title: A synthetic substrate to support early mesodermal differentiation of human embryonic stem cells.
abstract: Our ability to guide differentiation of human pluripotent stem cells (hPSCs) toward desired lineages efficiently and reproducibly in xeno-free conditions is the key to advancing hPSC technology from the laboratory to clinical use. Here we report an engineered biomimetic substrate functionalized with both peptide ligands for α5β1 and α6β1 integrins to support efficient early mesodermal differentiation of human embryonic stem cells (hESCs) when cultured in a differentiation medium containing BMP4. In contrast, mesodermal differentiation is not induced on substrates functionalized with either ligand alone even though the culture medium is identical. Mesodermal differentiation was characterized by immunofluorescent staining, flow cytometric analysis, and RT-PCR analysis of early mesodermal markers Brachyury, Mixl1, and Wnt3. The early mesodermal progenitors derived on the substrate functionalized with both integrin ligands have the normal developmental potential to further differentiate along the hemato-endothelial and cardiac lineages. Immobilized ligands for α5β1 and α6β1 integrins both are permissive, necessary, and sufficient insoluble ligands in this engineered system to support early mesodermal differentiation of hESCs. This synthetic substrate, in conjunction with defined soluble factors, constructs a well-controlled and xeno-free early mesodermal differentiation niche that offers advantages over the previously reported niche constructed with the Matrigel-coated substrate.
authors.affiliation: Department of Biomedical Engineering, University of Minnesota, Minneapolis, MN 55455, USA.
authors.forename: Yang
authors.initials: Y
authors.lastname: Liu
authors.forename: Xintong
authors.initials: X
authors.lastname: Wang
authors.forename: Dan S
authors.initials: DS
authors.lastname: Kaufman
authors.forename: Wei
authors.initials: W
authors.lastname: Shen
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D015415:Biomarkers / Q000378:metabolism; D001773:Blood Cells / Q000166:cytology / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects*; D019070:Cell Lineage / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects; D005326:Fetal Proteins / Q000235:genetics / Q000378:metabolism; D005434:Flow Cytometry; D050991:GATA4 Transcription Factor / Q000235:genetics / Q000378:metabolism; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D016023:Integrins / Q000378:metabolism; D008024:Ligands; D026161:MADS Domain Proteins / Q000235:genetics / Q000378:metabolism; D064326:MEF2 Transcription Factors; D008648:Mesoderm / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D009206:Myocardium / Q000166:cytology; D018006:Myogenic Regulatory Factors / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000494:pharmacology*; D020825:T-Box Domain Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D000098908:Brachyury Protein
chemical_list: D018952:Antigens, CD34; D015415:Biomarkers; D005326:Fetal Proteins; D050991:GATA4 Transcription Factor; D016023:Integrins; D008024:Ligands; D026161:MADS Domain Proteins; D064326:MEF2 Transcription Factors; D018006:Myogenic Regulatory Factors; D010455:Peptides; D020825:T-Box Domain Proteins; D000098908:Brachyury Protein; C484170:GATA4 protein, human; C578035:MEF2A protein, human