title: Decellularized feeders: an optimized method for culturing pluripotent cells.
abstract: Pluripotent cells such as human embryonic stem cells and human induced pluripotent stem cells are useful in the field of regenerative medicine because they can proliferate indefinitely and differentiate into all cell types. However, a limiting factor for maintaining and propagating stem cells is the need for inactivated fibroblasts as a growth matrix, since these may potentially cause cross-contamination. In this study, we aimed to maintain stem cells on the extracellular matrix (ECM) of either nonirradiated or γ-irradiated fibroblasts. It has been demonstrated that the ECM contains factors and proteins vital for the adhesion, proliferation, and differentiation of pluripotent cells. In order to preserve the ECM, the cell layers of the fibroblasts were decellularized by treatment with 0.05% sodium dodecyl sulfate (SDS), which resulted in an absence of DNA as compared with conventional feeder culture. However, SDS treatment did not cause a detectable change in the ECM architecture and integrity. Furthermore, immunohistochemistry demonstrated that expressions of major ECM proteins, such as fibronectin, collagen, and laminin, remained unaltered. The human pluripotent cells cultured on this decellularized matrix maintained gene expression of the pluripotency markers NANOG and OCT4 and had the potency to differentiate to three germ layers. The in vitro culture system shown here has an excellent potential since the main allogeneic components (i.e., DNA of the feeder cells) are removed. It is also a technically easy, fast, safe, and cheap method for maintaining a refined feeder-free stem cell culture for further cell differentiation studies.
authors.affiliation: Advanced Center for Translational Regenerative Medicine, Department for Clinical Science, Intervention and Technology, and.
authors.forename: Mei Ling
authors.initials: ML
authors.lastname: Lim
authors.forename: Philipp
authors.initials: P
authors.lastname: Jungebluth
authors.forename: Sebastian
authors.initials: S
authors.lastname: Sjöqvist
authors.forename: Hero
authors.initials: H
authors.lastname: Nikdin
authors.forename: Kristín Rós
authors.initials: KR
authors.lastname: Kjartansdóttir
authors.forename: Christian
authors.initials: C
authors.lastname: Unger
authors.forename: Ivan
authors.initials: I
authors.lastname: Vassliev
authors.forename: Paolo
authors.initials: P
authors.lastname: Macchiarini
mesh_terms: D015415:Biomarkers / Q000378:metabolism; D002448:Cell Adhesion; D002454:Cell Differentiation; D002460:Cell Line; D049109:Cell Proliferation; D048430:Cell Shape; D018920:Coculture Techniques; D004247:DNA / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism*; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D061252:Feeder Cells; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism / Q000502:physiology* / Q000528:radiation effects; D018507:Gene Expression Regulation, Developmental; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D000071317:Nanog Homeobox Protein; D050814:Octamer Transcription Factor-3 / Q000235:genetics / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000378:metabolism / Q000502:physiology*; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D013501:Surface-Active Agents / Q000494:pharmacology
chemical_list: D015415:Biomarkers; D016326:Extracellular Matrix Proteins; D018398:Homeodomain Proteins; C486780:NANOG protein, human; D000071317:Nanog Homeobox Protein; D050814:Octamer Transcription Factor-3; C494173:POU5F1 protein, human; D013501:Surface-Active Agents; D012967:Sodium Dodecyl Sulfate; D004247:DNA
keywords: Cell culture; Extracellular matrix; Pluripotent stem cells; Scaffold attachment region; Serum-free media; Stem cell; Stem cell culture; Technology