Endothelial actin cytoskeleton remodeling during mechanostimulation with fluid shear stress

Fluid shear stress stimulation induces endothelial cells to elongate and align in the direction of applied flow. Using the complementary techniques of photoactivation of fluorescence and fluorescence recovery after photobleaching, we have characterized endothelial actin cytoskeleton dynamics during the alignment process in response to steady laminar fluid flow and have correlated these results to motility. Alignment requires 24 h of exposure to fluid flow, but the cells respond within minutes to flow and diminish their movement by 50%. Although movement slows, the actin filament turnover rate increases threefold and the percentage of total actin in the polymerized state decreases by 34%, accelerating actin filament remodeling in individual cells within a confluent endothelial monolayer subjected to flow to levels used by dispersed nonconfluent cells under static conditions for rapid movement. Temporally, the rapid decrease in filamentous actin shortly after flow stimulation is preceded by an increase in actin filament turnover, revealing that the earliest phase of the actin cytoskeletal response to shear stress is net cytoskeletal depolymerization. However, unlike static cells, in which cell motility correlates positively with the rate of filament turnover and negatively with the amount polymerized actin, the decoupling of enhanced motility from enhanced actin dynamics after shear stress stimulation supports the notion that actin remodeling under these conditions favors cytoskeletal remodeling for shape change over locomotion. Hours later, motility returned to pre-shear stress levels but actin remodeling remained highly dynamic in many cells after alignment, suggesting continual cell shape optimization. We conclude that shear stress initiates a cytoplasmic actin-remodeling response that is used for endothelial cell shape change instead of bulk cell translocation.

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Faculty Opinions recommendation of Endothelial actin cytoskeleton remodeling during mechanostimulation with fluid shear stress.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.13409065.14780078 ◽

2011 ◽

Author(s):

Martin A Schwartz

Keyword(s):

Shear Stress ◽

Actin Cytoskeleton ◽

Fluid Shear Stress ◽

Fluid Shear ◽

Cytoskeleton Remodeling

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Binding of ADAMTS13 to Von Willebrand Factor Under Static Conditions and Under Fluid Shear Stress

Blood ◽

10.1182/blood.v112.11.258.258 ◽

2008 ◽

Vol 112 (11) ◽

pp. 258-258

Author(s):

Hendrik B Feys ◽

Patricia J Anderson ◽

J. Evan Sadler

Keyword(s):

Shear Stress ◽

Von Willebrand Factor ◽

Time Course ◽

Fluid Shear Stress ◽

Proteolytic Processing ◽

Full Length ◽

Fluid Shear ◽

Static Conditions ◽

Von Willebrand ◽

Willebrand Factor

Abstract ADAMTS13 is a plasma metalloprotease that is essential for the normal proteolytic processing of von Willebrand factor (VWF). Dysfunctional ADAMTS13 may lead to thrombotic thrombocytopenic purpura, as uncleaved and unusually large VWF multimers accumulate in the blood and cause intravascular platelet aggregation. Many studies indicate that proteolysis of multimeric VWF involves conformational changes in the VWF A2 domain that expose the Y1605-M1606 scissile bond and also allow substrate binding to multiple exosites on ADAMTS13. For example, VWF is resistant to proteolysis by ADAMTS13 unless the VWF is subjected to fluid shear stress, mild denaturation with guanidine or urea, or adsorption onto a surface. However, the functional interactions between shear stress, various ADAMTS13 binding sites and VWF cleavage are not understood. Therefore, we investigated the effect of fluid shear stress and ADAMTS13 structure on ADAMTS13-VWF binding and VWF cleavage. Upon mixing recombinant VWF (rVWF) and ADAMTS13 in a physiological buffer (50 mM HEPES, 5 mM CaCl2, 1 μM ZnCl2, 150 mM NaCl, pH 7.4), we found that immunoprecipitation with anti-VWF also pulled down substantial amounts of ADAMTS13. Although less striking, a similar result was obtained with purified plasma VWF. Therefore, ADAMTS13 can bind VWF without gaining access to the cleavage site in VWF domain A2. When fluid shear stress was applied for 2 min with a bench-top vortexer, ADAMTS13 binding increased 3-fold and VWF was also cleaved. Lowering the ionic strength markedly increased the rate of VWF cleavage but did not affect ADAMTS13 binding, which suggests that cleavage and binding depend on distinct VWF-ADAMTS13 interactions. Shear-induced binding was reversible slowly upon removal of unbound ADAMTS13 or rapidly by addition of SDS. ADAMTS13-VWF binding was stable for at least 24 h after cessation of shear stress, indicating that the structural change in VWF that promotes binding was not readily reversible. Using a catalytically inactive ADAMTS13 variant to simplify the analysis of binding assays, 30 nM ADAMTS13(E231Q) bound to 30 μg/ml rVWF (120 nM subunits) with a stoichiometry of 0.012 ± 0.004 under static conditions and 0.098 ± 0.023 after shearing (mean ± SD, n = 3, P = 0.019). With 120 nM ADAMTS13(E231Q) the stoichiometry increased to 0.086 ± 0.036 under static conditions and 0.469 ± 0.033 after shearing for 2 min. Recombinant ADAMTS13 truncated after TSP-1 repeat 8 (lacking the C-terminal CUB domains, delCUB), or truncated after the Spacer domain (consisting of domains MDTCS), did not bind rVWF under static conditions, implicating the CUB domains in binding to VWF. In contrast, full-length ADAMTS13, delCUB and MDTCS bound similarly to rVWF after shearing. In a previous study, delCUB and MDTCS did not cleave VWF subjected to fluid shear stress (Zhang et al, Blood2007; 110: 1887–1894). However, under the conditions employed in these experiments, MDTCS and delCUB displayed significant proteolytic activity, cleaving VWF at a rate comparable to that of full length ADAMTS13 when shear stress was applied over a time course of 0–160 sec. We conclude that ADAMTS13 CUB domains contribute to binding a few sites on multimeric VWF under static conditions, whereas ADAMTS13 MDTCS domains are sufficient to bind many sites in an altered conformation of VWF that is induced by fluid shear stress. Binding of ADAMTS13 to unsheared VWF multimers may facilitate the cleavage of VWF within a growing thrombus.

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Fluid Shear Stress Reduces Simvastatin Induced Adhesion Molecule Expression in Cytokine Activated Endothelial Cells

ASME 2009 Summer Bioengineering Conference, Parts A and B ◽

10.1115/sbc2009-206539 ◽

2009 ◽

Author(s):

Joanna Rossi ◽

Léonie Rouleau ◽

Jean-Claude Tardif ◽

Richard L. Leask

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Shear Stress ◽

Adhesion Molecule ◽

Fluid Shear Stress ◽

Lipid Lowering ◽

Adhesion Molecule Expression ◽

Fluid Shear ◽

Endothelial Cell Function ◽

Static Conditions

Although originally designed as inhibitors of cholesterol biosynthesis, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, or statins, are now known to also have non-lipid lowering benefits [1]. Statins have been reported to modulate gene expression in endothelial cells, however, the effect of statins on adhesion molecule expression is contradictory. Some studies report a decrease in adhesion molecule mRNA and/or protein after statin treatment [2], while others have shown that statins potentiate the effect of tumor necrosis factor alpha (TNFα) [3]. To the best of our knowledge, the effects of statins on gene expression in cultured endothelial cells has been done in static conditions only and no study has examined the effect of blood flow. This is particularly important since fluid shear stress is a strong regulator of endothelial cell function and phenotype [4]. The purpose of this study was to clarify the effects of statins on vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) expression in endothelial cells by evaluating their biological response under fluid flow.

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Podocytes are sensitive to fluid shear stress in vitro

AJP Renal Physiology ◽

10.1152/ajprenal.00196.2005 ◽

2006 ◽

Vol 291 (4) ◽

pp. F856-F865 ◽

Cited By ~ 76

Author(s):

Colin Friedrich ◽

Nicole Endlich ◽

Wilhelm Kriz ◽

Karlhans Endlich

Keyword(s):

Shear Stress ◽

Tyrosine Kinases ◽

Fluid Shear Stress ◽

Focal Adhesions ◽

Fluid Shear ◽

Cell Contacts ◽

Tk Inhibitors ◽

Static Conditions ◽

Cell Cell

Podocytes are exposed to mechanical forces arising from glomerular capillary pressure and filtration. It has been shown that stretch affects podocyte biology in vitro and plays a significant role in the development of glomerulosclerosis in vivo. However, whether podocytes are sensitive to fluid shear stress is completely unknown. In the present study, we therefore exposed cells of a recently generated conditionally immortalized mouse podocyte cell line to defined fluid shear stress in a flow chamber, mimicking flow of the glomerular ultrafiltrate over the surface of podocytes in Bowman's space. Shear stress above 0.25 dyne/cm2 resulted in dramatic loss of podocytes but not of proximal tubular epithelial cells (LLC-PK1 cells) after 20 h. At 0.015–0.25 dyne/cm2, lamellipodia formation in podocytes was enhanced and the actin nucleation protein cortactin was redistributed to the cell margins. Shear stress further diminished stress fibers and the presence of vinculin in focal adhesions. Linear zonula occludens-1 distribution at cell-cell contacts remained unaffected at low shear stress. At 0.25 dyne/cm2, the monolayer was broken up and remaining cell-cell contacts were reinforced by F-actin and α-actinin. Because the cytoskeletal changes induced by shear stress suggested the involvement of tyrosine kinases (TKs), we tested several TK inhibitors that were all without effect on podocyte number under static conditions. At 0.25 dyne/cm2, however, the TK inhibitors genistein and AG 82 were associated with marked podocyte loss. Our data demonstrate that podocytes are highly sensitive to fluid shear stress. Shear stress induces a reorganization of the actin cytoskeleton and activates specific tyrosine kinases that are required to withstand fluid shear stress.

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The eosinophil actin cytoskeleton undergoes rapid rearrangement in response to fluid shear stress

Journal of Leukocyte Biology ◽

10.1002/jlb.1ma0320-349rr ◽

2020 ◽

Vol 108 (1) ◽

pp. 129-137

Author(s):

Kiho Son ◽

Mike Small ◽

Roma Sehmi ◽

Luke Janssen

Keyword(s):

Shear Stress ◽

Actin Cytoskeleton ◽

Fluid Shear Stress ◽

Fluid Shear

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Colonization of distant organs by tumor cells generating circulating homotypic clusters adaptive to fluid shear stress

Scientific Reports ◽

10.1038/s41598-021-85743-z ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Manabu Maeshiro ◽

Satoru Shinriki ◽

Rin Liu ◽

Yutaka Nakachi ◽

Yoshihiro Komohara ◽

...

Keyword(s):

Shear Stress ◽

Tumor Cells ◽

Fluid Shear Stress ◽

Dna Barcode ◽

Dependent Manner ◽

Fluid Shear ◽

Cortical Actin ◽

Metastatic Recurrence ◽

Static Conditions ◽

E Cadherin

AbstractOnce disseminated tumor cells (DTCs) arrive at a metastatic organ, they remain there, latent, and become seeds of metastasis. However, the clonal composition of DTCs in a latent state remains unclear. Here, we applied high-resolution DNA barcode tracking to a mouse model that recapitulated the metastatic dormancy of head and neck squamous cell carcinoma (HNSCC). We found that clones abundantly circulated peripheral blood dominated DTCs. Through analyses of multiple barcoded clonal lines, we identified specific subclonal population that preferentially generated homotypic circulating tumor cell (CTC) clusters and dominated DTCs. Despite no notable features under static conditions, this population significantly generated stable cell aggregates that were resistant to anoikis under fluid shear stress (FSS) conditions in an E-cadherin-dependent manner. Our data from various cancer cell lines indicated that the ability of aggregate-constituting cells to regulate cortical actin-myosin dynamics governed the aggregates’ stability in FSS. The CTC cluster-originating cells were characterized by the expression of a subset of E-cadherin binding factors enriched with actin cytoskeleton regulators. Furthermore, this expression signature was associated with locoregional and metastatic recurrence in HNSCC patients. These results reveal a biological selection of tumor cells capable of generating FSS-adaptive CTC clusters, which leads to distant colonization.

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Effect of Fluid Shear Stress on Endocytosis of Heparan Sulfate and Low-density Lipoproteins

Journal of Biomedicine and Biotechnology ◽

10.1155/2007/65136 ◽

2007 ◽

Vol 2007 ◽

pp. 1-8 ◽

Cited By ~ 8

Author(s):

Irmeli Barkefors ◽

Cyrus K. Aidun ◽

E. M. Ulrika Egertsdotter

Keyword(s):

Shear Stress ◽

Heparan Sulfate ◽

Fluid Shear Stress ◽

Critical Factor ◽

Low Density Lipoproteins ◽

Confocal Imaging ◽

Low Density ◽

Fluid Shear ◽

Main Component ◽

Static Conditions

Hemodynamic stress is a critical factor in the onset of atherosclerosis such that reduced rates of shear stress occurring at regions of high curvature are more prone to disease. The level of shear stress has direct influence on the thickness and integrity of the glycocalyx layer. Here we show that heparan sulfate, the main component of the glycocalyx layer, forms an intact layer only on cell surfaces subjected to shear, and not under static conditions. Furthermore, receptor-mediated endocytosis of heparan sulfate and low-density liporoteins is not detectable in cells exposed to shear stress. The internalized heparan sulfate and low-density lipoproteins are colocalized as shown by confocal imaging.

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