Transcriptional and functional modulation of the endothelial cell inflammatory response by a biomechanical stimulus

The non-random distribution of the early atherosclerotic lesions suggests an important role for hemodynamic forces in atherogenesis. The putative protective effects of certain hemodynamic forces in lesion-resistant areas can be modeled in vitro by the application of laminar shear stress (LSS) to cultured endothelial monolayers. This thesis tested the hypothesis that preconditioning with a physiological level of steady LSS can act to modulate the pattern of gene expression and functional responsiveness of endothelial cells to an inflammatory stimulus, interleukin-1beta (IL-1). Various patterns of modulation attributed to the inflammatory stimulus, the biomechanical stimulus, or their combination were revealed on a genomic scale using transcriptional profiling. Development of an improved statistical test for detecting reproducibly regulated genes in replicated microarray data (Zpool), along with software to automate data processing and provide interactive analysis of results ( Argus), facilitated the selection of genes for further study. Preconditioning with LSS appeared to have a selective rather than global effect, resulting in the reproducible modulation of a subset of IL-1 induced genes. Validation experiments identified TNF-related apoptosis-inducing ligand (TRAIL) and plasminogen activator inhibitor-2 (PAI-2) as genes whose IL-1 induction was most strongly and reproducibly modulated by LSS preconditioning. IL-1 stimulation upregulated TRAIL mRNA and protein levels, and LSS preconditioning completely blocked this upregulation. Sensitivity to apoptosis and activation from exogenous TRAIL was dramatically inhibited by preconditioning with LSS or with a shear stress waveform reproduced from an "athero-protected" region of the human carotid artery. Modulation of the TRAIL pathway by biomechanical stimuli in vitro suggests that TRAIL may play a role in various flow-dependent apoptotic processes in the vasculature during atherogenesis, angiogenesis, or vascular remodeling. In summary, the demonstration that endothelial gene expression is modulated by preconditioning with LSS suggests that the endothelial cell can integrate multiple humoral and biomechanical stimuli in manner that may be relevant to endothelial function in health and disease.