Matrix Remodeling Maintains Embryonic Stem Cell Self‐Renewal by Activating Stat3

While a variety of natural and synthetic matrices have been used to influence embryonic stem cell (ESC) self‐renewal or differentiation, and ESCs also deposit a rich matrix of their own, the mechanisms behind how extracellular matrix affects cell fate are largely unexplored. The ESC matrix is continuously remodeled by matrix metalloproteinases (MMPs), a process that we find is enhanced by the presence of mouse embryonic fibroblast feeders in a paracrine manner. Matrix remodeling by MMPs aids in the self‐renewal of ESCs, as inhibition of MMPs inhibits the ability of ESCs to self‐renew. We also find that addition of the interstitial collagenase MMP1 is sufficient to maintain long‐term leukemia inhibitory factor (LIF)‐independent mouse ESC (mESC) self‐renewal in a dose‐dependent manner. This remarkable ability is due to the presence of endogenously produced self‐renewal‐inducing signals, including the LIF‐family ligand ciliary neurotrophic factor, that are normally trapped within the ECM and become exposed upon MMP‐induced matrix remodeling to signal through JAK and Stat3. These results uncover a new role for feeder cells in maintaining self‐renewal and show that mESCs normally produce sufficient levels of autocrine‐acting pro‐self‐renewal ligands. STEM Cells 2013;31:1097–1106

[1]  Donald Metcalf,et al.  Myeloid leukaemia inhibitory factor maintains the developmental potential of embryonic stem cells , 1988, Nature.

[2]  John K. Heath,et al.  Inhibition of pluripotential embryonic stem cell differentiation by purified polypeptides , 1988, Nature.

[3]  L. Matrisian,et al.  Metalloproteinases and their inhibitors in matrix remodeling. , 1990, Trends in genetics : TIG.

[4]  G. Yancopoulos,et al.  Ciliary neurotrophic factor maintains the pluripotentiality of embryonic stem cells. , 1993, Development.

[5]  J. Nichols,et al.  Maintenance of the pluripotential phenotype of embryonic stem cells through direct activation of gp130 signalling pathways , 1994, Mechanisms of Development.

[6]  B. Schryver,et al.  Properties of Wnt-1 protein that enable cell surface association. , 1996, Oncogene.

[7]  M. Flannery,et al.  Expression and function of matrix metalloproteinases and their inhibitors at the maternal-embryonic boundary during mouse embryo implantation. , 1996, Development.

[8]  Jussi Taipale,et al.  Growth factors in the extracellular matrix , 1997, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[9]  Thiennu H. Vu,et al.  Matrix metalloproteinases: effectors of development and normal physiology. , 2000, Genes & development.

[10]  D. Lauffenburger,et al.  Leukemia inhibitory factor (LIF) concentration modulates embryonic stem cell self-renewal and differentiation independently of proliferation. , 2000, Biotechnology and bioengineering.

[11]  R. Jaenisch,et al.  Hybrid vigor, fetal overgrowth, and viability of mice derived by nuclear cloning and tetraploid embryo complementation , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[12]  S. Rafii,et al.  Recruitment of Stem and Progenitor Cells from the Bone Marrow Niche Requires MMP-9 Mediated Release of Kit-Ligand , 2002, Cell.

[13]  J. Nichols,et al.  BMP Induction of Id Proteins Suppresses Differentiation and Sustains Embryonic Stem Cell Self-Renewal in Collaboration with STAT3 , 2003, Cell.

[14]  D. Page,et al.  Dppa3 / Pgc7 / stella is a maternal factor and is not required for germ cell specification in mice , 2004, BMC Developmental Biology.

[15]  P. Heinrich,et al.  Principles of interleukin (IL)-6-type cytokine signalling and its regulation. , 2003, The Biochemical journal.

[16]  Eun-sook Kim,et al.  TGF-beta-induced upregulation of MMP-2 and MMP-9 depends on p38 MAPK, but not ERK signaling in MCF10A human breast epithelial cells. , 2004, International journal of oncology.

[17]  D. Scadden,et al.  The stem-cell niche as an entity of action , 2006, Nature.

[18]  C. Streuli.  Faculty Opinions recommendation of Beta 4 integrin amplifies ErbB2 signaling to promote mammary tumorigenesis. , 2006 .

[19]  B. Graham-Evans,et al.  Murine Embryonic Stem Cells Secrete Cytokines/Growth Modulators That Enhance Cell Survival/Anti‐Apoptosis and Stimulate Colony Formation of Murine Hematopoietic Progenitor Cells , 2006, Stem cells.

[20]  G. Inghirami,et al.  β4 Integrin Amplifies ErbB2 Signaling to Promote Mammary Tumorigenesis , 2006, Cell.

[21]  S. Yamanaka,et al.  Induction of Pluripotent Stem Cells from Mouse Embryonic and Adult Fibroblast Cultures by Defined Factors , 2006, Cell.

[22]  R. Jaenisch,et al.  In vitro reprogramming of fibroblasts into a pluripotent ES-cell-like state , 2007, Nature.

[23]  M. Trotter,et al.  Derivation of pluripotent epiblast stem cells from mammalian embryos , 2007, Nature.

[24]  R. McKay,et al.  New cell lines from mouse epiblast share defining features with human embryonic stem cells , 2007, Nature.

[25]  P. Zandstra,et al.  LIF‐mediated control of embryonic stem cell self‐renewal emerges due to an autoregulatory loop , 2007, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[26]  P. Gray,et al.  Identification of potential pluripotency determinants for human embryonic stem cells following proteomic analysis of human and mouse fibroblast conditioned media. , 2007, Journal of proteome research.

[27]  T. Ichisaka,et al.  Generation of germline-competent induced pluripotent stem cells , 2007, Nature.

[28]  N. D. Clarke,et al.  Integration of External Signaling Pathways with the Core Transcriptional Network in Embryonic Stem Cells , 2008, Cell.

[29]  R. Nusse,et al.  Wnt signaling mediates self-organization and axis formation in embryoid bodies. , 2008, Cell stem cell.

[30]  D. Discher,et al.  Matrix Elasticity, Cytoskeletal Tension, and TGF-β: The Insoluble and Soluble Meet , 2008, Science Signaling.

[31]  K. Hochedlinger,et al.  Defining molecular cornerstones during fibroblast to iPS cell reprogramming in mouse. , 2008, Cell stem cell.

[32]  M. Singer,et al.  Culture of mouse embryonic stem cells. , 2008, Current protocols in stem cell biology.

[33]  B. Doble,et al.  The ground state of embryonic stem cell self-renewal , 2008, Nature.

[34]  Rudolf Jaenisch,et al.  Analysis of histone 2B-GFP retention reveals slowly cycling hematopoietic stem cells , 2009, Nature Biotechnology.

[35]  Richard O. Hynes,et al.  The Extracellular Matrix: Not Just Pretty Fibrils , 2009, Science.

[36]  J. Nichols,et al.  Klf4 reverts developmentally programmed restriction of ground state pluripotency , 2009, Development.

[37]  Herbert Schulz,et al.  Novel STAT3 Target Genes Exert Distinct Roles in the Inhibition of Mesoderm and Endoderm Differentiation in Cooperation with Nanog , 2009, Stem cells.

[38]  Ning Wang,et al.  Soft Substrates Promote Homogeneous Self-Renewal of Embryonic Stem Cells via Downregulating Cell-Matrix Tractions , 2010, PloS one.

[39]  R. Nusse,et al.  A study on the interactions between heparan sulfate proteoglycans and Wnt proteins , 2009, Developmental dynamics : an official publication of the American Association of Anatomists.

[40]  J. Rossant,et al.  Heparan Sulfation–Dependent Fibroblast Growth Factor Signaling Maintains Embryonic Stem Cells Primed for Differentiation in a Heterogeneous State , 2009, Stem cells.

[41]  Ying Jin,et al.  Calcineurin-NFAT signaling critically regulates early lineage specification in mouse embryonic stem cells and embryos. , 2011, Cell stem cell.

[42]  H. Hirai,et al.  Regulation of embryonic stem cell self-renewal and pluripotency by leukaemia inhibitory factor. , 2011, The Biochemical journal.

[43]  T. Blauwkamp,et al.  Embryonic stem cells require Wnt proteins to prevent differentiation to epiblast stem cells , 2011, Nature Cell Biology.

[44]  Albert J. Keung,et al.  Rho GTPases Mediate the Mechanosensitive Lineage Commitment of Neural Stem Cells , 2011, Stem cells.

[45]  J. Gutkind,et al.  Modulation of canonical Wnt signaling by the extracellular matrix component biglycan , 2011, Proceedings of the National Academy of Sciences.

[46]  M. Wolfe,et al.  Self‐Renewal Versus Lineage Commitment of Embryonic Stem Cells: Protein Kinase C Signaling Shifts the Balance , 2011, Stem cells.

[47]  S. Kahl,et al.  Quantitative and semiquantitative immunoassay of growth factors and cytokines in the conditioned medium of STO and CF-1 mouse feeder cells , 2011, In Vitro Cellular & Developmental Biology - Animal.

[48]  Joel Voldman,et al.  Attenuation of extrinsic signaling reveals the importance of matrix remodeling on maintenance of embryonic stem cell self-renewal , 2012, Proceedings of the National Academy of Sciences.

[49]  S. Roy,et al.  ETS-1 Protein Regulates Vascular Endothelial Growth Factor-induced Matrix Metalloproteinase-9 and Matrix Metalloproteinase-13 Expression in Human Ovarian Carcinoma Cell Line SKOV-3* , 2012, The Journal of Biological Chemistry.

[50]  C. Merry,et al.  E‐cadherin and, in Its Absence, N‐cadherin Promotes Nanog Expression in Mouse Embryonic Stem Cells via STAT3 Phosphorylation , 2012, Stem cells.