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. Author manuscript; available in PMC: 2016 Apr 1.
Published in final edited form as: Biomaterials. 2015 Jan 22;47:1–12. doi: 10.1016/j.biomaterials.2014.12.043

Figure 4. Validation of endothelial cell differentiation, and tube formation by CPCs was dependent on hydrogel stiffness and adhesion peptide density.

Figure 4

a, Representative confocal microscopy images of immunostaining of endothelial cell marker CD31, and acetylated low-density lipoprotein (Ac-LDL) uptake by the cells after 12 days of culture in HyA, HyA-P, HyA-PH and HyA-PHT hydrogels. HyA-P treated with an equivalent concentration of soluble TGFβ1 as delivered in HyA-PHT was included as a control. Both TGFβ1 and heparin were necessary in the HyA hydrogels to promote endothelial cell differentiation, as determined by CD31 expression and Ac-LDL uptake, and tubule formation in HyA-PHT hydrogel (850Pa modulus, 380μM bsp-RGD(15), 0.03wt% heparin, and 40nM TGFβ1). b, Vascular-like tube formation of CD31 positive cells in the HyA-PHT hydrogel was observed as a function of bsp-RGD(15) peptide density (120–380 μM) and gel modulus (15–850 Pa). RSM plots of the combined effects of peptide ligand and material modulus on c, total tube length, and d, average tube thickness. e, A confocal reconstruction of the 3D network structure of the CD31 positive cells and f, the appearance of a central lumen within the network structures suggested they were hollow nascent vessels, e.g. tubular structures.