Tissue-engineering scaffolds were designed to mimic several features of the extracellular matrix using a combination of the synthetic polymer, PEG diacrylate, and bioactive factors. Scaffolds were formed by exposing aqueous solutions of PEG diacrylate and bioactive factors modified with PEG monoacrylate to ultraviolet or visible light in the presence of a suitable photoinitiator. Light exposure generated free radicals that targeted acrylate groups in the monomer and in PEG conjugated bioactive factors resulting in crosslinked hydrogel scaffolds with bioactive factors covalently incorporated. This study extended the capability for directing cell behavior using PEG-based hydrogels to include control over the spatial distribution of bioactive factors and the presentation of the growth factor, bFGF. Additionally, PEG hydrogels were modified with a degradable peptide sequence to enable cells to remodel the scaffold by secreting matrix metalloproteinases (MMPs). A continuous linear gradient was formed by simultaneously using a gradient maker to combine hydrogel precursor solutions with photopolymerization, which locks the gradient in place. Coomassie blue staining confirmed the formation of protein gradients. Fibroblast cells responded to covalently immobilized gradients of the adhesive peptide, RGD, by changing their morphology to align in the direction of increasing RGD concentration and by migrating differentially on RGD-gradient hydrogels compared to control hydrogels. Next, bFGF was covalently immobilized to hydrogels with retention of its mitogenic and chemotactic effects on smooth muscle cells (SMCs). A covalently immobilized bFGF gradient was also formed using the gradient maker and shown to increase linearly along the hydrogel's length by silver staining. SMCs responded to these bFGF-gradient hydrogels by aligning in the direction of increasing bFGF concentration and by migrating differentially, up the concentration gradient, compared to migration on control hydrogels. Finally, the MMP-sensitive peptide sequence, GPQGILGQ, was inserted into the main polymer chain's backbone to allow targeted degradation by cell-secreted proteases. Cells were observed to change their morphology and migrate when seeded within these degradable hydrogel scaffolds, but not in scaffolds lacking this degradable peptide sequence. This hydrogel system is expected to be useful for studying tissue formation leading eventually to an improved understanding of the factors needed to form engineered tissues
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