Abstract 580: The Production of a Bio-Engineered Endothelial Intima From Cultured Cells Using Whole Cardiac Cadaveric Extracellular Matrix.
Background; Bioengineered solutions to failing cardiac tissue have been difficult to achieve due partially to adverse interactions between circulating blood and the engineered surface. The aim of this study was to determine if by using naturally-derived ECM and cultured endothelial cells, a bioengineered whole-heart vascular intima could be generated. The matrix substrate for organ culture was produced by a perfusion-based detergent decelluarization of cadaveric rat heart. This process maintained ECM protein integrity as indicated by a glycosaminoglycan assay, with ~ equivalent amounts present relative to cadaveric rat heart. Its acellular nature was confirmed by loss of > 96% DNA (p = 0.001) compared to normal rat heart. In vitro infusion of aqueous dye or Mercox resin suggested a complete arterial tree, with structural preservation of vascular conduits. In vivo perfusability of the ECM was demonstrated by heterotopic transplantation with anticoagulation (n=4) into RNU rats for 7 days. Recellularization of the vascular tree was attempted by In Vitro Langendorff perfusion of 2 x 107 rat aortal endothelial cells (ECs) followed by a 7d incubation with escalating pulsatile flow in a 3D bioreactor. CellTracker Green assessed EC viability and permitted visualization of engrafted cells by fluorescent microscopy. Vessels of different diameters contained “patches” of confluent endothelium with complete circumferential lining of many of the matrix conduits. ECs lining both chamber walls and trabeculae were also observed. Nuclear staining showed 537.8 +/− 67.6 ECs / mm2 on endocardial surfaces, as well as 311.7 +/− 61.8 ECs / mm2 in vessels. To enhance the delivery of cells into the ventricular walls, a microcanulization of the brachiocephalic artery with sustained aortal perfusion was undertaken. This technique diverted more cells to the vasculature and more broadly distributed the cells in each area resulting in a lower cell density; 199.8 +/− 25.0 ECs / mm2 in vessels vs 125.8 +/− 43.4 ECs / mm2 on endocardial surfaces. In conclusion, these data suggest that by using detergent prepared acellular ECM of a whole organ, generation of a complete endothelial lining of vascular structures may be possible.