Enhanced differentiation of human embryonic stem cells to mesenchymal progenitors by inhibition of TGF-β/activin/nodal signaling using SB-431542

Mahmood, Amer, Harkness, Linda, Schroder, Henrik Daa, Abdallah, Basem M. and Kassem, Moustapha (2010) Enhanced differentiation of human embryonic stem cells to mesenchymal progenitors by inhibition of TGF-β/activin/nodal signaling using SB-431542. Journal of Bone and Mineral Research, 25 6: 1216-1233. doi:10.1002/jbmr.34


Author Mahmood, Amer
Harkness, Linda
Schroder, Henrik Daa
Abdallah, Basem M.
Kassem, Moustapha
Title Enhanced differentiation of human embryonic stem cells to mesenchymal progenitors by inhibition of TGF-β/activin/nodal signaling using SB-431542
Journal name Journal of Bone and Mineral Research   Check publisher's open access policy
ISSN 0884-0431
1523-4681
Publication date 2010-01-01
Year available 2010
Sub-type Article (original research)
DOI 10.1002/jbmr.34
Open Access Status
Volume 25
Issue 6
Start page 1216
End page 1233
Total pages 18
Place of publication Hoboken, NJ, United States
Publisher Wiley-Blackwell Publishing
Language eng
Abstract Directing differentiation of human embryonic stem cells (hESCs) into specific cell types using an easy and reproducible protocol is a prerequisite for the clinical use of hESCs in regenerative-medicine procedures. Here, we report a protocol for directing the differentiation of hESCs into mesenchymal progenitor cells. We demonstrate that inhibition of transforming growth factor β (TGF-β)/activin/nodal signaling during embryoid body (EB) formation using SB-431542 (SB) in serum-free medium markedly upregulated paraxial mesodermal markers (TBX6, TBX5) and several myogenic developmental markers, including early myogenic transcriptional factors (Myf5, Pax7), as well as myocyte-committed markers [NCAM, CD34, desmin, MHC (fast), α-smooth muscle actin, Nkx2.5, cTNT]. Continuous inhibition of TGF-β signaling in EB outgrowth cultures (SB-OG) enriched for myocyte progenitor cells; markers were PAX7 + (25%), MYOD1 + (52%), and NCAM + (CD56) (73%). DNA microarray analysis revealed differential upregulation of 117 genes (>2-fold compared with control cells) annotated to myogenic development and function. Moreover, these cells showed the ability to contract (80% of the population) and formed myofibers when implanted intramuscularly in vivo. Interestingly, SB-OG cells cultured in 10% fetal bovine serum (FBS) developed into a homogeneous population of mesenchymal progenitors that expressed CD markers characteristic of mesenchymal stem cells (MSCs): CD44 + (100%), CD73 + (98%), CD146 + (96%), and CD166 + (88%) with the ability to differentiate into osteoblasts, adipocytes, and chondrocytes in vitro and in vivo. Furthermore, microarray analysis of these cells revealed downregulation of genes related to myogenesis: MYH3 (-167.9-fold), ACTA1 (-161-fold), MYBPH (-139-fold), ACTC (-100.3-fold), MYH8 (-45.5-fold), and MYOT (-41.8-fold) and marked upregulation of genes related to mesoderm-derived cell lineages. In conclusion, our data provides a simple and versatile protocol for directing the differentiation of hESCs into a myogenic lineage and then further into mesenchymal progenitors by blocking the TGF-β signaling pathway.
Keyword Human embryonic stem cells
Mesenchymal differentiation
Osteoblast differentiation
Skeletal myoblasts
TGF-β signaling
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status Non-UQ

Document type: Journal Article
Sub-type: Article (original research)
Collection: Australian Institute for Bioengineering and Nanotechnology Publications
 
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Created: Thu, 24 Sep 2015, 01:35:35 EST by Linda Harkness on behalf of Scholarly Communication and Digitisation Service