טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentNseir Nora
SubjectEngineering Vascular-Like Conduit Embedded in
a Pre-Vascularized Bulk
DepartmentDepartment of Mechanical Engineering
Supervisors Professor Eyal Zussman
Professor Shulamit Levenberg
Full Thesis textFull thesis text - English Version


Abstract


Innate properties supportive of oxygen and nutrient perfusion are critical when designing engineering vascularized bulk tissue constructs. In order to induce vascularization (angiogenesis), a naturally occurring process in which new blood vessels sprout from preexisting ones in vivo, we aimed to generate a vascular-like conduit embedded in a vascularized bulk surrounding in vitro. Tubular albumin scaffolds were embedded in a synthetic polymer PLLA/PLGA bulk graft. The albumin layer was to serve as a template for evolving blood vessel structures. The PLLA/PLGA graft was salt-leached to a porosity of approximately 90%, with a pore size range of 200-600 μm. The inner albumin layer of an average thickness of 100 ?m was fabricated using an electrospinning technique. The morphology and mechanical features of the resultant scaffold were characterized. Comparative biocompatibility analyses were performed on albumin fibers both in vitro and in vivo. Several multi-cellular cultures, including fibroblasts and endothelial cells, were seeded on the designed scaffolds. Scaffold fate was also assessed in a specially designed bioreactor system, which allowed for application of pulsatile perfusion conditions, mimicking blood perfusion. Finally, the impact of the scaffold design on cellular organization was comprehensively examined, under both static and dynamic conditions. Wet albumin fibers resembled soft ECM fibers, e.g. elastin, with mechanical properties such as stiffness of 0.006 GPa and 1.1% extensibility. Albumin biodegradability (>50%) was demonstrated within three weeks of implantation, and paralleled rapid and thick fibrosis formation, characteristic of a late stage of wound healing. Cells demonstrated strong affinity to the albumin layer, seemingly mediated by albumin binding proteins (ABP) or by its negatively charged fibers. The tubular scaffolds guided the formation of blood vessel-like structures comprised of fibroblasts lined with endothelial cells (ECs). The fibroblast layer differentiated into smooth muscle-like cells, resembling the native blood vessel multilayered configuration. In addition, ECs organized into small lumen-like structures throughout the bulk surrounding, creating a pre-vascularized bulk projected to promote vascularization, when implanted in vivo. It was further demonstrated, that dynamic conditions fostered homogenous cell spreading and increased proliferation rates throughout the bulk, as well as the formation of a packed cell-dense area within the tube lumen, when compared to static conditions. The described construct provides an extra vascular supply to engineered tissues and is expected to facilitate tissue development and maintenance of their functions, in vitro. In addition, the developed cell-seeded scaffold can be further utilized as a model for studying angiogenesis in vitro.