Abstract 266: Disturbed Flow Promotes Endothelial Cell Injury via the Induction of Developmental Genes

Abstract

Introduction: Atherosclerosis is influenced by blood flow patterns which exert wall shear stress (WSS) on endothelial cells (EC). Low oscillatory WSS promotes atherosclerosis by inducing EC apoptosis and activation, whereas high unidirectional WSS is protective. We used microarray technology coupled to computational fluid dynamics to study the transcriptome of EC at regions of the porcine aorta exposed to low, oscillatory or high, unidirectional WSS. The study revealed differential expression of multiple genes that co-ordinate embryonic development. We hypothesised that this gene set includes WSS-sensitive regulators of EC survival. Methods and Results: EC were isolated from distinct anatomical regions of the porcine aorta using collagenase (n=6), and quantitative RT-PCR revealed elevated expression of developmental genes (GATA4, HAND2, TWIST1, FZD5, BMP2, SLIT2, PDGFRA, and FBN2) at the inner (low WSS) compared to the outer curvature (high WSS) (all p<0.05). Similarly, en face staining demonstrated that expression of GATA4 protein in EC was higher at the inner compared to the outer curvature of the murine aortic arch (p<0.05). Studies using cultured porcine aortic EC (n=7) or human umbilical vein EC (n=7) revealed that GATA4, HAND2, TWIST1, FZD5, BMP2, SLIT2, PDGFRA, and FBN2 were induced in HUVEC or PAEC exposed to low, oscillatory WSS for 72h using either the orbital shaking (210 rpm; +/-WSS 4 dyn/cm2) or IBIDI™ (+/-5 dyn/cm2 at 2Hz) systems (all p<0.05). The rate of caspase-3 activation was significantly higher in EC exposed to low, oscillatory WSS compared to high, unidirectional WSS (3.1% vs 0.7%; p<0.0001; n=10). Silencing of GATA4, FZD5 and BMP2 significantly reduced apoptosis in EC exposed to low, oscillatory WSS (1.2%, 0.6% and 1.6%; p<0.05). Conclusions: We conclude that WSS influences EC viability through the induction of GATA4, FZD5 and BMP2; molecules that have a well-defined role in embryonic development. Further work is required to define the molecular mechanisms that underlie the induction of apoptosis by these molecules. Our observations illuminate the molecular mechanisms that regulate the focal nature of vascular injury and atherosclerosis.