Abstract 1039: Endothelial Deletion Of Nck1 Reduces Atherosclerosis In Apoe-/- Mice Through The Nck1-atf3 Axis

Abstract

The Nck family of signaling adaptors, including both Nck1 and Nck2, classically regulate cell motility during vascular development and postnatal angiogenesis. While Nck1 and Nck2 play redundant roles in this regard with deletion of both isoforms required to affect tissue remodeling, we demonstrated that global Nck1 deletion was sufficient to reduce atherosclerotic plaque formation and selective Nck1 inhibition reduces proinflammatory endothelial activation in response to oscillatory shear stress. Therefore, we sought to determine the role of endothelial Nck1 in atherosclerosis and ischemic angiogenesis using novel endothelial-specific Nck1 knockout mice. Endothelial-specific Nck1 deletion significantly reduced Western diet-induced atherosclerotic plaque formation at multiple sites compared to mice expressing endothelial-specific Cre alone or to endothelial-specific Nck2 knockout mice. This reduced plaque formation was characterized by diminished macrophage and smooth muscle incorporation despite enhanced weight gain in endothelial Nck1 knockout mice. RNA Sequencing analysis of endothelial cells in culture showed that Nck1 depletion reduces the expression of a variety of proinflammatory pathways but enhances the expression of genes involved in central carbon metabolism and proliferation. While Nck1 deletion is sufficient to blunt atherogenic inflammation, Nck1 inhibition did not affect angiogenesis or limb perfusion in the femoral artery ligation model of hind limb ischemia. To further characterized Nck1’s proinflammatory role, we utilized affinity precipitation-mass spectrometry and identified Activating Transcription Factor 3 ATF3 as a Nck1-selective binding partner. While no studies to date have examined endothelial ATF3 signaling in atherosclerosis, our preliminary data show that Nck1 is required for ATF3 expression in response to disturbed flow in vitro and in vivo , but the mechanism by which disturbed flow stimulates ATF3 expression remains unknown. ATF3 knockdown reduces disturbed flow-induced proinflammatory gene expression, suggesting that the Nck1-ATF3 signaling axis is a novel regulator of flow-induced inflammation.