Abstract 2084: Endothelial Cells With Silenced Flrt2 Expression Exhibit Enhanced Retention On Vascular Graft Materials Under Fluid Shear Stress In Vitro

Abstract

Myocardial infraction is a leading cause of death, and surgical intervention is often required to treat severe coronary artery disease. Autologous vessel grafts are the only option for bypass graft procedures. Synthetic vascular grafts less than 6 mm diameter fail due to unacceptable patency rates (~60% at 1 year). The loss of patency is accredited to platelet activation, thrombosis, and neointimal hyperplasia. A promising strategy to improve patency is to endothelialize synthetic vascular grafts ex-vivo, but previous attempts have met with modest success due to detachment of cells upon exposure to fluid shear stress. We previously used RNA-seq to characterized gene expression of endothelial cells (ECs) which remained adherent to synthetic grafts post shear stress. We found that fibronectin leucine-rich transmembrane protein 2 (FLRT2) was significantly downregulated in adherent subpopulations. We therefore hypothesize that suppressing FLRT2 expression in ECs will result in better retention on synthetic vascular grafts upon exposure to shear stress. We used silencing RNA approach using lipofectamine and confirmed successful knockdown of FLRT2 expression (Si-FLRT2). We then seeded Si-FLRT2 cells onto a commercially available Goretex graft material and subjected them to fluid shear stress of for 20 min. We then stained the samples with DAPI. We observed increased retention of Si-FLRT2 ECs on Goretex graft material in comparison to non-specifically targeted scramble siRNA containing ECs. In addition, we compared both vascular cell adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1) levels to assess endothelial cell activation and found that silencing of FLRT2 lead to decreased levels of both VCAM-1 (by 49.2%±3.5%) and ICAM-1 (by 57.1%±3.4%) suggestive of decreased endothelial cell activation. In conclusion, we show that FLRT2 plays an important role in adhesion of ECs onto a commercially available synthetic graft material and silencing its expression ex-vivo may have beneficial clinical outcomes. In addition, we found that FLRT2 silencing may lead to decreased endothelial cell activation and we would like to further investigate the molecular mechanisms underlying the role of FLRT2 in both EC adhesion and activation.