Mechanisms of induction of endothelial cell E-selectin expression by smooth muscle cells and its inhibition by shear stress

Abstract

E-selectin is a major adhesion molecule expressed by endothelial cells (ECs), which are exposed to shear stress and neighboring smooth muscle cells (SMCs). We investigated the mechanisms underlying the modulation of EC E-selectin expression by SMCs and shear stress. SMC coculture induced rapid and sustained increases in expression of E-selectin and phosphorylation of interleukin-1 (IL-1) receptor-associated kinase and glycopro-tein-130, as well as the downstream mitogen-activated protein kinases (MAPKs) and Akt. By using specific in-hibitors, dominant-negative mutants, and small interfering RNA, we demonstrated that activations of c-Jun-NH 2-terminal kinase (JNK) and p38 of the MAPK pathways are critical for the cocul-ture-induced E-selectin expression. Gel shifting and chromatin immunoprecipita-tion assays showed that SMC coculture increased the nuclear factor-B (NF-B)-promoter binding activity in ECs; inhibition of NF-B activation by p65-anti-sense, lactacystin, and N-acetyl-cysteine blocked the coculture-induced E-selectin promoter activity. Protein arrays and blocking assays using neutralizing anti-bodies demonstrated that IL-1 and IL-6 produced by EC/SMC cocultures are major contributors to the coculture induction of EC signaling and E-selectin expression. Preshearing of ECs at 12 dynes/cm 2 inhibited the coculture-induced EC signaling and E-selectin expression. Our findings have elucidated the molecular mechanisms underlying the SMC induction of EC E-selectin expression and the shear stress protection against this SMC induction. (Blood. 2007; Introduction During the development of atherosclerotic lesions, vascular smooth muscle cells (SMCs) change from their physiologic contractile phenotype to the pathophysiologic synthetic phenotype and migrate into the intima, where they release proinflam-matory cytokines and interact with vascular endothelial cells (ECs) to regulate their gene expression and function, including the modulation of leukocyte recruitment. 1-3 ECs are constantly subjected to blood flow-induced shear stress, which can modulate leukocyte-EC interaction and the subsequent leukocyte extravasation into inflamed tissue, mainly by modulating EC surface expression of adhesion molecules. 3,4 E-selectin is a major EC adhesion molecule that regulates binding and extravasation of leukocytes from the bloodstream to sites of inflammation. The effects of cytokines and shear stress on EC E-selectin expression have been extensively studied. The E-selectin gene is rapidly expressed by ECs in response to proinflammatory cytokines, and it is more responsive to disturbed and oscillatory flows 5,6 than to laminar shear stress. 7 Most studies on effects of shear stress on EC gene expression have been performed on EC monolayers, which may not reflect the in vivo environment of ECs, which exist in close proximity to SMCs. By using our newly developed EC/SMC coculture flow system 8 in which ECs and SMCs are grown on opposite sides of a porous membrane, we demonstrated that coculture with SMCs induced E-selectin expr...

PDGF-BB and TGF-β1 on cross-talk between endothelial and smooth muscle cells in vascular remodeling induced by low shear stress

“…Shear stress acting on ECs changes the phenotype of the cocultured VSMCs, with the subsequent alterations in the EC release of proinflammatory cytokines, as well as proliferation, apoptosis, and gene expression that induce atherogenesis (22,23). These studies revealed that the interactions between ECs and VSMCs play an important role in vascular remodeling induced by LowSS.…”

“…After 8 h for attachment, the membrane was placed with the EC side down in the six-well plate, and VSMCs were seeded on the inside of the membrane at a density of 2 × 10 5 cells per cup. NSS and LowSS were subjected to ECs in the flow chamber system (20)(21)(22).…”

“…There are 1.6 million pores/cm 2 on the membrane, and each pore is 0.4 μm in diameter. The porous structure permitted the interaction between ECs and VSMCs and allowed the respective analysis on these two kinds of cells (20)(21)(22).…”

Oscillatory Shear Stress Induces Mitochondrial Superoxide Production: Implication of NADPH Oxidase and c-Jun NH2-Terminal Kinase Signaling

“…8c). Unlike our in vitro cultured BAEC model, human coronary arteries were constantly exposed to the circulating cytokines and growth factors as well as the paracrine effects of SMCs, all of which may contribute to JNK activation (5,8).…”

IL-8 reduces VCAM-1 secretion of smooth muscle cells by increasing p-ERK expression when 3-D co-cultured with vascular endothelial cells

“…Using this model, co-cultured HUVECs were shown to secrete IL-8, which prevented HUASMCs from expressing VCAM-1. Suppression of VCAM-1 expression in HUASMC cells is mediated by the p-ERK signaling pathway [21,22].…”