Thrombotic occlusion of coronary and peripheral small-diameter (<6 mm) arteries is one of major causes of morbidity and mortality worldwide. The use of autologous saphenous veins as bypass grafts for occluded vessels are constrained by unavailability in patients with systemic vascular disease, and synthetic graft are also limited since the materials (e.g., ePTFE, Dacron) induce unnatural cell responses leading to thrombosis and graft stenosis. While tissue-engineered vascular conduits can potentially overcome these challenges, they currently fail to incorporate the biological and architectural components of vascular matrix due to inherently poor elastin synthesis by post-neonatal cells. Recently, we have shown that tetrameric fragments of hyaluronan (HA-o), a glycosaminoglycan in vascular extracellular matrix, and TGF-21 significantly upregulate elastin precursor synthesis and elastic matrix maturation by adult rat vascular smooth muscle cells (RASMCs). However, application of this method to engineer vascular constructs can benefit from (a) recruiting a young and potentially more inherently elastogenic source of autologous cells within a peritoneal cavity and (b) utilizing elastogenic factor (HA-o and TGF-21) functionalized electrospun meshes, with sub-micron diameter aligned polymer fibers, to further enhance elastin deposition and guide elastic fiber formation and alignment. Additional goals of developing intra-peritoneal autologous tissue are to provide components of a physiologic microenvironment for cells, while circumventing the need with in vitro bioreactors to isolate primary cells from biopsied healthy tissues and expand them in culture prior to cell seeding. Our objective is to produce a patent, elastic fiber rich small-diameter vascular graft by recruiting autologous peritoneal cells to an electrospun meshes tethered with elastogenic factors. The study will test four hypotheses: (a) the graft can be produced by recruiting rat peritoneal cells to an electrospun conduit, (b) HA-o and TGF- 21 tethered-meshes will prompt peritoneal cell differentiation into smooth muscle-like, though more elastogenic cells, (c) circumferentially oriented electrospun fibers will direct cellular deposition of aligned elastic fibers, and (d) grafts will remain viable when autologously implanted in rat abdominal aortae. These hypotheses will be tested with two specific aims.
Aim 1 A will investigate separately (a) the extent of aligned elastic fiber generation by RASMCs on non- functionalized meshes and (b) the density of HA-o and TGF-21 immobilized on 2-D films that prompts maximum elastic matrix synthesis and maturation.
Aim 1 B will determine the benefits of factor functionalized 3-D electrospun sheets, over functionalized films and non-functionalized electrospun sheets.
Aim 2 A, B will test (a) recruited intra-peritoneal cell differentiation into smooth muscle-like cells on an electrospun conduit and the time required to (b) generate an elastic-matrix rich graft replicating aortal structure and mechanics.
Aim 2 C will determine graft patency through autologous transplantation in a rat aorta. The significance of this proposal is in engineering vascular tissue replacements with elastic matrix similar to native tissue.
Coronary artery disease and diseases of other small arteries are one of major causes of death and disability worldwide, and the primary graft to bypass blood around blocked arteries (i.e other blood vessels from the patient) are frequently unavailable. Alternative grafts for surgical treatment have significant complications because they lack a component of a blood vessel that provides its rubbery properties, mature elastic fibers. The significance of this proposal lies in its combined use of factors (i.e. hyaluronan oligomers and transforming growth factor-21), a mesh with small synthetic fibers (i.e. an aligned electrospun mesh), and a patient's own body to develop vascular replacements with mature elastic fibers and properties similar to actual blood vessels. This should allow the graft to survive in the long-term during future large animal trials.
|Bashur, Chris A; Ramamurthi, Anand (2014) Composition of intraperitoneally implanted electrospun conduits modulates cellular elastic matrix generation. Acta Biomater 10:163-72|
|Bashur, Chris A; Eagleton, Matthew J; Ramamurthi, Anand (2013) Impact of electrospun conduit fiber diameter and enclosing pouch pore size on vascular constructs grown within rat peritoneal cavities. Tissue Eng Part A 19:809-23|