title: Fibrin hydrogels induce mixed dorsal/ventral spinal neuron identities during differentiation of human induced pluripotent stem cells.
abstract: We hypothesized that generating spinal motor neurons (sMNs) from human induced pluripotent stem cell (hiPSC)-derived neural aggregates (NAs) using a chemically-defined differentiation protocol would be more effective inside of 3D fibrin hydrogels compared to 2D poly-L-ornithine(PLO)/laminin-coated tissue culture plastic surfaces. We performed targeted RNA-Seq using next generation sequencing to determine the substrate-specific differences in gene expression that regulate cell phenotype. Cells cultured on both substrates expressed sMN genes CHAT and MNX1, though persistent WNT signaling contributed to a higher expression of genes associated with interneurons in NAs cultured in 3D fibrin scaffolds. Cells in fibrin also expressed lower levels of astrocyte progenitor genes and higher levels of the neuronal-specific gene TUBB3, suggesting a purer population of neurons compared to 2D cultures.
  
  
  
  Fibrin scaffolds can support the neuronal differentiation of pluripotent stem cells. This study provides insight into how fibrin hydrogels affect neuronal induction by analyzing of the signaling pathways activated during the differentiation process. These insights can then be used to tailor the properties of these hydrogels to optimize the generation of sMNs for regenerative medicine applications.
authors.affiliation: Department of Biomedical Engineering, University of Victoria, British Columbia V8W2Y2, Canada.
authors.forename: John M
authors.initials: JM
authors.lastname: Edgar
authors.affiliation: Department of Biomedical Engineering, University of Victoria, British Columbia V8W2Y2, Canada.
authors.forename: Meghan
authors.initials: M
authors.lastname: Robinson
authors.affiliation: Department of Mechanical Engineering, University of Victoria, British Columbia V8W2Y2, Canada; Division of Medical Science, University of Victoria, British Columbia V8W2Y2, Canada; International Collaboration on Repair Discoveries (ICORD), Vancouver, British Columbia V5Z1M9, Canada. Electronic address: willerth@uvic.ca.
authors.forename: Stephanie M
authors.initials: SM
authors.lastname: Willerth
mesh_terms: D000818:Animals; D002417:Cattle; D002449:Cell Aggregation / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects* / Q000235:genetics; D019070:Cell Lineage / Q000187:drug effects / Q000235:genetics; D005337:Fibrin / Q000494:pharmacology*; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects; D053263:Gene Regulatory Networks / Q000187:drug effects; D006801:Humans; D020100:Hydrogels / Q000494:pharmacology*; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D007797:Laminin / Q000494:pharmacology; D008297:Male; D009046:Motor Neurons / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D010455:Peptides / Q000494:pharmacology; D017423:Sequence Analysis, RNA; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D013116:Spinal Cord / Q000166:cytology*; D013997:Time Factors
chemical_list: D020100:Hydrogels; D007797:Laminin; D010455:Peptides; C008973:polyornithine; D005337:Fibrin
keywords: Fibrin; Gene expression; Laminin; Motor neuron; Neural progenitor; RNA-Seq; Stem cells