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    VEGF BOUND TO HEPARIN-CONJUGATED, ELECTROSPUN FIBRIN MICROFIBER SUPPORTS ECM FORMATION AND ORIENTATION ON 3D, IN VITRO MICROVASCULAR MODEL

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    DAVENPORT-THESIS-2014.pdf (1.719Mb)
    Date
    2014-08-08
    Author
    Davenport, Matthew James
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    Abstract
    Current three dimensional (3D) models for microvasculature fail to understand the combinatorial role nanotopography, curvature, and bound growth factors have on extracellular matrix (ECM) deposition and cell alignment. Previous research shows that endothelial cell and ECM alignment is critical for managing shear stresses and contractile forces in vivo. We developed a novel, in vitro 3D model for microvasculature by using electrospun fibrin-alginate microfibers for endothelial cell proliferation and attachment with subsequent ECM deposition. Our preliminary studies showed that endothelial colony forming cells (ECFCs) cultured on our microfibers aligned in the direction of the stretched nanotopography. We found the ECM produced by ECFCs aligns perpendicular to the cell layer and circumferentially around the fibers themselves. The fiber diameter has a direct effect on ECM orientation, as fibers larger than 400 µm lead to random ECM orientations compared to the circumferentially wrapped ECM seen in 100 to 400 µm fibers. The endothelial layer is able to support the co-culture of vSMCs on our microfibers. Additionally, we successfully immobilized vascular endothelial growth factor (VEGF) on our fibrin system. We established a reductive amination protocol to conjugate fibrinogen to heparin, which is a known VEGF-binding molecule. The VEGF-bound fiber catalyzed ECM production without additional VEGF in solution, and also produced an ECM and cell orientation comparable to the stock fibrin microfibers. Summarily, we designed a dynamic, 3D fibrin-based platform for studying ECM deposition and cell interactions in-vitro in the presence of bound and unbound VEGF.
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    http://jhir.library.jhu.edu/handle/1774.2/38138
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