Atherosclerosis is an inflammatory disease of the vasculature that can be regulated by regional differences in arterial hemodynamics. Specific blood flow shear stress waveforms have been characterized as either atheroprotective or atheroprone, where an atheroprone region is more susceptible to the development of the disease. Smooth muscle cells (SMCs) and endothelial cells (ECs) play equally important roles in maintaining homeostasis of the vasculature and both cell types are intimately involved in the progression of atherosclerosis. SMCs, in particular, are known to switch from a contractile phenotype to a proliferative/remodeling phenotype during atherogenesis leading to neointima invasion, formation of the fibrous cap and eventually vessel luminal narrowing. A hallmark event of SMC phenotypic switching is suppression of contractile proteins that define the differentiated SMC, including smooth muscle myosin heavy chain (SMMHC), smooth muscle alpha actin (SMaA), and myocardin. Atheroprone hemodynamic shear stresses on the endothelium in vivo and in vitro are known to promote a pro-inflammatory phenotype;​ however, regulation of the SMC phenotype due to hemodynamic forces exerted on ECs is unknown. Thus, the hypothesis that ECs play a key role in regulating classic SMC phenotypic modulation as a function of the local hemodynamic environment was tested.

A novel in vitro hemodynamic EC/SMC co-culture model was developed to determine the role of hemodynamic regulation of EC and SMC phenotypes. Human ECs and SMCs were plated on a synthetic elastic lamina and human-derived atheroprone and atheroprotective shear stresses were imposed on ECs. Initial characterization of the model indicated that atheroprotective, but not atheroprone, flow caused EC alignment with the direction of flow and SMC alignment more perpendicular to flow, similar to in vivo vessel organization. Furthermore, phenotypic modulation of each cell type was observed as a result of differential flow environments. Atheroprone flow decreased genes associated with differentiated ECs (endothelial nitric oxide synthase, Tie2, and Kruppel-like factor 2) and SMCs (SMaA and myocardin) and induced a proinflammatory phenotype in ECs and SMCs (VCAM-1, IL-8, and monocyte chemoattractant protein-1). Atheroprone flow-induced changes in SMC differentiation markers were regulated at the chromatin level, as indicated by decreased serum response factor (SRF) binding to the smooth muscle a-actin-CC(A/T)₆GG (CArG) promoter region and decreased histone H4 acetylation.

EC/SMC cross talk via secreted factors was additionally examined within the system, where the mechanism by which EC-derived IL-8 influences SMC expression of VCAM-1 was studied. Interestingly, despite the known IL-8 inflammatory function, blocking its activity in this model resulted in a further increase in VCAM-1 by 3.2-fold. Cell culture studies demonstrated that IL-8 can serve to limit or resolve VCAM-1 expression in SMCs. We have thus identified a novel role for IL-8 to act in a negative feedback loop to prevent further SMC inflammation during early atherogenesis.

Together, these results provide a novel mechanism whereby modulation of the EC phenotype by hemodynamic shear stresses, atheroprone or atheroprotective, play a critical role in mechanical-transcriptional coupling and regulation of the SMC and EC phenotype.