Nanotopographical cues augment mesenchymal differentiation of human embryonic stem cells.

The production of bone-forming osteogenic cells for research purposes or transplantation therapies remains a significant challenge. Using planar polycarbonate substrates lacking in topographical cues and substrates displaying a nanotopographical pattern, mesenchymal differentiation of human embryonic stem cells is directed in the absence of chemical factors and without induction of differentiation by embryoid body formation. Cells incubated on nanotopographical substrates show enhanced expression of mesenchymal or stromal markers and expression of early osteogenic progenitors at levels above those detected in cells on planar substrates in the same basal media. Evidence of epithelial-to-mesenchymal transition during substrate differentiation and DNA methylation changes akin to chemical induction are also observed. These studies provide a suitable approach to overcome regenerative medical challenges and describe a defined, reproducible platform for human embryonic stem cell differentiation.

[1]  F. Dell’Accio,et al.  Mesenchymal differentiation propensity of a human embryonic stem cell line , 2011, Cell proliferation.

[2]  E. Wolvetang,et al.  Small Molecule Mesengenic Induction of Human Induced Pluripotent Stem Cells to Generate Mesenchymal Stem/Stromal Cells , 2012, Stem cells translational medicine.

[3]  P. Stern,et al.  Epithelial-mesenchymal transition events during human embryonic stem cell differentiation. , 2007, Cancer research.

[4]  Charles P. Lin,et al.  Ex vivo glycan engineering of CD44 programs human multipotent mesenchymal stromal cell trafficking to bone , 2008, Nature Medicine.

[5]  R. Sherwood,et al.  Genetic targeting of the endoderm with claudin-6CreER , 2008, Developmental dynamics : an official publication of the American Association of Anatomists.

[6]  Reine Bareille,et al.  Altered nanofeature size dictates stem cell differentiation , 2012, Journal of Cell Science.

[7]  Laura A. Smith,et al.  The enhancement of human embryonic stem cell osteogenic differentiation with nano-fibrous scaffolding. , 2010, Biomaterials.

[8]  Sanjin Zvonic,et al.  Immunophenotype of Human Adipose‐Derived Cells: Temporal Changes in Stromal‐Associated and Stem Cell–Associated Markers , 2006, Stem cells.

[9]  J. Thomson,et al.  Embryonic stem cell lines derived from human blastocysts. , 1998, Science.

[10]  G. Karsenty,et al.  Osf2/Cbfa1: A Transcriptional Activator of Osteoblast Differentiation , 1997, Cell.

[11]  C. Wilkinson,et al.  The control of human mesenchymal cell differentiation using nanoscale symmetry and disorder. , 2007, Nature materials.

[12]  Rong Fan,et al.  Nanotopography influences adhesion, spreading, and self-renewal of human embryonic stem cells. , 2012, ACS nano.

[13]  A. G. Herreros,et al.  The transcription factor Snail is a repressor of E-cadherin gene expression in epithelial tumour cells , 2000, Nature Cell Biology.

[14]  Gavin Jell,et al.  Comparative materials differences revealed in engineered bone as a function of cell-specific differentiation. , 2009, Nature materials.

[15]  M. Fraga,et al.  The transcription factor Slug represses E-cadherin expression and induces epithelial to mesenchymal transitions: a comparison with Snail and E47 repressors , 2003, Journal of Cell Science.

[16]  Ali H. Brivanlou,et al.  Neural induction, the default model and embryonic stem cells , 2002, Nature Reviews Neuroscience.

[17]  K. Anseth,et al.  Small functional groups for controlled differentiation of hydrogel-encapsulated human mesenchymal stem cells. , 2008, Nature materials.

[18]  M. S. Kallos,et al.  Reduced differentiation efficiency of murine embryonic stem cells in stirred suspension bioreactors. , 2010, Stem cells and development.

[19]  Markus J. Buehler,et al.  Hierarchical Structure Controls Nanomechanical Properties of Vimentin Intermediate Filaments , 2009, PloS one.

[20]  Sungho Jin,et al.  Stem cell fate dictated solely by altered nanotube dimension , 2009, Proceedings of the National Academy of Sciences.

[21]  K. Mizuseki,et al.  Induction of Midbrain Dopaminergic Neurons from ES Cells by Stromal Cell–Derived Inducing Activity , 2000, Neuron.

[22]  Ya-jun Liu,et al.  Role of nucleostemin in growth regulation of gastric cancer, liver cancer and other malignancies. , 2004, World journal of gastroenterology.

[23]  D G Wilkinson,et al.  Control of cell behavior during vertebrate development by Slug, a zinc finger gene. , 1994, Science.

[24]  T. Takada,et al.  Transient suppression of PPARγ directed ES cells into an osteoblastic lineage , 2006 .

[25]  Yoshiakira Kanai,et al.  Depletion of definitive gut endoderm in Sox17-null mutant mice. , 2002, Development.

[26]  R. McKay,et al.  A nucleolar mechanism controlling cell proliferation in stem cells and cancer cells. , 2002, Genes & development.

[27]  J. Pitts,et al.  PPARgamma: observations in the hematopoietic system. , 2000, Prostaglandins & other lipid mediators.

[28]  Francisco Portillo,et al.  The transcription factor Snail controls epithelial–mesenchymal transitions by repressing E-cadherin expression , 2000, Nature Cell Biology.

[29]  D. van der Kooy,et al.  Embryonic stem cells assume a primitive neural stem cell fate in the absence of extrinsic influences , 2006, The Journal of cell biology.

[30]  Richard O C Oreffo,et al.  Bridging the regeneration gap: stem cells, biomaterials and clinical translation in bone tissue engineering. , 2008, Archives of biochemistry and biophysics.

[31]  Raymond H. W. Lam,et al.  Mechanics Regulates Fate Decisions of Human Embryonic Stem Cells , 2012, PloS one.

[32]  S. Gronthos,et al.  The STRO-1+ fraction of adult human bone marrow contains the osteogenic precursors. , 1994, Blood.

[33]  K. Mizuseki,et al.  Directed differentiation of telencephalic precursors from embryonic stem cells , 2005, Nature Neuroscience.

[34]  P. Robson,et al.  Osteogenic differentiation within intact human embryoid bodies result in a marked increase in osteocalcin secretion after 12 days of in vitro culture, and formation of morphologically distinct nodule-like structures. , 2005, Tissue & cell.

[35]  Raghu Kalluri,et al.  The epithelial–mesenchymal transition: new insights in signaling, development, and disease , 2006, The Journal of cell biology.

[36]  B. Herrmann,et al.  The T genes in embryogenesis. , 1994, Trends in genetics : TIG.

[37]  Derrick E Rancourt,et al.  Induction of chondro-, osteo- and adipogenesis in embryonic stem cells by bone morphogenetic protein-2: Effect of cofactors on differentiating lineages , 2005, BMC Developmental Biology.

[38]  P. Monk,et al.  STRO-1, HOP-26 (CD63), CD49a and SB-10 (CD166) as markers of primitive human marrow stromal cells and their more differentiated progeny: a comparative investigation in vitro , 2003, Cell and Tissue Research.

[39]  Ning Wang,et al.  Cell material property dictates stress-induced spreading and differentiation in embryonic stem cells , 2009, Nature materials.

[40]  S. Kanzaki,et al.  Secretion of osteocalcin and its propeptide from human osteoblastic cells: Dissociation of the secretory patterns of osteocalcin and its propeptide , 1993, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[41]  E. Kroon,et al.  Efficient differentiation of human embryonic stem cells to definitive endoderm , 2005, Nature Biotechnology.

[42]  T. Kudo,et al.  Localized phosphorylation of vimentin by Rho‐kinase in neuroblastoma N2a cells , 2000, Genes to cells : devoted to molecular & cellular mechanisms.

[43]  N. Gadegaard,et al.  Nanoscale surfaces for the long-term maintenance of mesenchymal stem cell phenotype and multipotency. , 2011, Nature materials.

[44]  Wojciech Wojakowski,et al.  Mobilization of bone marrow-derived Oct-4+ SSEA-4+ very small embryonic-like stem cells in patients with acute myocardial infarction. , 2009, Journal of the American College of Cardiology.

[45]  R. Perlingeiro,et al.  SSEA-4 identifies mesenchymal stem cells from bone marrow. , 2007, Blood.

[46]  Virginie Sottile,et al.  In vitro osteogenic differentiation of human ES cells. , 2003, Cloning and stem cells.

[47]  N. Allen,et al.  Neural differentiation of mouse embryonic stem cells in chemically defined medium , 2005, Brain Research Bulletin.

[48]  A. Hollander,et al.  Nucleostemin Is a Marker of Proliferating Stromal Stem Cells in Adult Human Bone Marrow , 2006, Stem cells.

[49]  G. Boland,et al.  Cross‐talk between Wnt signaling pathways in human mesenchymal stem cells leads to functional antagonism during osteogenic differentiation , 2007, Journal of cellular biochemistry.

[50]  Peng Wang,et al.  Enhancing osteogenic differentiation of mouse embryonic stem cells by nanofibers. , 2009, Tissue engineering. Part A.

[51]  S. Sen,et al.  Matrix Elasticity Directs Stem Cell Lineage Specification , 2006, Cell.

[52]  Judith M Curran,et al.  The guidance of human mesenchymal stem cell differentiation in vitro by controlled modifications to the cell substrate. , 2006, Biomaterials.

[53]  Milan Mrksich,et al.  Geometric cues for directing the differentiation of mesenchymal stem cells , 2010, Proceedings of the National Academy of Sciences.

[54]  B. Spiegelman,et al.  mPPAR gamma 2: tissue-specific regulator of an adipocyte enhancer. , 1994, Genes & development.

[55]  Janghwan Kim,et al.  Characterization of DNA methylation change in stem cell marker genes during differentiation of human embryonic stem cells. , 2007, Biochemical and biophysical research communications.

[56]  C. Semino,et al.  Osteogenic differentiation of mouse embryonic stem cells and mouse embryonic fibroblasts in a three-dimensional self-assembling peptide scaffold. , 2006, Tissue engineering.

[57]  K. Schenke-Layland,et al.  Mapping the first stages of mesoderm commitment during differentiation of human embryonic stem cells , 2010, Proceedings of the National Academy of Sciences.

[58]  E. Fearon,et al.  The SLUG zinc-finger protein represses E-cadherin in breast cancer. , 2002, Cancer research.

[59]  Ali Khademhosseini,et al.  Cultivation of Human Embryonic Stem Cells Without the Embryoid Body Step Enhances Osteogenesis In Vitro , 2006, Stem cells.

[60]  M. Wendel,et al.  Bone matrix formation in osteogenic cultures derived from human embryonic stem cells in vitro. , 2007, Stem cells and development.

[61]  J. Chan,et al.  Human First‐Trimester Fetal MSC Express Pluripotency Markers and Grow Faster and Have Longer Telomeres Than Adult MSC , 2007, Stem cells.

[62]  Christopher S. Chen,et al.  Cell shape, cytoskeletal tension, and RhoA regulate stem cell lineage commitment. , 2004, Developmental cell.

[63]  D. Rancourt,et al.  Microenvironment Modulates Osteogenic Cell Lineage Commitment in Differentiated Embryonic Stem Cells , 2010, PloS one.

[64]  Peter X Ma,et al.  The influence of three-dimensional nanofibrous scaffolds on the osteogenic differentiation of embryonic stem cells. , 2009, Biomaterials.

[65]  H. Kleinman,et al.  Osteonectin, a bone-specific protein linking mineral to collagen , 1981, Cell.

[66]  Nikolaj Gadegaard,et al.  Using nanotopography and metabolomics to identify biochemical effectors of multipotency. , 2012, ACS nano.