Immobilization of Heparan Sulfate on Electrospun Meshes to Support Embryonic Stem Cell Culture and Differentiation *

Toin H Van Kuppevelt,Kate A Meade,Andrew Marson,Anthony J Day,Kathryn J White,Catherine L.R Merry,Rebecca J Holley,Claire E Pickford,Jason D Whittle,Donna Tillotson

doi:10.1074/jbc.m112.423012

Abstract

As our understanding of what guides the behavior of multi- and pluripotent stem cells deepens, so too does our ability to utilize certain cues to manipulate their behavior and maximize their therapeutic potential. Engineered, biologically functionalized materials have the capacity to influence stem cell behavior through a powerful combination of biological, mechanical, and topographical cues. Here, we present the development of a novel electrospun scaffold, functionalized with glycosaminoglycans (GAGs) ionically immobilized onto the fiber surface. Bound GAGs retained the ability to interact with GAG-binding molecules and, crucially, presented GAG sulfation motifs fundamental to mediating stem cell behavior. Bound GAG proved to be biologically active, rescuing the neural differentiation capacity of heparan sulfate-deficient mouse embryonic stem cells and functioning in concert with FGF4 to facilitate the formation of extensive neural processes across the scaffold surface. The combination of GAGs with electrospun scaffolds creates a biomaterial with potent applicability for the propagation and effective differentiation of pluripotent stem cells.

Highlights

Glycosaminoglycans influence stem cell fate but their combination with biomaterials remains to be optimized
Electrospun Microfiber Meshes Have Similar Dimensions to the Fibrous Components of Natural extracellular matrix (ECM)—Electrospinning of poly(lacticco-glycolic acid) (PLGA) created a reproducible scaffold with fiber diameter predominantly between 0.1–1.2 ␮m (Fig. 1A), dimensions that mimic those within the natural ECM [34]. plasma polymerization of allylamine (ppAm) did not alter average fiber diameter or morphology (Fig. 1B) with no discernible difference between the fiber diameter distribution of coated and uncoated microfiber scaffolds
Allylamine-coating Technology Is Transferrable to a Threedimensional Scaffold—Scaffolds coated with ppAm were more hydrophilic than uncoated microfiber meshes, as determined by water contact analysis

Summary

Introduction

Glycosaminoglycans influence stem cell fate but their combination with biomaterials remains to be optimized. Results: GAG bound to scaffolds presented essential sulfation epitopes and proved biologically active. Engineered, biologically functionalized materials have the capacity to influence stem cell behavior through a powerful combination of biological, mechanical, and topographical cues. Bound GAGs retained the ability to interact with GAG-binding molecules and, crucially, presented GAG sulfation motifs fundamental to mediating stem cell behavior. Bound GAG proved to be biologically active, rescuing the neural differentiation capacity of heparan sulfate-deficient mouse embryonic stem cells and functioning in concert with FGF4 to facilitate the formation of extensive neural processes across the scaffold surface. The combination of GAGs with electrospun scaffolds creates a biomaterial with potent applicability for the propagation and effective differentiation of pluripotent stem cells

Objectives

Results

Conclusion