A Thromboresistant Cell-Derived Biomaterial Modification for Vascular Graft

Cardiovascular diseases are responsible for as many as 17.3 million deaths per year worldwide. In cases where arterial bypass is necessary, the most common surgical treatment involves the use of the patient's own saphenous vein. Unfortunately, individuals requiring such grafts often do not possess sufficiently healthy vessels for bypass, or, if vascular harvest is possible, the extraction of a graft will result in additional morbidity at the donor site. Despite the great need for alternatives to autologous arterial grafts, none are clinically available. While some promising alternatives are currently being tested as dialysis access vessels, failure due to thrombosis remains an obstacle, even in this relatively low-stakes application. This thesis examines the use of thrombospondin-2 knock out extracellular matrix (TSP2 KO ECM) as a nonthrombogenic modification for the lumen of vascular grafts and proposes a mechanism for its unique thromboresistance.;Previous reports have shown that mice lacking TSP2 possess irregular collagen fibril morphology and present with a bleeding diathesis. In the current work, we investigated the contribution of TSP2 KO extracellular matrix (ECM) to these defects. Bone marrow transplants and endothelial denudation studies revealed that an ECM defect heavily contributed to the bleeding diathesis. Moreover, we report that decellularized TSP2 KO ECM produced by dermal fibroblasts in vitro had decreased von Willebrand Factor (vWF) binding compared to WT ECM. This finding was probed via immunofluorescence after whole blood flow over ECM to determine quantity of vWF adhesion, and via atomic force microscopy to determine vWF binding force to ECM to determine the quality of vWF adhesion (53.85+/-8.44pN on WT ECM compared to 16.47+/-6.17pN on KO ECM). This unique property presents an opportunity to use TSP2 KO ECM as a hemocompatible modification to the lumen of vascular grafts, potentially circumventing thrombosis using simple biological processes.;Herein, we probed the feasibility and efficacy of TSP2 KO ECM modification of decellularized vascular grafts in vitro and in vivo. Tensile testing via 1NSTRON and suture strength analysis of decellularized aortas with and without ECM modification showed that ECM modification did not alter graft mechanical properties. Scanning electron microscopy and immunofluorescent studies clearly indicated that TSP2 KO ECM readily modifies the lumen of decellularized vascular grafts. Moreover, exposure of platelets to unmodified and ECM modified decellularized aortas and subsequent analysis via SEM and fluorescent staining revealed significantly decreased platelet adhesion and activation on TSP2 KO ECM modified grafts compared to unmodified and wild-type ECM modified controls (1.77+/-0.65% platelet coverage vs. 28.7+/-8.02% and 22.6+/-3.87%, respectively). In addition, implantation of TSP2 KO ECM coated vascular grafts in an infra-renal aortic interposition model resulted in increased endothelial and smooth muscle-like cell recruitment to the graft, and decreased the rate of graft failure.;Together, these data indicate that TSP2 plays a key role in the formation of collagen fibrils such that vWF is unable to adhere properly to them. This property may be leveraged to produce a non-thombogenic vascular graft modification that has the potential to aid in the creation of the ideal arterial graft.