Small Molecule Mesengenic Induction of Human Induced Pluripotent Stem Cells to Generate Mesenchymal Stem/Stromal Cells

The translational potential of mesenchymal stem/stromal cells (MSCs) is limited by their rarity in somatic organs, heterogeneity, and need for harvest by invasive procedures. Induced pluripotent stem cells (iPSCs) could be an advantageous source of MSCs, but attempts to derive MSCs from pluripotent...

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Published inStem cells translational medicine Vol. 1; no. 2; pp. 83 - 95
Main Authors Chen, Yen Shun, Pelekanos, Rebecca A., Ellis, Rebecca L., Horne, Rachel, Wolvetang, Ernst J., Fisk, Nicholas M.
Format Journal Article
LanguageEnglish
Published United States AlphaMed Press 01.02.2012
Oxford University Press
Subjects
Antigens, CD - genetics
Antigens, CD - metabolism
Benzamides - pharmacology
Biomarkers - metabolism
Bone marrow
Cadherins
Cadherins - genetics
Cadherins - metabolism
Cell adhesion & migration
Cell culture
Cell Culture Techniques - methods
Cell Differentiation
Cell Proliferation
Cell Shape
Cells, Cultured
Culture Media, Serum-Free
Cytology
Data analysis
Differentiation
Dioxoles - pharmacology
Embryo cells
Embryonic stem cells
Embryonic Stem Cells - cytology
Embryonic Stem Cells - metabolism
Epithelial-Mesenchymal Transition
Fetuses
Fibroblasts
Gene Expression Regulation
Growth factors
Humans
Immunohistochemistry
Immunophenotyping
Induced pluripotent stem cells
Induced Pluripotent Stem Cells - cytology
Induced Pluripotent Stem Cells - drug effects
Induced Pluripotent Stem Cells - metabolism
Inhibitory postsynaptic potentials
Mesenchymal stem cells
Mesenchymal Stromal Cells - cytology
Mesenchymal Stromal Cells - metabolism
Mesenchyme
Mesoderm - cytology
Mesoderm - metabolism
Msx2 protein
Neural cell adhesion molecule
Oct-4 protein
Octamer Transcription Factor-3 - genetics
Octamer Transcription Factor-3 - metabolism
Original and Reviews
Pluripotency
Pluripotent stem cells
Regenerative Medicine - methods
Stem cells
Stromal cells
Time Factors
Transforming growth factor
Transforming growth factor-b
Vimentin
Vimentin - genetics
Vimentin - metabolism
Online AccessGet full text
ISSN2157-6564
2157-6580
DOI10.5966/sctm.2011-0022

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Abstract The translational potential of mesenchymal stem/stromal cells (MSCs) is limited by their rarity in somatic organs, heterogeneity, and need for harvest by invasive procedures. Induced pluripotent stem cells (iPSCs) could be an advantageous source of MSCs, but attempts to derive MSCs from pluripotent cells have required cumbersome or untranslatable techniques, such as coculture, physical manipulation, sorting, or viral transduction. We devised a single‐step method to direct mesengenic differentiation of human embryonic stem cells (ESCs) and iPSCs using a small molecule inhibitor. First, epithelial‐like monolayer cells were generated by culturing ESCs/iPSCs in serum‐free medium containing the transforming growth factor‐β pathway inhibitor SB431542. After 10 days, iPSCs showed upregulation of mesodermal genes (MSX2, NCAM, HOXA2) and downregulation of pluripotency genes (OCT4, LEFTY1/2). Differentiation was then completed by transferring cells into conventional MSC medium. The resultant development of MSC‐like morphology was associated with increased expression of genes, reflecting epithelial‐to‐mesenchymal transition. Both ESC‐ and iPSC‐derived MSCs exhibited a typical MSC immunophenotype, expressed high levels of vimentin and N‐cadherin, and lacked expression of pluripotency markers at the protein level. Robust osteogenic and chondrogenic differentiation was induced in vitro in ES‐MSCs and iPS‐MSCs, whereas adipogenic differentiation was limited, as reported for primitive fetal MSCs and ES‐MSCs derived by other methods. We conclude that treatment with SB431542 in two‐dimensional cultures followed by culture‐induced epithelial‐to‐mesenchymal transition leads to rapid and uniform MSC conversion of human pluripotent cells without the need for embryoid body formation or feeder cell coculture, providing a robust, clinically applicable, and efficient system for generating MSCs from human iPSCs.
AbstractList The translational potential of mesenchymal stem/stromal cells (MSCs) is limited by their rarity in somatic organs, heterogeneity, and need for harvest by invasive procedures. Induced pluripotent stem cells (iPSCs) could be an advantageous source of MSCs, but attempts to derive MSCs from pluripotent cells have required cumbersome or untranslatable techniques, such as coculture, physical manipulation, sorting, or viral transduction. We devised a single-step method to direct mesengenic differentiation of human embryonic stem cells (ESCs) and iPSCs using a small molecule inhibitor. First, epithelial-like monolayer cells were generated by culturing ESCs/iPSCs in serum-free medium containing the transforming growth factor-β pathway inhibitor SB431542. After 10 days, iPSCs showed upregulation of mesodermal genes (MSX2, NCAM, HOXA2) and downregulation of pluripotency genes (OCT4, LEFTY1/2). Differentiation was then completed by transferring cells into conventional MSC medium. The resultant development of MSC-like morphology was associated with increased expression of genes, reflecting epithelial-to-mesenchymal transition. Both ESC- and iPSC-derived MSCs exhibited a typical MSC immunophenotype, expressed high levels of vimentin and N-cadherin, and lacked expression of pluripotency markers at the protein level. Robust osteogenic and chondrogenic differentiation was induced in vitro in ES-MSCs and iPS-MSCs, whereas adipogenic differentiation was limited, as reported for primitive fetal MSCs and ES-MSCs derived by other methods. We conclude that treatment with SB431542 in two-dimensional cultures followed by culture-induced epithelial-to-mesenchymal transition leads to rapid and uniform MSC conversion of human pluripotent cells without the need for embryoid body formation or feeder cell coculture, providing a robust, clinically applicable, and efficient system for generating MSCs from human iPSCs.
The translational potential of mesenchymal stem/stromal cells (MSCs) is limited by their rarity in somatic organs, heterogeneity, and need for harvest by invasive procedures. Induced pluripotent stem cells (iPSCs) could be an advantageous source of MSCs, but attempts to derive MSCs from pluripotent cells have required cumbersome or untranslatable techniques, such as coculture, physical manipulation, sorting, or viral transduction. We devised a single-step method to direct mesengenic differentiation of human embryonic stem cells (ESCs) and iPSCs using a small molecule inhibitor. First, epithelial-like monolayer cells were generated by culturing ESCs/iPSCs in serum-free medium containing the transforming growth factor-β pathway inhibitor SB431542. After 10 days, iPSCs showed upregulation of mesodermal genes (MSX2, NCAM, HOXA2) and downregulation of pluripotency genes (OCT4, LEFTY1/2). Differentiation was then completed by transferring cells into conventional MSC medium. The resultant development of MSC-like morphology was associated with increased expression of genes, reflecting epithelial-to-mesenchymal transition. Both ESC- and iPSC-derived MSCs exhibited a typical MSC immunophenotype, expressed high levels of vimentin and N-cadherin, and lacked expression of pluripotency markers at the protein level. Robust osteogenic and chondrogenic differentiation was induced in vitro in ES-MSCs and iPS-MSCs, whereas adipogenic differentiation was limited, as reported for primitive fetal MSCs and ES-MSCs derived by other methods. We conclude that treatment with SB431542 in two-dimensional cultures followed by culture-induced epithelial-to-mesenchymal transition leads to rapid and uniform MSC conversion of human pluripotent cells without the need for embryoid body formation or feeder cell coculture, providing a robust, clinically applicable, and efficient system for generating MSCs from human iPSCs.The translational potential of mesenchymal stem/stromal cells (MSCs) is limited by their rarity in somatic organs, heterogeneity, and need for harvest by invasive procedures. Induced pluripotent stem cells (iPSCs) could be an advantageous source of MSCs, but attempts to derive MSCs from pluripotent cells have required cumbersome or untranslatable techniques, such as coculture, physical manipulation, sorting, or viral transduction. We devised a single-step method to direct mesengenic differentiation of human embryonic stem cells (ESCs) and iPSCs using a small molecule inhibitor. First, epithelial-like monolayer cells were generated by culturing ESCs/iPSCs in serum-free medium containing the transforming growth factor-β pathway inhibitor SB431542. After 10 days, iPSCs showed upregulation of mesodermal genes (MSX2, NCAM, HOXA2) and downregulation of pluripotency genes (OCT4, LEFTY1/2). Differentiation was then completed by transferring cells into conventional MSC medium. The resultant development of MSC-like morphology was associated with increased expression of genes, reflecting epithelial-to-mesenchymal transition. Both ESC- and iPSC-derived MSCs exhibited a typical MSC immunophenotype, expressed high levels of vimentin and N-cadherin, and lacked expression of pluripotency markers at the protein level. Robust osteogenic and chondrogenic differentiation was induced in vitro in ES-MSCs and iPS-MSCs, whereas adipogenic differentiation was limited, as reported for primitive fetal MSCs and ES-MSCs derived by other methods. We conclude that treatment with SB431542 in two-dimensional cultures followed by culture-induced epithelial-to-mesenchymal transition leads to rapid and uniform MSC conversion of human pluripotent cells without the need for embryoid body formation or feeder cell coculture, providing a robust, clinically applicable, and efficient system for generating MSCs from human iPSCs.
The translational potential of mesenchymal stem/stromal cells (MSCs) is limited by their rarity in somatic organs, heterogeneity, and need for harvest by invasive procedures. Induced pluripotent stem cells (iPSCs) could be an advantageous source of MSCs, but attempts to derive MSCs from pluripotent cells have required cumbersome or untranslatable techniques, such as coculture, physical manipulation, sorting, or viral transduction. We devised a single-step method to direct mesengenic differentiation of human embryonic stem cells (ESCs) and iPSCs using a small molecule inhibitor. First, epithelial-like monolayer cells were generated by culturing ESCs/iPSCs in serum-free medium containing the transforming growth factor-β pathway inhibitor SB431542. After 10 days, iPSCs showed upregulation of mesodermal genes ( MSX2, NCAM, HOXA2 ) and downregulation of pluripotency genes ( OCT4, LEFTY1/2 ). Differentiation was then completed by transferring cells into conventional MSC medium. The resultant development of MSC-like morphology was associated with increased expression of genes, reflecting epithelial-to-mesenchymal transition. Both ESC- and iPSC-derived MSCs exhibited a typical MSC immunophenotype, expressed high levels of vimentin and N-cadherin, and lacked expression of pluripotency markers at the protein level. Robust osteogenic and chondrogenic differentiation was induced in vitro in ES-MSCs and iPS-MSCs, whereas adipogenic differentiation was limited, as reported for primitive fetal MSCs and ES-MSCs derived by other methods. We conclude that treatment with SB431542 in two-dimensional cultures followed by culture-induced epithelial-to-mesenchymal transition leads to rapid and uniform MSC conversion of human pluripotent cells without the need for embryoid body formation or feeder cell coculture, providing a robust, clinically applicable, and efficient system for generating MSCs from human iPSCs.
Author Pelekanos, Rebecca A.
Ellis, Rebecca L.
Chen, Yen Shun
Horne, Rachel
Fisk, Nicholas M.
Wolvetang, Ernst J.
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  givenname: Rebecca L.
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  givenname: Nicholas M.
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BackLink https://www.ncbi.nlm.nih.gov/pubmed/23197756$$D View this record in MEDLINE/PubMed
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Snippet The translational potential of mesenchymal stem/stromal cells (MSCs) is limited by their rarity in somatic organs, heterogeneity, and need for harvest by...
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StartPage 83
SubjectTerms Antigens, CD - genetics
Antigens, CD - metabolism
Benzamides - pharmacology
Biomarkers - metabolism
Bone marrow
Cadherins
Cadherins - genetics
Cadherins - metabolism
Cell adhesion & migration
Cell culture
Cell Culture Techniques - methods
Cell Differentiation
Cell Proliferation
Cell Shape
Cells, Cultured
Culture Media, Serum-Free
Cytology
Data analysis
Differentiation
Dioxoles - pharmacology
Embryo cells
Embryonic stem cells
Embryonic Stem Cells - cytology
Embryonic Stem Cells - metabolism
Epithelial-Mesenchymal Transition
Fetuses
Fibroblasts
Gene Expression Regulation
Growth factors
Humans
Immunohistochemistry
Immunophenotyping
Induced pluripotent stem cells
Induced Pluripotent Stem Cells - cytology
Induced Pluripotent Stem Cells - drug effects
Induced Pluripotent Stem Cells - metabolism
Inhibitory postsynaptic potentials
Mesenchymal stem cells
Mesenchymal Stromal Cells - cytology
Mesenchymal Stromal Cells - metabolism
Mesenchyme
Mesoderm - cytology
Mesoderm - metabolism
Msx2 protein
Neural cell adhesion molecule
Oct-4 protein
Octamer Transcription Factor-3 - genetics
Octamer Transcription Factor-3 - metabolism
Original and Reviews
Pluripotency
Pluripotent stem cells
Regenerative Medicine - methods
Stem cells
Stromal cells
Time Factors
Transforming growth factor
Transforming growth factor-b
Vimentin
Vimentin - genetics
Vimentin - metabolism
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Title Small Molecule Mesengenic Induction of Human Induced Pluripotent Stem Cells to Generate Mesenchymal Stem/Stromal Cells
URI https://onlinelibrary.wiley.com/doi/abs/10.5966%2Fsctm.2011-0022
https://www.ncbi.nlm.nih.gov/pubmed/23197756
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