Roles for GP IIb/IIIa and αvβ3 integrins in MDA-MB-231 cell invasion and shear flow-induced cancer cell mechanotransduction

Adhesion of cancer cell to endothelial cells and the subsequent trans-endothelial migration are key steps in hematogenous metastasis. However, the molecular mechanisms of cancer cell/endothelial cell interaction under hemodynamic shear flow and how shear flow-induced cancer cell mechanotransduction are yet to be fully defined. In this study, we identified that the integrins of both platelet glycoprotein IIb/IIIa (GP and alpha v beta 3 were crucial for hematogenous metastasis of human breast carcinoma MDA-MB-231cells. The cell migration and invasion were studied by using Millicell cell culture insert system. The numbers of invaded MDA-MB-231 cells significantly increased by thrombin-activated platelets and reduced by eptifibatide, a platelet inhibitor. Meanwhile, RGDWE peptides, a specific inhibitor of alpha v beta 3 integrin, also inhibited MDA-MB-231 cell invasion. We further used a parallel-plate flow chamber to investigate MDA-MB-231 cell adhesion under flow conditions. Alike in static condition, the adhesion capability of MDA-MB-231 cells to endothelial monolayer was also significantly affected by GP Ilb/Illa and alpha v beta 3 integrins. The expression of matrix metalloproteinase-2 (MMP-2), MMP-9 and alpha v beta 3 integrin in MDA-MB-231 cells were up-regulated after low shear stress exposure (1.84 dynes/cm(2), 2 h). Moreover, we also demonstrated that low shear stress induced a sustained activation of p85 (a regulatory subunit of PI3K) and Akt. Pre-treating MDA-MB-231 cells with the specific PI3K inhibitor of LY294002 abolished the shear stress induced-Akt activation, and the expression of MMP-2, MMP-9, vascular endothelial growth factor (VEGF) and ,v133 integrin were also down-regulated. Immunofluorescence assay showed that low shear stress also induced alpha v beta 3 integrin clustering and nuclear factor-kappa B (NF-kappa B) activation. Interestingly, shear stress-induced activation of Akt and NF-kappa B was attenuated by LM609, a specific antibody of alpha v beta 3 integrin. It suggests that alpha v beta 3 integrin might be as a mechanosensor to trigger both PI3K/Akt and NF-kappa beta signaling pathways. Taken together, these results establish that GP lIb/Illa and alpha v beta 3 integrins are essential mediators, and provide insight into how shear stress-induced alpha v beta 3 integrin activation and the downstream pathways for contribution to MDA-MB-231 cell adhesion, migration and invasion. (C) 2013 Elsevier Ireland Ltd. All rights reserved.