Recellularization of Porcine Internal Thoracic Arteries as a Tissue Engineered Small-Diameter Vascular Graft Alternative

Small-diameter vascular grafts are the leading treatment for myocardial infarctions resulting from atherosclerosed coronary vasculature. A potential alternative to using patient-derived grafts is the use of tissue engineered vessels. The removal of native cells from xenogenic vessels allows for recellularization with human cells types; however, the decellularization process depends on the use of cytotoxic reagents that must be removed prior to recellularization. The recellularization process must produce a functional graft that contains the multiple cell types found within arteries. We hypothesized that porcine internal thoracic arteries decellularized using a combination of detergents could be processed to support human endothelial and smooth muscle cell growth. We also hypothesized that in order to optimize in vitro co-culture conditions during recellularization, a mix of media types would be necessary to simultaneously support endothelial and smooth muscle cell growth. A cytotoxicity assay was performed to assess the effects of residual detergents on endothelial cells seeded onto the scaffolds. A relationship between the degree of detergent rinsing and cellular viability was identified via a resazurin reduction assay with more extensive rinsing significantly enhancing cell viability. This same cell viability assay was used to identify media combinations that supported growth of both endothelial and smooth muscle cells. Both cell types were able to grow in a 50:50 mixture of their media types without any loss of viability or effect on morphology. Furthermore, endothelial cells grown in the mixed medium maintained their characteristic CD31 expression. Taken together, these results show that human cells native to arteries can remain viable within the extracellular matrix of porcine internal thoracic artery scaffolds after thorough scaffold detoxification. Additionally, the co-culture conditions established can support the growth of both endothelial and smooth muscle cell types found within arteries. Future work will focus on simultaneously culturing the cells within scaffolds to build toward the goal of tissue engineering small-diameter vascular graft alternatives.https://digitalcommons.winthrop.edu/sureposters/1005/thumbnail.jp

Optimization of Culture Conditions for the Simultaneous Recellularization of Porcine Internal Thoracic Artery Scaffolds with Multiple Cell Types

The increasing prevalence of ischemic diseases has generated a growing need for heart bypass surgeries. The goal of our research is to tissue engineer vascular grafts from scaffolds derived from decellularized porcine internal thoracic arteries that can be recellularized with patient-specific cells and restore function more effectively than current methods. The ability to recellularize scaffolds with multiple cell types, including endothelial cells (ECs), smooth muscle cells (SMCs), and adipose-derived mesenchymal stem cells (ADSCs) is important for imparting function to an engineered vessel. However, an important question that arises is what type of cell culture media should be used to allow the different cell types to grow together. We hypothesized that mixtures of two media types that support growth of two of the cell types of interest could be identified. To test our hypothesis, we grew ECs in different combinations of 1) EC and SMC medium and 2) EC and ADSC medium. We also grew ADSCs in different combinations of EC and ADSC medium. We monitored the viability of the cells and assessed the expression of CD31 by ECs cultured in different media combinations. In all cases, the growth of the cell type of interest was no different in a 50:50 combination of its medium and the other medium than growth in 100% of its own medium. In addition, CD31 expression was maintained by ECs under all experimental conditions. These results suggest that 50:50 mixtures of culture medium will support the growth of two cell types within our vascular scaffolds