Vara, D.; (2010) Development of a coronary artery bypass graft with the aid of tissue engineering: investigation of gene expression on seeded compliant nanocomposite conduits. Doctoral thesis , UCL (University College London).

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Abstract

BACKGROUND. Coronary artery bypass graft surgery is a commonly performed procedure. The internal thoracic or mammary artery is gaining widespread preference as the bypassing conduit. Synthetic grafts used for large diameter substitutes are successful but have dismal patency as small diameter (< 6mm) grafts due to compliance mismatch and thrombogeniecity. To overcome this, cell adhesion to synthetic scaffolds is used to construct tissue engineered grafts. For this to be successful a precise understanding of the behaviour of cells at the synthetic graft surface is required under static and haemodynamic conditions. The aim of this research was to investigate gene expression of seeded human umbilical vein endothelial cells (HUVEC) on the novel compliance conduit under physiological flow condition; furthermore various physiological shear stress preconditioning was used to investigate adhesion of HUVEC on the conduit. METHODS. HUVEC seeding of a novel polymer nanocomposite was undertaken. An optimal method for extracting mRNA from HUVEC seeded onto conduits was then validated. The optimal seeding conditions for the conduits were delineated. Haemodynamic conditions were applied to the seeded conduits and gene expression was investigated using polymerase chain reaction (PCR). Shear stress was used to assess the ideal preconditioning environment. RESULTS. Studies of nanocomposite graft material and cultured HUVEC proved that the novel nanocomposite polymer was non-toxic to cells and supported good rates of growth. To provide useful flow studies an extrusion-phase inversion method was used to reproducibly fabricate conduits of this nanocomposite with compliance similar to the native artery. The optimal seeding density of the conduits was found to be 1.2 x 104 cells/cm2. It was demonstrated that RNA can be extracted from seeded conduits and I succeeded in showing the optimal technique. This study culminates in the combination of all these techniques when the gene expression of HUVEC under flow was studied after physiological shear stress was applied on the conduits. Genes significantly upregulated included TGF-β1, COL-1 and PECAM-1. Low shear stress demonstrated the optimal preconditioning environment with increasing expression of VEGFR-1 and VEGFR-2 genes. CONCLUSION. This thesis demonstrated that novel nanocomposite small diameter bypass graft can be seeded with human endothelial cells.

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