Davies, Peter F

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Now showing 1 - 6 of 6
  • 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.
  • Publication
    Imaging Live Cells Under Mechanical Stress
    (2003-03-01) Helmke, Brian P; Davies, Peter F
    Cellular responses to mechanical stimuli are implicated in the structural and functional adaptation of many tissues. For example, cellular mechanisms mediate bone and skeletal muscle remodeling during mechanical loading, lung function during ventilator-induced injury, hearing loss in the inner ear, and blood flow-mediated cardiovascular pathophysiology. Since much of our own work investigates vascular biomechanics, we will focus in this chapter on the techniques used to study vascular endothelial cells in vitro; however, similar techniques can be used to study other cell types.
  • Publication
    The convergence of haemodynamics, genomics, and endothelial structure in studies of the focal origin of atherosclerosis
    (2002-04-01) Davies, Peter F; Shi, Congzhu; Polacek, Denise C; Helmke, Brian P
    The completion of the Human Genome Project and ongoing sequencing of mouse, rat and other genomes has led to an explosion of genetics-related technologies that are finding their way into all areas of biological research; the field of biorheology is no exception. Here we outline how two disparate modern molecular techniques, microarray analyses of gene expression and real-time spatial imaging of living cell structures, are being utilized in studies of endothelial mechanotransduction associated with controlled shear stress in vitro and haemodynamics in vivo. We emphasize the value of such techniques as components of an integrated understanding of vascular rheology. In mechanotransduction, a systems approach is recommended that encompasses fluid dynamics, cell biomechanics, live cell imaging, and the biochemical, cell biology and molecular biology methods that now encompass genomics. Microarrays are a useful and powerful tool for such integration by identifying simultaneous changes in the expression of many genes associated with interconnecting mechanoresponsive cellular pathways.
  • Publication
    The Cytoskeleton Under External Fluid Mechanical Forces: Hemodynamic Forces Acting on the Endothelium
    (2002-03-01) Helmke, Brian P; Davies, Peter F
    The endothelium, a single layer of cells that lines all blood vessels, is the focus of intense interest in biomechanics because it is the principal recipient of hemodynamic shear stress. In arteries, shear stress has been demonstrated to regulate both acute vasoregulation and chronic adaptive vessel remodeling and is strongly implicated in the localization of atherosclerotic lesions. Thus, endothelial biomechanics and the associated mechanotransduction of shear stress are of great importance in vascular physiology and pathology. Here we discuss the important role of the cytoskeleton in a decentralization model of endothelial mechanotransduction. In particular, recent studies of four-dimensional cytoskeletal motion in living cells under external fluid mechanical forces are summarized together with new data on the spatial distribution of cytoskeletal strain. These quantitative studies strongly support the decentralized distribution of luminally imposed forces throughout the endothelial cell.
  • Publication
    Mapping Mechanical Strain of an Endogenous Cytoskeletal Network in Living Endothelial Cells
    (2003-04-01) Helmke, Brian P.; Rosen, Amy B.; Davies, Peter F
    A central aspect of cellular mechanochemical signaling is a change of cytoskeletal tension upon the imposition of exogenous forces. Here we report measurements of the spatiotemporal distribution of mechanical strain in the intermediate filament cytoskeleton of endothelial cells computed from the relative displacement of endogenous green fluorescent protein (GFP)-vimentin before and after onset of shear stress. Quantitative image analysis permitted computation of the principal values and orientations of Lagrangian strain from 3-D high-resolution fluorescence intensity distributions that described intermediate filament positions. Spatially localized peaks in intermediate filament strain were repositioned after onset of shear stress. The orientation of principal strain indicated that mechanical stretching was induced across cell boundaries. This novel approach for intracellular strain mapping using an endogenous reporter demonstrates force transfer from the lumenal surface throughout the cell.