Abstract 3057: Tissue Engineered Three-layered Artery Using Hemodynamically-equivalent Pulsatile Bioreactor

Abstract

Introduction: Due to lack of anti-thrombogenicity in small diameter vascular prostheses, there is a strong demand for tissue-engineering autologous small-diameter graft, which can function in arterial high pressure and flow environments. Here, we investigated a potential to engineer a robust and three-layered artery. Methods: Endothelial cells (ECs), smooth muscle cells (SMCs) and fibroblasts were isolated with collagenase from bovine artery, then each cell were primary cultured. A polyglycolic acid (PGA) sheet seeded with SMCs was wrapped on silicon tube with a diameter of 6 mm. After 4 days of incubation, a polycaprolactone sheet seeded with SMCs was over-wrapped. Then, after incubation for another 3 weeks, a PGA sheet seeded with fibroblasts was over-wrapped and incubated for another 2 days. The luminal tube was then removed out, and the tubular tissue was mounted on a specially designed chamber. The luminal areas of engineered tubular structure were seeded with half a million of ECs and incubated for another 2days. Finally, bioreactor culture was performed for more 2weeks (n=10). Pulsatile flow and pressure were gradually increased from mean flow rate of 0.2(0.5/0) L/min and mean pressure of 20(40/15) mmHg to 0.6(1.4/0.2) L/min and 100(120/80) mmHg. Results: Scanning electron microscopic examination and von Willebrand factor stain revealed that ECs lined over the lumen and oriented by flow stimulation. Stain of Elastica Van Gieson and Masson Tricrome showed that a lot of elastin components and collagen were produced in the tissue engineered grafts. Stain for α-SMA, calponin, and vimentin showed that engineered arteries were constructed from SMC and fibroblasts regions and morphologically organized as three layered. Tensile tests demonstrated that ultimate strength of engineered arteries were increased by 6.7 times as compared with that of biodegradable polymers (n=8) and become equivalent to that of the native arteries (Native: 882±133kPa, Engineered: 827±155kPa) (each n=8). Conclusion: A robust and three-layered small-diameter elastic artery was engineered from vascular cells and biodegradable polymers using the hemodynamically-equivalent pulsatile bioreactor.