A pro-inflammatory lipid mediator, lysophosphatidic acid (LPA), is a potent activator of the RhoA/Rho kinase signaling pathway and has been shown to induce the expression of cell adhesion molecules (CAMs). LPA levels are elevated in the circulation of patients with acute coronary syndrome and has also been described inside human atherosclerotic specimens. However, the involvement of Rho-kinase isoforms (i.e., ROCK1, ROCK2) in LPA-mediated expression of CAMs in endothelial cells is not fully understood. In this study, we investigated isoform-specific roles of Rho-kinase in LPA-mediated E-selectin expression using human aortic endothelial cells (HAEC). Exposure of HAEC to LPA resulted in an increase of E-selectin expression at both mRNA (real-time PCR) and protein (Western blot) levels. We confirmed increased Rho-kinase activity in the LPA-stimulated cells. Y-27632, a specific Rho-kinase inhibitor, completely abolished E-selectin protein expression, indicating that LPA induces E-selectin expression via Rho-kinase activation. LPA-induced E-selectin gene expression was strongly suppressed by a chemical inhibitor of NF-κB, indicating the contribution of NF-κB transactivation in this pathway. Intriguingly, phosphorylation of NF-κB RelA/p65 was decreased by Rho-kinase inhibition in the LPA-stimulated cells. Furthermore, using isoform specific siRNAs, we found that Rho-kinase isoform ROCK2 is the key mediator of LPA-induced E-selectin expression and cell adhesion between HAECs and human monocytic cells (THP-1). In the present study, we demonstrate that ROCK2 mediates LPA-induced expression of E-selectin via NF-κB signaling in endothelial cells. ROCK2 could be an important therapeutic target against atherosclerosis.
Y. Takeda: None. K. Matoba: None. D. Kawanami: None. Y. Nagai: None. T. Akamine: None. S. Ishizawa: None. Y. Kanazawa: None. T. Yokota: None. K. Utsunomiya: None.