title: Polymeric nanofibrous substrates stimulate pluripotent stem cells to form three-dimensional multilayered patty-like spheroids in feeder-free culture and maintain their pluripotency.
abstract: Expansion of pluripotent stem cells in defined media devoid of animal-derived feeder cells to generate multilayered three-dimensional (3D) bulk preparations or spheroids, rather than two-dimensional (2D) monolayers, is advantageous for many regenerative, biological or disease-modelling studies. Here we show that electrospun polymer matrices comprised of nanofibres that mimic the architecture of the natural fibrous extracellular matrix allow for feeder-free expansion of pluripotent human induced pluripotent stem cells (IPSCs) and human embryonic stem cells (HESCs) into multilayered 3D 'patty-like' spheroid structures in defined xeno-free culture medium. The observation that IPSCs and HESCs readily revert to 2D growth in the absence of the synthetic nanofibre membranes suggests that this 3D expansion behaviour is mediated by the physical microenvironment and artificial niche provided by the nanofibres only. Importantly, we could show that such 3D growth as patties maintained the pluripotency of cells as long as they were kept on nanofibres. The generation of complex multilayered 3D structures consisting of only pluripotent cells on biodegradable nanofibre matrices of the desired shape and size will enable both industrial-scale expansion and intricate organ-tissue engineering applications with human pluripotent stem cells, where simultaneous coupling of differentiation pathways of all germ layers from one stem cell source may be required for organ formation.
authors.affiliation: Clem Jones Research Centre for Stem Cells and Tissue Regenerative Therapies, Bond University, Gold Coast, Queensland, Australia.|Faciomaxillary and Regenerative Surgery, Griffith University, Gold Coast, Queensland, Australia.
authors.forename: Mohammad A
authors.initials: MA
authors.lastname: Alamein
authors.affiliation: Australian Institute for Bioengineering and Nanotechnology, University of Queensland, Brisbane, Queensland, Australia.
authors.forename: Ernst J
authors.initials: EJ
authors.lastname: Wolvetang
authors.affiliation: Australian Institute for Bioengineering and Nanotechnology, University of Queensland, Brisbane, Queensland, Australia.
authors.forename: Dmitry A
authors.initials: DA
authors.lastname: Ovchinnikov
authors.affiliation: Clem Jones Research Centre for Stem Cells and Tissue Regenerative Therapies, Bond University, Gold Coast, Queensland, Australia.|Faciomaxillary and Regenerative Surgery, Griffith University, Gold Coast, Queensland, Australia.
authors.forename: Sebastien
authors.initials: S
authors.lastname: Stephens
authors.affiliation: Faciomaxillary and Regenerative Surgery, Griffith University, Gold Coast, Queensland, Australia.
authors.forename: Katherine
authors.initials: K
authors.lastname: Sanders
authors.affiliation: Clem Jones Research Centre for Stem Cells and Tissue Regenerative Therapies, Bond University, Gold Coast, Queensland, Australia.|Faciomaxillary and Regenerative Surgery, Griffith University, Gold Coast, Queensland, Australia.
authors.forename: Patrick H
authors.initials: PH
authors.lastname: Warnke
mesh_terms: D018929:Cell Culture Techniques; D008855:Microscopy, Electron, Scanning; D057139:Nanofibers / Q000737:chemistry*; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000648:ultrastructure; D011108:Polymers / Q000737:chemistry*
chemical_list: D011108:Polymers
keywords: PLGA; PMEDSAH; electrospinning; extracellular matrix; feeder-free; human embryonic stem cells (HESCs); induced pluripotent stem cells (IPSCs); nanofibres; pluripotency; three-dimensional; xeno-free