Upregulation of chemokine receptor CXCR4 and caspase-dependent apoptosis genes can synergistically favour activation of endothelial cells exposed to low shear stress

Purpose: In the pathogenesis of atherosclerosis, a central role is represented by the development of inflammation with influx of chemokine-mediated leukocytes in the vascular wall. Macrophage migration inhibitory factor (MIF) promotes the atherogenic and inflammatory recruitment of T cells through the chemokine receptor CXCR4. Whether local shear stresses generated by blood flow may modulate activation of CXCR4/MIF axis is yet unknown. Aims of this study were to evaluate the effect of different shear stresses on gene expression of molecules involved in homeostatic and inflammatory cell migration processes, and to assess potential synergy with up-regulation of apoptotic processes. Methods: Human Umbilical Vein Endothelial cells were submitted to 0, 1 5 and 10 dyne/cm2 in a Laminar Flow Bioreactor for 24 hours in order to compare gene expression modulation at normal or reduced shear stress by microarray techniques. Total RNA, extracted from each experimental set, was analyzed by Affimetrix technology. Genes were sorted for differential expression based on oneway ANOVA and different expressed genes were identified as those genes having adjusted p values of < 0.05 with fold changed of at least 2 fold in modulus. Results: The microarray analysis highlighted a strong up-regulation of the chemokine receptor CXCR4 at low (1 dyne/cm2) compared to intermediate and high shear stresses (5 and 10 dyne/cm2); also Caspase-8 apoptotic gene was markedly upregulated at low shear stress. Conversely, a marked downregulation of TNFAIP3 gene was observed. TNFAIP3 is able to prevent TNF-induced cytotoxicity and reduce monocyte-cell interaction, its downregulation may favour monocyte adhesion to vessel wall. Conclusions: Endothelial chemokine receptor CXCR4 activation at low shear stress can play a role in the development of atherosclerosis. This receptor acts as an important mediator of the endothelial response to damage, being able to interact with MIF, potent mediator of T cell recruitment. The concomitant overexpression of Caspase-8, involved in early phase of apoptotic process, and the downregulation of TNFAIP3 suggest that all these gene modulations, observed at low shear stress, can synergistically promote an initial inflamed cell phenotype with a cascade of events that precede the endothelial activation through adhesion molecules expression.