scholarly journals Magnitude-dependent effects of cyclic stretch on HGF- and VEGF-induced pulmonary endothelial remodeling and barrier regulation

2008 ◽  
Vol 295 (4) ◽  
pp. L612-L623 ◽  
Author(s):  
Anna A. Birukova ◽  
Nurgul Moldobaeva ◽  
Junjie Xing ◽  
Konstantin G. Birukov
Keyword(s):  
Growth Factor ◽  
Lung Injury ◽  
Synergistic Effects ◽  
Endothelial Permeability ◽  
Small Gtpase ◽  
Cyclic Stretch ◽  
Dependent Manner ◽  
Protective Effects ◽  
Gap Formation ◽  
Barrier Dysfunction

Mechanical ventilation at high tidal volumes compromises the blood-gas barrier and increases lung vascular permeability, which may lead to ventilator-induced lung injury and pulmonary edema. Using pulmonary endothelial cell (ECs) exposed to physiologically [5% cyclic stretch (CS)] and pathologically (18% CS) relevant magnitudes of CS, we evaluated the potential protective effects of hepatocyte growth factor (HGF) on EC barrier dysfunction induced by CS and vascular endothelial growth factor (VEGF). In static culture, HGF enhanced EC barrier function in a Rac-dependent manner and attenuated VEGF-induced EC permeability and paracellular gap formation. The protective effects of HGF were associated with the suppression of Rho-dependent signaling triggered by VEGF. Five percent CS promoted HGF-induced enhancement of the cortical F-actin rim and activation of Rac-dependent signaling, suggesting synergistic barrier-protective effects of physiological CS and HGF. In contrast, 18% CS further enhanced VEGF-induced EC permeability, activation of Rho signaling, and formation of actin stress fibers and paracellular gaps. These effects were attenuated by HGF pretreatment. EC preconditioning at 5% CS before HGF and VEGF further promoted EC barrier maintenance. Our data suggest synergistic effects of HGF and physiological CS in the Rac-mediated mechanisms of EC barrier protection. In turn, HGF reduced the barrier-disruptive effects of VEGF and pathological CS via downregulation of the Rho pathway. These results support the importance of HGF-VEGF balance in control of acute lung injury/acute respiratory distress syndrome severity via small GTPase-dependent regulation of lung endothelial permeability.

scholarly journals AM966, an Antagonist of Lysophosphatidic Acid Receptor 1, Increases Lung Microvascular Endothelial Permeability through Activation of Rho Signaling Pathway and Phosphorylation of VE-Cadherin

2017 ◽  
Vol 2017 ◽  
pp. 1-12 ◽  
Author(s):  
Junting Cai ◽  
Jianxin Wei ◽  
Shuang Li ◽  
Tomeka Suber ◽  
Jing Zhao
Keyword(s):  
Endothelial Cells ◽  
Lung Injury ◽  
Lysophosphatidic Acid ◽  
Endothelial Permeability ◽  
Endothelial Barrier ◽  
Dependent Manner ◽  
Gap Formation ◽  
Barrier Dysfunction ◽  
Lpa Receptor ◽  
Microvascular Endothelial

Maintenance of pulmonary endothelial barrier integrity is important for reducing severity of lung injury. Lysophosphatidic acid (LPA) regulates cell motility, cytoskeletal rearrangement, and cell growth. Knockdown of LPA receptor 1 (LPA1) has been shown to mitigate lung injury and pulmonary fibrosis. AM966, an LPA1 antagonist exhibiting an antifibrotic property, has been considered to be a future antifibrotic medicine. Here, we report an unexpected effect of AM966, which increases lung endothelial barrier permeability. An electric cell-substrate sensing (ECIS) system was used to measure permeability in human lung microvascular endothelial cells (HLMVECs). AM966 decreased the transendothelial electrical resistance (TEER) value immediately in a dose-dependent manner. VE-cadherin and f-actin double immunostaining reveals that AM966 increases stress fibers and gap formation between endothelial cells. AM966 induced phosphorylation of myosin light chain (MLC) through activation of RhoA/Rho kinase pathway. Unlike LPA treatment, AM966 had no effect on phosphorylation of extracellular signal-regulated kinases (Erk). Further, in LPA1 silencing cells, we observed that AM966-increased lung endothelial permeability as well as phosphorylation of VE-cadherin and focal adhesion kinase (FAK) were attenuated. This study reveals that AM966 induces lung endothelial barrier dysfunction, which is regulated by LPA1-mediated activation of RhoA/MLC and phosphorylation of VE-cadherin.

scholarly journals Mechanosensitive Rap1 activation promotes barrier function of lung vascular endothelium under cyclic stretch

2019 ◽  
Vol 30 (8) ◽  
pp. 959-974 ◽  
Author(s):  
Yunbo Ke ◽  
Pratap Karki ◽  
Chenou Zhang ◽  
Yue Li ◽  
Trang Nguyen ◽  
...  
Keyword(s):  
Mechanical Ventilation ◽  
Endothelial Cell ◽  
Lung Injury ◽  
Tidal Volume ◽  
Small Gtpase ◽  
Cyclic Stretch ◽  
Cell Junctions ◽  
Protective Effects ◽  
Barrier Dysfunction

Mechanical ventilation remains an imperative treatment for the patients with acute respiratory distress syndrome, but can also exacerbate lung injury. We have previously described a key role of RhoA GTPase in high cyclic stretch (CS)–induced endothelial cell (EC) barrier dysfunction. However, cellular mechanotransduction complexes remain to be characterized. This study tested a hypothesis that recovery of a vascular EC barrier after pathologic mechanical stress may be accelerated by cell exposure to physiologic CS levels and involves Rap1-dependent rearrangement of endothelial cell junctions. Using biochemical, molecular, and imaging approaches we found that EC pre- or postconditioning at physiologically relevant low-magnitude CS promotes resealing of cell junctions disrupted by pathologic, high-magnitude CS. Cytoskeletal remodeling induced by low CS was dependent on small GTPase Rap1. Protective effects of EC preconditioning at low CS were abolished by pharmacological or molecular inhibition of Rap1 activity. In vivo, using mice exposed to mechanical ventilation, we found that the protective effect of low tidal volume ventilation against lung injury caused by lipopolysaccharides and ventilation at high tidal volume was suppressed in Rap1 knockout mice. Taken together, our results demonstrate a prominent role of Rap1-mediated signaling mechanisms activated by low CS in acceleration of lung vascular EC barrier restoration.

scholarly journals Cyclic stretch induces alveolar epithelial barrier dysfunction via calpain-mediated degradation of p120-catenin

2011 ◽  
Vol 301 (2) ◽  
pp. L197-L206 ◽  
Author(s):  
Yuelan Wang ◽  
Richard D. Minshall ◽  
David E. Schwartz ◽  
Guochang Hu
Keyword(s):  
Lung Injury ◽  
Alveolar Epithelial Cells ◽  
Cyclic Stretch ◽  
Epithelial Barrier ◽  
Gap Formation ◽  
Barrier Dysfunction ◽  
P120 Catenin ◽  
Alveolar Epithelial ◽  
Ventilator Induced Lung Injury ◽  
Epithelial Barrier Dysfunction

Lung hyperinflation is known to be an important contributing factor in the pathogenesis of ventilator-induced lung injury. Mechanical stretch causes epithelial barrier dysfunction and an increase in alveolar permeability, although the precise mechanisms have not been completely elucidated. p120-catenin is an adherens junction-associated protein that regulates cell-cell adhesion. In this study, we determined the role of p120-catenin in cyclic stretch-induced alveolar epithelial barrier dysfunction. Cultured alveolar epithelial cells (MLE-12) were subjected to uniform cyclic (0.5 Hz) biaxial stretch from 0 to 8 or 20% change in surface area for 0, 1, 2, or 4 h. At the end of the experiments, cells were lysed to determine p120-catenin expression by Western blot analysis. Immunofluorescence staining of p120-catenin and F-actin was performed to assess the integrity of monolayers and interepithelial gap formation. Compared with unstretched control cells, 20% stretch caused a significant loss in p120-catenin expression, which was coupled to interepithelial gap formation. p120-Catenin knockdown with small interfering RNA (siRNA) dose dependently increased stretch-induced gap formation, whereas overexpression of p120-catenin abolished stretch-induced gap formation. Furthermore, pharmacological calpain inhibition or depletion of calpain-1 with a specific siRNA prevented p120-catenin loss and subsequent stretch-induced gap formation. Our findings demonstrate that p120-catenin plays a critical protective role in cyclic stretch-induced alveolar barrier dysfunction, and, thus, maintenance of p120-catenin expression may be a novel therapeutic strategy for the prevention and treatment of ventilator-induced lung injury.

ID: 114: AN ANTAGONIST OF LYSOPHOSPHATIDIC ACID RECEPTOR1, AM966, INCREASES PULMONARY ENDOTHELIAL PERMEABILITY THROUGH ACTIVATION OF VE-CADHERIN

2016 ◽  
Vol 64 (4) ◽  
pp. 965.3-966
Author(s):  
J Cai ◽  
J Wei ◽  
AM Jacko ◽  
J Zhao
Keyword(s):  
Lysophosphatidic Acid ◽  
Barrier Function ◽  
Endothelial Permeability ◽  
Heart Association ◽  
Endothelial Barrier ◽  
Dependent Manner ◽  
Gap Formation ◽  
Endothelial Barrier Function ◽  
Barrier Dysfunction ◽  
Barrier Integrity

BackgroundMaintenance of pulmonary endothelial barrier integrity is important for reducing severity of lung injury. VE-cadherin is a major component of cell–cell adherens junctions in endothelium. In response to inflammatory stimuli, VE-cadherin is tyrosine phosphorylated, resulting in dissociation with catenins, which links to f-actin. Lysophosphatidic acid (LPA) is a bioactive lysophospholipid, which regulates cell motility. LPA has been shown to increase lung epithelial barrier integrity, while it reduces endothelial barrier function. AM966 is an antagonist exhibiting an anti-fibrotic property. However, the effect of AM966 on pulmonary endothelial barrier integrity has not been well studied.Methods and ResultsTo investigate endothelial barrier integrity, electric cell-substrate sensing (ECIS) system was used to measure permeability in human lung microvascular endothelial cells (HLMVECs). Similar to the effect of LPA, AM966 increases permeability immediately in a dose dependent manner. To investigate the molecular mechanism by which regulates AM966-mediated reduction of endothelial barrier function, HLMVECs were treated with AM966, and then phosphorylation of myosin light chain (MLC) and VE-cadherin were determined by immunoblotting. AM966 increased phosphorylation of MLC and VE-cadherin. VE-cadherin and f-actin double immunostaining revealed that AM966 induces gap formation and f-actin stress fibers as well as dissociation between VE-cadherin and f-actin.ConclusionThis study reveals that AM966 induces lung endothelial barrier dysfunction, which is regulated by phosphorylation of VE-cadherin.This work was supported by the National Institutes of Health (R01GM115389 to J.Z.), American Heart Association 12SDG9050005 (J.Z.), American Lung Association Biomedical Research Grant RG350146 (J.Z.).

Integrin-linked kinase and PINCH: partners in regulation of cell-extracellular matrix interaction and signal transduction

1999 ◽  
Vol 112 (24) ◽  
pp. 4485-4489 ◽  
Author(s):  
C. Wu
Keyword(s):  
Growth Factor ◽  
Protein Kinase ◽  
Signaling Pathways ◽  
Focal Adhesion ◽  
Adaptor Protein ◽  
Small Gtpase ◽  
Beta Catenin ◽  
Dependent Manner ◽  
Mesenchymal Transition ◽  
Integrin Linked Kinase

Integrin-linked kinase (ILK) is a focal adhesion serine/threonine protein kinase that is emerging as a key signaling protein functioning at one of the early convergence points of integrin- and growth factor-signaling pathways. ILK binds to PINCH through the N-terminal ankyrin (ANK) repeat domain and the PINCH binding is crucial for focal adhesion localization of ILK. The ILK-PINCH interaction also connects ILK to Nck-2, an SH2-SH3-containing adaptor protein that interacts with components of growth factor and small GTPase signaling pathways. The kinase activity of ILK is regulated by both cell adhesion and growth factors in a phosphoinositide 3-kinase (PI3K)-dependent manner. ILK phosphorylates downstream targets such as protein kinase B (PKB, also known as Akt) and glycogen synthase kinase 3 (GSK-3) and regulates their activities. Overexpression of ILK in epithelial cells leads to striking morphological changes mimicking epithelial-mesenchymal transition, including upregulation of integrin-mediated fibronectin matrix assembly and downregulation of cell-cell adhesions. Furthermore, ILK regulates nuclear translocation of (beta)-catenin and gene expression, and promotes cell cycle progression and tumor formation. Recent genetic studies in Drosophila melanogaster and Caenorhabditis elegans have shown that lack of expression of ILK or PINCH results in phenotypes resembling those of integrin-null mutants, which demonstrates that ILK and PINCH are indispensable for integrin function during embryonic development.

SEB is cytotoxic and alters EC barrier function through protein tyrosine phosphorylation in vitro

1997 ◽  
Vol 273 (1) ◽  
pp. L31-L39 ◽  
Author(s):  
W. N. Campbell ◽  
M. Fitzpatrick ◽  
X. Ding ◽  
M. Jett ◽  
P. Gemski ◽  
...  
Keyword(s):  
Barrier Function ◽  
Tyrosine Kinases ◽  
Polymyxin B ◽  
Time Dependent ◽  
Dependent Manner ◽  
Gap Formation ◽  
Barrier Dysfunction ◽  
Effector Cells ◽  
Protein Tyrosine

We studied whether Staphylococcal enterotoxin B (SEB) has direct effects on endothelial cells (EC) in the absence of effector cells or their products. Bovine or human pulmonary artery EC were grown to confluence on filters mounted in chemotaxis chambers. Barrier function was assessed by placing [14C]bovine serum albumin in the chamber and sampling the lower well for 14C activity. SEB exposures induced a significant (P < 0.001) dose- and time-dependent increase in albumin flux across both bovine and human EC monolayers. Albumin flux was temperature dependent, and cycloheximide pretreatment of the monolayers did not block the SEB-induced increase in permeability. Preincubation of SEB with trypsin or anti-SEB antibody significantly (P < 0.0001) reduced the effect, whereas pretreatment with polymyxin B did not. SEB at > or = 10 micrograms/ml significantly (P < 0.03) increased EC injury as measured by 51Cr release in a dose- and time-dependent manner. Herbimycin and genistein, inhibitors of protein tyrosine kinases, each protected against SEB-induced cytotoxicity, barrier dysfunction, and intercellular gap formation. We conclude that SEB perturbs endothelial barrier function and viability in the absence of effector cells or their mediators.

scholarly journals NEMO-Binding Domain Peptide Attenuates Lipopolysaccharide-Induced Acute Lung Injury by Inhibiting the NF-κB Signaling Pathway

2016 ◽  
Vol 2016 ◽  
pp. 1-11 ◽  
Author(s):  
Jianhua Huang ◽  
Li Li ◽  
Weifeng Yuan ◽  
Linxin Zheng ◽  
Zhenhui Guo ◽  
...  
Keyword(s):  
Acute Lung Injury ◽  
Lung Injury ◽  
Signaling Pathway ◽  
Binding Domain ◽  
Dependent Manner ◽  
Protective Effects ◽  
Lps Challenge ◽  
Domain Peptide ◽  
Nemo Binding Domain ◽  
Dose Dependent

The aim of the present study is to investigate the protective effects and relevant mechanisms exerted by NEMO-binding domain peptide (NBD) against lipopolysaccharide- (LPS-) induced acute lung injury (ALI) in mice. The ALI model was induced by intratracheally administered atomized LPS (5 mg/kg) to BABL/c mice. Half an hour before LPS administration, we treated the mice with increasing concentrations of intratracheally administered NBD or saline aerosol. Two hours after LPS administration, each group of mice was sacrificed. We observed that NBD pretreatment significantly attenuated LPS-induced lung histopathological injury in a dose-dependent manner. Western blotting established that NBD pretreatment obviously attenuated LPS-induced IκB-αand NF-κBp65 activation and NOX1, NOX2, and NOX4 overexpression. Furthermore, NBD pretreatment increased SOD and T-AOC activity and decreased MDA levels in lung tissue. In addition, NBD also inhibited TNF-αand IL-1βsecretion in BALF after LPS challenge. In conclusion, NBD protects against LPS-induced ALI in mice.

Magnitude-dependent regulation of pulmonary endothelial cell barrier function by cyclic stretch

2003 ◽  
Vol 285 (4) ◽  
pp. L785-L797 ◽  
Author(s):  
Konstantin G. Birukov ◽  
Jeffrey R. Jacobson ◽  
Alejandro A. Flores ◽  
Shui Q. Ye ◽  
Anna A. Birukova ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Map Kinases ◽  
Critical Role ◽  
Barrier Properties ◽  
Small Gtpase ◽  
Cyclic Stretch ◽  
Cell Permeability ◽  
Dependent Manner ◽  
Sphingosine 1 Phosphate ◽  
Mlc Phosphorylation

Ventilator-induced lung injury syndromes are characterized by profound increases in vascular leakiness and activation of inflammatory processes. To explore whether excessive cyclic stretch (CS) directly causes vascular barrier disruption or enhances endothelial cell sensitivity to edemagenic agents, human pulmonary artery endothelial cells (HPAEC) were exposed to physiologically (5% elongation) or pathologically (18% elongation) relevant levels of strain. CS produced rapid (10 min) increases in myosin light chain (MLC) phosphorylation, activation of p38 and extracellular signal-related kinase 1/2 MAP kinases, and actomyosin remodeling. Acute (15 min) and chronic (48 h) CS markedly enhanced thrombin-induced MLC phosphorylation (2.1-fold and 3.2-fold for 15-min CS at 5 and 18% elongation and 2.1-fold and 3.1-fold for 48-h CS at 5 and 18% elongation, respectively). HPAEC preconditioned at 18% CS, but not at 5% CS, exhibited significantly enhanced thrombin-induced reduction in transendothelial electrical resistance but did not affect barrier protective effect of sphingosine-1-phosphate (0.5 μM). Finally, expression profiling analysis revealed a number of genes, including small GTPase rho, apoptosis mediator ZIP kinase, and proteinase activated receptor-2, to be regulated by CS in an amplitude-dependent manner. Thus our study demonstrates a critical role for the magnitude of CS in regulation of agonist-mediated pulmonary endothelial cell permeability and strongly suggests phenotypic regulation of HPAEC barrier properties by CS.

scholarly journals Regulation of lung endothelial permeability and inflammatory responses by prostaglandin A2: role of EP4 receptor

2017 ◽  
Vol 28 (12) ◽  
pp. 1622-1635 ◽  
Author(s):  
Tomomi Ohmura ◽  
Yufeng Tian ◽  
Nicolene Sarich ◽  
Yunbo Ke ◽  
Angelo Meliton ◽  
...  
Keyword(s):  
Inflammatory Responses ◽  
Endothelial Permeability ◽  
Protective Effects ◽  
Barrier Dysfunction ◽  
P120 Catenin ◽  
Ep4 Receptor ◽  
Rac1 Gtpase ◽  
Prostaglandin A2

The role of prostaglandin A2 (PGA2) in modulation of vascular endothelial function is unknown. We investigated effects of PGA2 on pulmonary endothelial cell (EC) permeability and inflammatory activation and identified a receptor mediating these effects. PGA2 enhanced the EC barrier and protected against barrier dysfunction caused by vasoactive peptide thrombin and proinflammatory bacterial wall lipopolysaccharide (LPS). Receptor screening using pharmacological and molecular inhibitory approaches identified EP4 as a novel PGA2 receptor. EP4 mediated barrier-protective effects of PGA2 by activating Rap1/Rac1 GTPase and protein kinase A targets at cell adhesions and cytoskeleton: VE-cadherin, p120-catenin, ZO-1, cortactin, and VASP. PGA2 also suppressed LPS-induced inflammatory signaling by inhibiting the NFκB pathway and expression of EC adhesion molecules ICAM1 and VCAM1. These effects were abolished by pharmacological or molecular inhibition of EP4. In vivo, PGA2 was protective in two distinct models of acute lung injury (ALI): LPS-induced inflammatory injury and two-hit ALI caused by suboptimal mechanical ventilation and injection of thrombin receptor–activating peptide. These protective effects were abolished in mice with endothelial-specific EP4 knockout. The results suggest a novel role for the PGA2–EP4 axis in vascular EC protection that is critical for improvement of pathological states associated with increased vascular leakage and inflammation.

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