title: The β-catenin/Tcf4/survivin signaling maintains a less differentiated phenotype and high proliferative capacity of human corneal epithelial progenitor cells.
abstract: It is clear that the microenvironment or niche plays an important role in determining the fate of stem cells: being stem cells or differentiated. However, the intrinsic pathways controlling the fate of adult stem cells in different niches are largely unknown. This study was to explore the role of β-catenin/Tcf4/survivin signaling in determining the fate of human corneal epithelial stem cells in different media. We observed that the low calcium serum-free media, especially CnT-20, promoted proliferative capacity, colony forming efficiency and stem cell-like phenotype of human corneal epithelial cells (HCECs) when compared with the cells cultured in a high calcium serum-containing medium SHEM. Three key factors in Wnt signaling, β-catenin, Tcf4 and survivin, were found to be expressed higher by HCECs grown in CnT-20 than those cultured in SHEM, as evaluated by real-time PCR, Western blotting and immunostaining. Transfection of siRNA-Tcf4 at 10-50nM knocked down Tcf4, and also significantly suppressed its down stream molecule survivin at both mRNA and protein levels in HCECs. Furthermore, Tcf4 silencing significantly suppressed the proliferative capacity of HCECs, measured by WST-1 assay, compared with the control groups, untreated or transfected with non-coding sequence siRNA-fluorescein. These findings demonstrate that low calcium serum free media promote ex vivo expansion of corneal epithelial progenitor cells that retain a less differentiated phenotype and high proliferative capacity via β-catenin/Tcf4/survivin signaling, a novel intrinsic pathway. This study may have high impact and clinic implication on the expansion of corneal epithelial stem cells in regenerative medicine, especially for ocular surface reconstruction.
authors.affiliation: Ocular Surface Center, Cullen Eye Institute, Department of Ophthalmology, Baylor College of Medicine, Houston, TX, USA.
authors.forename: Rong
authors.initials: R
authors.lastname: Lu
authors.forename: Fang
authors.initials: F
authors.lastname: Bian
authors.forename: Xiaobo
authors.initials: X
authors.lastname: Zhang
authors.forename: Hong
authors.initials: H
authors.lastname: Qi
authors.forename: Eliseu Y
authors.initials: EY
authors.lastname: Chuang
authors.forename: Stephen C
authors.initials: SC
authors.lastname: Pflugfelder
authors.forename: De-Quan
authors.initials: DQ
authors.lastname: Li
mesh_terms: D053687:Adult Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors / Q000235:genetics / Q000378:metabolism*; D002118:Calcium / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D019573:Epithelium, Corneal / Q000166:cytology*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D000077022:Survivin; D000073940:Transcription Factor 4; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism*
chemical_list: C107489:BIRC5 protein, human; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D016895:Culture Media, Serum-Free; D051034:Inhibitor of Apoptosis Proteins; D000077022:Survivin; C532096:TCF4 protein, human; D000073940:Transcription Factor 4; D014157:Transcription Factors; D051176:beta Catenin; D002118:Calcium