Biodegradable scaffolds seeded with bone marrow mononuclear cells (BMCs) are the earliest tissue-engineered vascular grafts (TEVGs) to be used clinically. These TEVGs transform into living blood vessels in vivo, with an endothelial cell (EC) lining invested by smooth muscle cells (SMCs); however, the process by which this occurs is unclear. To test if the seeded BMCs differentiate into the mature vascular cells of the neovessel, we implanted an immunodeficient mouse recipient with human BMC (hBMC)-seeded scaffolds. As in humans, TEVGs implanted in a mouse host as venous interposition grafts gradually transformed into living blood vessels over a 6-month time course. Seeded hBMCs, however, were no longer detectable within a few days of implantation. Instead, scaffolds were initially repopulated by mouse monocytes and subsequently repopulated by mouse SMCs and ECs. Seeded BMCs secreted significant amounts of monocyte chemoattractant protein-1 and increased early monocyte recruitment. These findings suggest TEVGs transform into functional neovessels via an inflammatory process of vascular remodeling.bone marrow | monocyte chemoattractant protein-1 | tissue engineering | neovascularization C ongenital heart disease is a leading cause of infant mortality, often requiring early surgical intervention to correct fatal cardiovascular malformations. Prosthetic vascular grafts are widely used in these reconstructive operations, but revisions are often necessary because of their inability to grow or effectively remodel within a growing child (1-3). A strategy to address this issue is the use of living tissue-engineered vascular grafts (TEVGs). Constructed from biodegradable polyester tubes seeded with autologous bone marrow mononuclear cells (BMCs), these grafts undergo extensive remodeling in animal recipients and appear to transform into living blood vessels, similar in morphology and function to the native veins into which they are interposed (4, 5). Ongoing clinical studies evaluating BMC-seeded grafts as venous conduits for congenital heart surgery report excellent safety profiles and 100% patency rates at 1-3 years of follow-up (6-8). Additionally, these grafts demonstrate growth potential, suggesting they may be more effective for the pediatric patient population than currently available vascular grafts (8,9).Although the functional efficacy and clinical utility of TEVGs are promising, little is known about how these BMC-seeded polyester tubes transform into living blood vessels in host recipients. It has been proposed that stem cells within the seeded BMC population differentiate into the endothelial cells (ECs) and smooth muscle cells (SMCs) of the developing neovessel, ultimately replacing the degrading polyester tube (10). This hypothesis, however, has not been directly examined.We recently developed a method for constructing small-diameter biodegradable synthetic scaffolds suitable for use as vascular grafts in mice (11). These tubular scaffolds are composed of the same materials and design used in clinical TEVGs...

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Tissue engineered vascular grafts (TEVGs) have the potential to overcome the issues faced by existing small diameter prosthetic grafts by providing a biodegradable scaffold where the patient’s own cells can engraft and form functional neotissue. However, applying classical approaches to create arterial TEVGs using slow degrading materials with supraphysiological mechanical properties, typically results in limited host cell infiltration, poor remodeling, stenosis, and calcification. The purpose of this study is to evaluate the feasibility of novel small diameter arterial TEVGs created using fast degrading material. A 1.0mm and 5.0mm diameter TEVGs were fabricated with electrospun polycaprolactone (PCL) and chitosan (CS) blend nanofibers. The 1.0mm TEVGs were implanted in mice (n = 3) as an unseeded infrarenal abdominal aorta interposition conduit., The 5.0mm TEVGs were implanted in sheep (n = 6) as an unseeded carotid artery (CA) interposition conduit. Mice were followed with ultrasound and sacrificed at 6 months. All 1.0mm TEVGs remained patent without evidence of thrombosis or aneurysm formation. Based on small animal outcomes, sheep were followed with ultrasound and sacrificed at 6 months for histological and mechanical analysis. There was no aneurysm formation or calcification in the TEVGs. 4 out of 6 grafts (67%) were patent. After 6 months in vivo, 9.1 ± 5.4% remained of the original scaffold. Histological analysis of patent grafts demonstrated deposition of extracellular matrix constituents including elastin and collagen production, as well as endothelialization and organized contractile smooth muscle cells, similar to that of native CA. The mechanical properties of TEVGs were comparable to native CA. There was a significant positive correlation between TEVG wall thickness and CD68+ macrophage infiltration into the scaffold (R2 = 0.95, p = 0.001). The fast degradation of CS in our novel TEVG promoted excellent cellular infiltration and neotissue formation without calcification or aneurysm. Modulating host macrophage infiltration into the scaffold is a key to reducing excessive neotissue formation and stenosis.

Bone marrow stromal cells (MSCs) are a promising cell source for a variety of tissue engineering applications, given their ready availability and ability to differentiate into multiple cell lineages. MSCs have been successfully used to create neotissue for cardiovascular, urological, and orthopedic reconstructive surgical procedures in preclinical studies. The ability to optimize seeding techniques of MSCs onto tissue engineering scaffolds and the ability to control neotissue formation in vitro will be important for the rational design of future tissue engineering applications using MSCs. In this study we investigated the effect of centrifugal force on seeding MSCs into a biodegradable polyester scaffold. MSCs were isolated and seeded onto porous scaffold sections composed of nonwoven polyglycolic acid mesh coated with poly(L-lactide-co-epsilon-caprolactone). Compared to standard static seeding techniques, centrifugal seeding increased the seeding efficiency by 38% (p < 0.007) and significantly improved cellular distribution throughout the scaffold. Overall, centrifugal seeding of MSCs enhances seeding efficiency and improves cellular penetration into scaffolds, making it a potentially useful technique for manipulating neotissue formation by MSCs for tissue engineering applications.

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