Disturbed flow promotes arterial stiffening through thrombospondin-1 (TSP-1) signaling pathways

Arterial stiffness is a prevalent, independent cardiovascular risk factor, but the underlying mechanisms are not well understood. Wall shear stress and shear-sensitive genes may promote arterial stiffening through clinically important signaling pathways. Our goal was to identify how disturbed blood flow leads to arterial stiffness using the mouse partial carotid ligation model. Here we used our in vivo partial carotid ligation model to induce d-flow in the LCA while the contralateral RCA continues to experience stable laminar flow using the C57BL/6x129SvEv mice, TSP-1 knockout (KO), and C57Bl/6J mice. We compared these to aged (80 week) mice which had increased arterial stiffness due to aging. Changes in gene expression were identified using microarrays that were performed on the endothelial-enriched RNA isolated from the carotids exposed to stable flow (RCA) and compared to disturbed flow (LCA). Arterial stiffness was determined ex vivo by biaxial mechanical testing and in vivo by ultrasound techniques. Myointimal hyperplasia and immunohistochemistry were performed in sectioned carotid arteries. In vitro testing of signaling pathways utilized oscillatory and laminar wall shear stress. Human arteries were tested ex vivo to validate critical results found in the animal model. Endothelial-enriched RNA was isolated from WT carotids exposed to stable flow or disturbed flow. Three replicates each.