Davies, Peter F
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Publication Role of lateral cell–cell border location and extracellular/transmembrane domains in PECAM/CD31 mechanosensation(2004-08-06) Kaufman, David A.; Albelda, Steven M.; Sun, Jing; Davies, Peter F.Phosphorylation of tyrosine residues on platelet–endothelial cell adhesion molecule-1 (PECAM-1), followed by signal trans- 13 duction events, has been described in endothelial cells following exposure to hyperosmotic and fluid shear stress. However, it is 14 unclear whether PECAM-1 functions as a primary mechanosensor in this process. Utilizing a PECAM-1–null EC-like cell line, we 15 examined the importance of cellular localization and the extracellular and transmembrane domains in PECAM-1 phosphorylation 16 responses to mechanical stress. Tyrosine phosphorylation of PECAM-1 was stimulated in response to mechanical stress in null cells 17 transfected either with full length PECAM-1 or with PECAM-1 mutants that do not localize to the lateral cell–cell adhesion site and 18 that do not support homophilic binding between PECAM-1 molecules. Furthermore, null cells transfected with a construct that 19 contains the intact cytoplasmic domain of PECAM-1 fused to the extracellular and transmembrane domains of the interleukin-2 20 receptor also underwent mechanical stress-induced PECAM-1 tyrosine phosphorylation. These findings suggest that mechano- 21 sensitive PECAM-1 may lie downstream of a primary mechanosensor that activates a tyrosine kinase.Publication Spatiotemporal Analysis of Flow-Induced Intermediate Filament Displacement in Living Endothelial Cells(2001-01-01) Helmke, Brian P.; Thakker, David B.; Goldman, Robert D.; Davies, Peter F.The distribution of hemodynamic shear stress throughout the arterial tree is transduced by the endothelium into local cellular responses that regulate vasoactivity, vessel wall remodeling, and atherogenesis. Although the exact mechanisms of mechanotransduction remain unknown, the endothelial cytoskeleton has been implicated in transmitting extracellular force to cytoplasmic sites of signal generation via connections to the lumenal, intercellular, and basal surfaces. Direct observation of intermediate filament (IF) displacement in cells expressing green fluorescent protein-vimentin has suggested that cytoskeletal mechanics are rapidly altered by the onset of fluid shear stress. Here, restored images from time-lapse optical sectioning fluorescence microscopy were analyzed as a four-dimensional intensity distribution function that represented IF positions. A displacement index, related to the product moment correlation coefficient as a function of time and subcellular spatial location, demonstrated patterns of IF displacement within endothelial cells in a confluent monolayer. Flow onset induced a significant increase in IF displacement above the nucleus compared with that measured near the coverslip surface, and displacement downstream from the nucleus was larger than in upstream areas. Furthermore, coordinated displacement of IF near the edges of adjacent cells suggested the existence of mechanical continuity between cells. Thus, quantitative analysis of the spatiotemporal patterns of flow-induced IF displacement suggests redistribution of intracellular force in response to alterations in hemodynamic shear stress acting at the lumenal surface.