title: GFAP-independent inflammatory competence and trophic functions of astrocytes generated from murine embryonic stem cells.
abstract: The directed generation of pure astrocyte cultures from pluripotent stem cells has proven difficult. Generation of defined pluripotent-stem-cell derived astrocytes would allow new approaches to the investigation of plasticity and heterogeneity of astrocytes. We here describe a two-step differentiation scheme resulting in the generation of murine embryonic stem cell (mESC) derived astrocytes (MEDA), as characterized by the upregulation of 19 astrocyte-associated mRNAs, and positive staining of most cells for GFAP (glial fibrillary acidic protein), aquaporin-4 or glutamine synthetase. The MEDA cultures could be cryopreserved, and they neither contained neuronal, nor microglial cells. They also did not react to the microglial stimulus lipopolysaccharide, while inflammatory activation by a complete cytokine mix (CCM) or its individual components (TNF-α, IL1-β, IFN-γ) was readily observed. MEDA, stimulated by CCM, became susceptible to CD95 ligand-induced apoptosis and produced NO and IL-6. This was preceded by NF-kB activation, and up-regulation of relevant mRNAs. Also GFAP-negative astrocytes were fully inflammation-competent. Neurotrophic support by MEDA was found to be independent of GFAP expression. In summary, we described here the generation and functional characterization of microglia-free murine astrocytes, displaying phenotypic heterogeneity as is commonly observed in brain astrocytes.
authors.affiliation: Doerenkamp-Zbinden Chair of In Vitro Toxicology and Biomedicine, Department of Biology, University of Konstanz, Konstanz, Germany.
authors.forename: Philipp B
authors.initials: PB
authors.lastname: Kuegler
authors.forename: Bettina A
authors.initials: BA
authors.lastname: Baumann
authors.forename: Bastian
authors.initials: B
authors.lastname: Zimmer
authors.forename: Sascha
authors.initials: S
authors.lastname: Keller
authors.forename: Andreas
authors.initials: A
authors.lastname: Marx
authors.forename: Suzanne
authors.initials: S
authors.lastname: Kadereit
authors.forename: Marcel
authors.initials: M
authors.lastname: Leist
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000235:genetics*; D002460:Cell Line; D019070:Cell Lineage / Q000235:genetics; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism / Q000473:pathology*; D005904:Glial Fibrillary Acidic Protein / Q000172:deficiency / Q000502:physiology*; D007249:Inflammation / Q000378:metabolism / Q000473:pathology*; D051379:Mice; D008807:Mice, Inbred BALB C; D009414:Nerve Growth Factors / Q000378:metabolism*; D010641:Phenotype; D061251:Primary Cell Culture
chemical_list: D005904:Glial Fibrillary Acidic Protein; D009414:Nerve Growth Factors