title: Hydrogels as feeder-free scaffolds for long-term self-renewal of mouse induced pluripotent stem cells.
abstract: Expanding undifferentiated induced pluripotent stem (iPS) cells in vitro is a basic requirement for application of iPS cells in both fundamental research and clinical regeneration. In this study, we intended to establish a simple, low cost and efficient method for the long-term self-renewal of mouse induced pluripotent stem (miPS) cells without using feeder-cells and adhesive proteins. Three scaffolds were selected for the long-term subculture of miPS cells over two months starting from passages 14 to 29: 1) a gelatin coated polystyrene (Gelatin-PS) that is a widely used scaffold for self-renewal of mouse embryonic stem (mES) cells; 2) a neutral hydrogel poly(N,N-dimethylacrylamide) (PDMAAm); and 3) a negatively charged hydrogel poly(2-acrylamido-2-methyl-propane sulfonic acid sodium salt) (PNaAMPS). Each passaged miPS cells on these scaffolds were cryopreserved successfully and the revived cells showed high viability and proliferation. The passaged miPS cells maintained a high undifferentiated state on all three scaffolds and a high level of pluripotency by expressing differentiation markers in vitro and forming teratomas in SCID mice with derivatives of all three germ layers. Compared to Gelatin-PS, the two hydrogels exhibited much better self-renewal performance in terms of high proliferation rate and level of expression of undifferentiated gene markers as well as efficiency in pluripotent teratoma formation. Furthermore, the PNaAMPS hydrogel demonstrated a slightly higher efficiency and simpler operation of cell expansion than the PDMAAm hydrogel. To conclude, PNaAMPS hydrogel is an excellent feeder-free scaffold because of its simplicity, low cost and high efficiency in expanding a large number of miPS cells in vitro.
authors.affiliation: Faculty of Advanced Life Science, Hokkaido University, Sapporo, 060-0810, Japan.
authors.forename: Jing Jing
authors.initials: JJ
authors.lastname: Yang
authors.forename: Jian Fang
authors.initials: JF
authors.lastname: Liu
authors.forename: Takayuki
authors.initials: T
authors.lastname: Kurokawa
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Kitada
authors.forename: Jian Ping
authors.initials: JP
authors.lastname: Gong
mesh_terms: D000818:Animals; D000943:Antigens, Differentiation / Q000096:biosynthesis*; D049109:Cell Proliferation; D020099:Coated Materials, Biocompatible / Q000138:chemical synthesis / Q000737:chemistry*; D005780:Gelatin / Q000737:chemistry; D020100:Hydrogels / Q000138:chemical synthesis / Q000737:chemistry*; D057026:Induced Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D011137:Polystyrenes / Q000737:chemistry; D054457:Tissue Scaffolds / Q000737:chemistry*
chemical_list: D000943:Antigens, Differentiation; D020099:Coated Materials, Biocompatible; D020100:Hydrogels; D011137:Polystyrenes; D005780:Gelatin