title: Mixl1 and Flk1 Are Key Players of Wnt/TGF-β Signaling During DMSO-Induced Mesodermal Specification in P19 cells.
abstract: Dimethyl sulfoxide (DMSO) is widely used to induce multilineage differentiation of embryonic and adult progenitor cells. To date, little is known about the mechanisms underlying DMSO-induced mesodermal specification. In this study, we investigated the signaling pathways and lineage-determining genes involved in DMSO-induced mesodermal specification in P19 cells. Wnt/β-catenin and TGF-β superfamily signaling pathways such as BMP, TGF-β and GDF1 signaling were significantly activated during DMSO-induced mesodermal specification. In contrast, Nodal/Cripto signaling pathway molecules, required for endoderm specification, were severely downregulated. DMSO significantly upregulated the expression of cardiac mesoderm markers but inhibited the expression of endodermal and hematopoietic lineage markers. Among the DMSO-activated cell lineage markers, the expression of Mixl1 and Flk1 was dramatically upregulated at both the transcript and protein levels, and the populations of Mixl1+, Flk1+ and Mixl1+/Flk1+ cells also increased significantly. DMSO modulated cell cycle molecules and induced cell apoptosis, resulting in significant cell death during EB formation of P19 cells. An inhibitor of Flk1, SU5416 significantly blocked expressions of TGF-β superfamily members, mesodermal cell lineage markers and cell cycle molecules but it did not affect Wnt molecules. These results demonstrate that Mixl1 and Flk1 play roles as key downstream or interacting effectors of Wnt/TGF-β signaling pathway during DMSO-induced mesodermal specification in P19 cells.
authors.affiliation: Department of Cardiology, Cardiovascular Center, Korea University Anam Hospital, Seoul, Republic of Korea.
authors.forename: Seung-Cheol
authors.initials: SC
authors.lastname: Choi
authors.forename: Ji-Hyun
authors.initials: JH
authors.lastname: Choi
authors.forename: Long-Hui
authors.initials: LH
authors.lastname: Cui
authors.forename: Ha-Rim
authors.initials: HR
authors.lastname: Seo
authors.forename: Jong-Ho
authors.initials: JH
authors.lastname: Kim
authors.forename: Chi-Yeon
authors.initials: CY
authors.lastname: Park
authors.forename: Hyung-Joon
authors.initials: HJ
authors.lastname: Joo
authors.forename: Jae-Hyoung
authors.initials: JH
authors.lastname: Park
authors.forename: Soon-Jun
authors.initials: SJ
authors.lastname: Hong
authors.forename: Cheol-Woong
authors.initials: CW
authors.lastname: Yu
authors.forename: Do-Sun
authors.initials: DS
authors.lastname: Lim
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D045744:Cell Line, Tumor; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D058732:Embryoid Bodies / Q000187:drug effects / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology; D005434:Flow Cytometry; D018398:Homeodomain Proteins / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D008648:Mesoderm / Q000166:cytology* / Q000187:drug effects; D060888:Real-Time Polymerase Chain Reaction; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism*; D051153:Wnt Proteins / Q000378:metabolism*
chemical_list: D018398:Homeodomain Proteins; C499230:MIXL1 protein, human; D016212:Transforming Growth Factor beta; D051153:Wnt Proteins; D040301:Vascular Endothelial Growth Factor Receptor-2; D004121:Dimethyl Sulfoxide