunori
authors.initials: K
authors.lastname: Utsunomiya
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Watabe
mesh_terms: D000818:Animals; D002454:Cell Differentiation; D018920:Coculture Techniques; D009186:Myelin Sheath / Q000378:metabolism*; D009414:Nerve Growth Factors / Q000494:pharmacology; D020890:Neuregulin-1 / Q000378:metabolism; D016501:Neurites / Q000502:physiology; D009474:Neurons / Q000166:cytology* / Q000378:metabolism; D016716:PC12 Cells; D051381:Rats; D012583:Schwann Cells / Q000166:cytology* / Q000378:metabolism
chemical_list: D009414:Nerve Growth Factors; D020890:Neuregulin-1