Differential effects of shear stress and cyclic stretch on focal adhesion remodeling, site-specific FAK phosphorylation, and small GTPases in human lung endothelial cells

Division of Pulmonary and Critical Care Medicine, Johns Hopkins University, School of Medicine, 5200 Eastern Avenue, Mason F. Lord Building, Center Tower, Room 677, Baltimore, MD 21224, USA.
Experimental Cell Research (Impact Factor: 3.25). 04/2005; 304(1):40-9. DOI: 10.1016/j.yexcr.2004.11.001
Source: PubMed


Regulation of endothelial cell (EC) permeability by bioactive molecules is associated with specific patterns of cytoskeletal and cell contact remodeling. A role for mechanical factors such as shear stress (SS) and cyclic stretch (CS) in cytoskeletal rearrangements and regulation of EC permeability becomes increasingly recognized. This paper examined redistribution of focal adhesion (FA) proteins, site-specific focal adhesion kinase (FAK) phosphorylation, small GTPase activation and barrier regulation in human pulmonary EC exposed to laminar shear stress (15 dyn/cm2) or cyclic stretch (18% elongation) in vitro. SS caused peripheral accumulation of FAs, whereas CS induced randomly distributed FAs attached to the ends of newly formed stress fibers. SS activated small GTPase Rac without effects on Rho, whereas 18% CS activated without effect on Rac. SS increased transendothelial electrical resistance (TER) in EC monolayers, which was further elevated by barrier-protective phospholipid sphingosine 1-phosphate. Finally, SS induced FAK phosphorylation at Y576, whereas CS induced FAK phosphorylation at Y397 and Y576. These results demonstrate for the first time differential effects of SS and CS on Rho and Rac activation, FA redistribution, site-specific FAK phosphorylation, and link them with SS-mediated barrier enhancement. Thus, our results suggest common signaling and cytoskeletal mechanisms shared by mechanical and chemical factors involved in EC barrier regulation.

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    • "For two other endothelial genes, less reduction occurred. In this set of materials, our research confirms the previous issues that different loadings are being sensed by different EC mechanoreceptors and cause different cell responses (Park, Chu, 2004, Shikata et al. , 2005). Previous researches have shown that UNCS strongly influences cell orientation and spreading although no direct measurement of cell height has been performed (Hsu et al. , 2009, Wei et al. , 2008). "
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    ABSTRACT: It has been well established that biomechanical environment can influence functionality of biological cells. There are evidences that show mechanical cyclic stretch can promote smooth muscle cell (SMC) markers in endothelial cells (ECs). The objective of this study was to determine whether mechanical stimuli in the forms of uniaxial and equiaxial cyclic stretches (UNCS and EQCS) can affect endothelial and smooth muscle gene expressions in mRNA level of human umbilical vein endothelial cells (HUVECs). For this purpose, 10% uniaxial UNCS and EQCS (60 cycles/min for 24 h) were applied on HUVECs, and using real-time PCR expressions of three EC specific markers, vascular endothelial growth factor receptor-2 (VEGFR-2, also known as FLK-1) , von Willebrand Factor (vWF) and vascular endothelial-cadherin (VE-cadherin) and two SMC specific genes, α-smooth muscle actin (α-SMA) and smooth muscle myosin heavy chain (SMMHC) were quantified. Moreover, alterations in cell height were analyzed by atomic force microscopy (AFM). Results showed that cyclic UNCS for 24 h downregulated the expression of all EC markers and upregulated the expression of all SMC markers while low effects on HUVECs height were observed. Cyclic EQCS in the same conditions resulted in minor effect on SMC gene expression in HUVECs, while led to strong reduction in vWF with no significant change in other two endothelial genes. Cyclic EQCS considerably elevated cell height. Results proposed that ECs can transdifferentiate to SMC phenotype under specific microenvironmental conditions.
    Cell Biology International 01/2015; 39(6). DOI:10.1002/cbin.10443 · 1.93 Impact Factor
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    • "The mechanochemical environment can significantly affect the severity of ALI/ARDS (Birukov 2009). An 18 % cyclic stretch enhances thrombin-induced Rho activation, whereas a 5 % cyclic stretch promotes Rac activation, critical for the recovery of endothelial barrier function (Shikata et al. 2005; Birukova et al. 2006). Another Rac1-activating agent, hepatocyte growth factor (HGF), has been shown to prevent endothelial barrier dysfunction induced by a cyclic stretch and VEGF (Birukova et al. 2008). "
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    ABSTRACT: Pulmonary endothelial permeability is an important determinant of vascular adaptation to changes in oxygen tension, blood pressure, levels of growth factors or inflammatory cytokines. The Ras homologous (Rho) family of guanosine triphosphate phosphatases (Rho GTPases), key regulators of the actin cytoskeleton, regulate endothelial barrier function in response to a variety of environmental factors and signalling agents via the reorganization of the actin cytoskeleton, changes in receptor trafficking or the phosphorylation of junctional proteins. This review provides a brief summary of recent knowledge on Rho-GTPase-mediated effects on pulmonary endothelial barrier function and focuses in particular on their role in pulmonary vascular disorders, including pulmonary hypertension, chronic obstructive pulmonary disease, acute lung injury and acute respiratory distress syndrome.
    Cell and Tissue Research 03/2014; 355(3). DOI:10.1007/s00441-014-1805-0 · 3.57 Impact Factor
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    • "Small GTP-binding genes play a crucial regulatory role in a number of cellular processes in both plants and ani-mals, such as vesicle-mediated intracellular trafficking, signal transduction, cytoskeletal organization, and cell division [68]. Differential expression of small GTP-binding proteins has been previously reported in various abiotic stresses in plant [68], animal [69], and bacteria [70], however, the functions of Small GTP-binding genes was little known in crustacea. 15 unigenes were enriched in GTPase mediated signal transduction process with a relatively low p-value implied that GTPase mediated signal transduction process plays an important role in Portunus trituberculatus salinity adaptation. "
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    ABSTRACT: The swimming crab, Portunus trituberculatus, which is naturally distributed in the coastal waters of Asia-Pacific countries, is an important farmed species in China. Salinity is one of the most important abiotic factors that influence not only the distribution and abundance of crustaceans, it is also an important factor for artificial propagation of the crab. To better understand the interaction between salinity stress and osmoregulation, we performed a transcriptome analysis in the gills of Portunus trituberculatus challenged with salinity stress, using the Illumina Deep Sequencing technology. We obtained 27,696,835, 28,268,353 and 33,901,271 qualified Illumina read pairs from low salinity challenged (LC), non-challenged (NC), and high salinity challenged (HC) Portunus trituberculatus cDNA libraries, respectively. The overall de novo assembly of cDNA sequence data generated 94,511 unigenes, with an average length of 644 bp. Comparative genomic analysis revealed that 1,705 genes differentially expressed in salinity stress compared to the controls, including 615 and 1,516 unigenes in NC vs LC and NC vs HC respectively. GO functional enrichment analysis results showed some differentially expressed genes were involved in crucial processes related to osmoregulation, such as ion transport processes, amino acid metabolism and synthesis processes, proteolysis process and chitin metabolic process. This work represents the first report of the utilization of the next generation sequencing techniques for transcriptome analysis in Portunus trituberculatus and provides valuable information on salinity adaptation mechanism. Results reveal a substantial number of genes modified by salinity stress and a few important salinity acclimation pathways, which will serve as an invaluable resource for revealing the molecular basis of osmoregulation in Portunus trituberculatus. In addition, the most comprehensive sequences of transcripts reported in this study provide a rich source for identification of novel genes in the crab.
    PLoS ONE 12/2013; 8(12):e82155. DOI:10.1371/journal.pone.0082155 · 3.23 Impact Factor
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