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EMBRYONIC STEM CELLS/INDUCED PLURIPOTENT STEM CELLS

A Developmentally Regulated Heparan Sulfate Epitope Defines a Subpopulation with Increased Blood Potential During Mesodermal Differentiation

^aUniversity of Manchester and Cancer Research UK Department of Medical Oncology, Manchester, United Kingdom;
^bDepartment of Biochemistry, Nijmegen Center for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands;
^cStem Cell Biology, Paterson Institute for Cancer Research, Manchester, United Kingdom;
^dSchool of Materials, University of Manchester, Manchester, United Kingdom

Key Words. Heparan sulfate • Hemangioblast • Embryonic stem cell • HS4C3 • Mesoderm • Hematopoiesis

Correspondence: Correspondence: Catherine Merry, Ph.D., School of Materials, University of Manchester, Manchester M1 7HS, U.K. Telephone: 0161-306-8871; Fax: 0161-306-3586; e-mail: catherine.merry{at}manchester.ac.uk

Received on March 26, 2008; accepted for publication on August 28, 2008.

First published online in STEM CELLS EXPRESS  September 11, 2008.

Heparan sulfate (HS) is a mandatory coreceptor for many growth factors and morphogens involved in embryonic development; its bioactivity is dictated by complex sulfation motifs embedded within the polymer chain. Using a panel of HS-specific antibodies we have identified a unique HS epitope recognized by antibody HS4C3 that is selectively expressed during differentiation of embryonic stem (ES) cells along the mesodermal lineage to the hemangioblast stage. The appearance of this high-affinity HS4C3-binding (HS4C3^high) epitope is transient; the epitope is specifically expressed within the emerging Brachyury⁺ (Bry⁺) population and marks those cells that will become fetal liver kinase 1 (Flk1)⁺. Fluorescence-activated cell sorting (FACS) separation and colony forming assays revealed that HS4C3^high/Flk1⁺ cells have a dramatically increased potential to form both blast and endothelial colonies, both of which depend upon the HS-binding growth factor vascular endothelial growth factor. Critically, expression of this HS epitope is tightly regulated, disappearing from the cell surface as the resultant hematopoietic lineages mature, in a similar manner to protein markers Bry and Flk1. In vivo studies showed a remarkable correlation with in vitro findings, with expression of HS4C3-binding epitopes restricted to newly formed mesodermal tissues during gastrulation. We believe this is the first time a defined HS epitope has been implicated in a specific developmental pathway and that this provides, in addition, a novel enrichment technique for the isolation of hemangioblasts from mixed differentiated ES cell cultures.

Disclosure of potential conflicts of interest is found at the end of this article.