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 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.

ID: 126: ROLE OF SYNDECAN-1 IN MECHANICAL STRESS-INDUCED LUNG ENDOTHELIAL CELL INFLAMMATORY RESPONSES MEDIATED BY INTEGRIN BETA4

2016 ◽  
Vol 64 (4) ◽  
pp. 972.2-973
Author(s):  
W Chen ◽  
RO Dull ◽  
JR Jacobson
Keyword(s):  
Endothelial Cell ◽  
Lung Injury ◽  
Mechanical Stress ◽  
Tyrosine Phosphorylation ◽  
Western Blotting ◽  
Inflammatory Responses ◽  
Cyclic Stretch ◽  
Protective Effects ◽  
Coa Reductase

RationaleSimvastatin, an HMG-CoA reductase inhibitor, has protective effects on mechanically stressed human lung endothelial cells (EC) that are mediated by the attenuation of agonist-induced integrin β4 (ITGB4) tyrosine phosphorylation. In addition, overexpression of ITGB4 constructs harboring mutations in tyrosine phosphorylation sites within the cytoplasmic tail results in decreased mechanical stress-induced inflammatory cytokine release by EC. However, the mechanisms by which ITGB4 phosphorylation is regulated is unknown. A molecule of interest in this context is syndecan-1, a cell-surface proteoglycan that binds extraceullar matrix components but also binds the ITGB4 cytoplasmic domain, is expressed by EC, and has been implicated as a mediator of acute lung injury (ALI). Thus, we hypothesized that syndecan-1 is an effector of lung endothelial cell inflammatory responses mediated by ITGB4 in response to mechanical stress.MethodsTo investigate the effects of simvastatin on ITGB4 and syndecan-1 expression human pulmonary artery EC lysates treated with simvastatin (5 µM, 16 h) were subjected to Western blotting for ITGB4 and syndecan-1. Simvastatin-treated EC were then used for immunoprecipitation (IP) of syndecan-1 followed by Western blotting for ITGB4. To study the role of syndecan-1 in EC inflammatory responses to mechanical stress mediated by ITGB4, EC were transfected with syndecan-1 siRNA prior to cyclic stretch (18% CS) and lysates were collected for immunoprecipitation of ITGB4 followed by Western blotting with a phospho-tyrosine antibody. Finally, syndecan-1-silenced EC were subjected to 18% CS and the media was collected for measurement of IL-6 and IL-8 levels.ResultsSimvastatin treatment of EC resulted in both a dramatic increase in ITGB4 and a marked decreased in syndecan-1 expression levels. In addition, syndecan-1 association with ITGB4 was markedly decreased in EC treated with simvastatin. In EC subjected to 18% CS, ITGB4 phosphorylation was significantly decreased after syndecan-1 knockdown. Finally, silencing of syndecan-1 was associated with a significant decrease in CS-induced EC IL-6 and IL-8 expression (76% and 66%, respectively, p<0.05 for both).ConclusionOur results implicate syndecan-1 as an important mediator of EC ITGB4 tyrosine phosphorylation affected by both simvastatin and mechanical-stress. These findings represent a novel area of investigation that may ultimately yield new therapeutic targets and strategies for patients with ventilator-induced lung injury, a form of ALI precipitated by excessive lung stretch.

scholarly journals Mechanosensitive cation channel Piezo1 contributes to ventilator-induced lung injury by activating RhoA/ROCK1 in rats

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Yang Zhang ◽  
Lulu Jiang ◽  
Tianfeng Huang ◽  
Dahao Lu ◽  
Yue Song ◽  
...  
Keyword(s):  
Mechanical Ventilation ◽  
Lung Injury ◽  
Tidal Volume ◽  
Cyclic Stretch ◽  
High Tidal Volume ◽  
Enzyme Linked Immunosorbent Assay ◽  
Sprague Dawley ◽  
Ventilator Induced Lung Injury ◽  
Enhanced Expression ◽  
Underlying Mechanisms

Abstract Background Mechanical ventilation can induce or aggravate lung injury, which is termed ventilator-induced lung injury (VILI). Piezo1 is a key element of the mechanotransduction process and can transduce mechanical signals into biological signals by mediating Ca2+ influx, which in turn regulates cytoskeletal remodeling and stress alterations. We hypothesized that it plays an important role in the occurrence of VILI, and investigated the underlying mechanisms. Methods High tidal volume mechanical ventilation and high magnitude cyclic stretch were performed on Sprague–Dawley rats, and A549 and human pulmonary microvascular endothelial cells, respectively, to establish VILI models. Immunohistochemical staining, flow cytometry, histological examination, enzyme-linked immunosorbent assay, western blotting, quantitative real-time polymerase chain reaction and survival curves were used to assess the effect of Piezo1 on induction of lung injury, as well as the signaling pathways involved. Results We observed that Piezo1 expression increased in the lungs after high tidal volume mechanical ventilation and in cyclic stretch-treated cells. Mechanistically, we observed the enhanced expression of RhoA/ROCK1 in both cyclic stretch and Yoda1-treated cells, while the deficiency or inhibition of Piezo1 dramatically antagonized RhoA/ROCK1 expression. Furthermore, blockade of RhoA/ROCK1 signaling using an inhibitor did not affect Piezo1 expression. GSMTx4 was used to inhibit Piezo1, which alleviated VILI-induced pathologic changes, water content and protein leakage in the lungs, and the induction of systemic inflammatory mediators, and improved the 7-day mortality rate in the model rats. Conclusions These findings indicate that Piezo1 affects the development and progression of VILI through promotion of RhoA/ROCK1 signaling.

scholarly journals Autophagy inhibitor 3-methyladenine protects against endothelial cell barrier dysfunction in acute lung injury

2018 ◽  
Vol 314 (3) ◽  
pp. L388-L396 ◽  
Author(s):  
Spencer A. Slavin ◽  
Antony Leonard ◽  
Valerie Grose ◽  
Fabeha Fazal ◽  
Arshad Rahman
Keyword(s):  
Acute Lung Injury ◽  
Endothelial Cell ◽  
Lung Injury ◽  
Vascular Injury ◽  
Barrier Dysfunction ◽  
Neutrophil Sequestration ◽  
Barrier Disruption ◽  
Proinflammatory Mediators ◽  
Autophagy Inhibitor

Autophagy is an evolutionarily conserved cellular process that facilitates the continuous recycling of intracellular components (organelles and proteins) and provides an alternative source of energy when nutrients are scarce. Recent studies have implicated autophagy in many disorders, including pulmonary diseases. However, the role of autophagy in endothelial cell (EC) barrier dysfunction and its relevance in the context of acute lung injury (ALI) remain uncertain. Here, we provide evidence that autophagy is a critical component of EC barrier disruption in ALI. Using an aerosolized bacterial lipopolysaccharide (LPS) inhalation mouse model of ALI, we found that administration of the autophagy inhibitor 3-methyladenine (3-MA), either prophylactically or therapeutically, markedly reduced lung vascular leakage and tissue edema. 3-MA was also effective in reducing the levels of proinflammatory mediators and lung neutrophil sequestration induced by LPS. To test the possibility that autophagy in EC could contribute to lung vascular injury, we addressed its role in the mechanism of EC barrier disruption. Knockdown of ATG5, an essential regulator of autophagy, attenuated thrombin-induced EC barrier disruption, confirming the involvement of autophagy in the response. Similarly, exposure of cells to 3-MA, either before or after thrombin, protected against EC barrier dysfunction by inhibiting the cleavage and loss of vascular endothelial cadherin at adherens junctions, as well as formation of actin stress fibers. 3-MA also reversed LPS-induced EC barrier disruption. Together, these data imply a role of autophagy in lung vascular injury and reveal the protective and therapeutic utility of 3-MA against ALI.

scholarly journals Pre-treatment with Isoflurane or Sevoflurane is not protective against high tidal volume induced lung injury in rats

2019 ◽  
Author(s):  
Florian Setzer ◽  
Lars Hueter ◽  
Barbara Schmidt ◽  
Konrad Schwarzkopf ◽  
Torsten Schreiber
Keyword(s):  
Mechanical Ventilation ◽  
Body Weight ◽  
Lung Injury ◽  
Tidal Volume ◽  
Lavage Fluid ◽  
Lung Lavage ◽  
Protective Effects ◽  
Markers Of Inflammation ◽  
In Vivo Animal Model

Abstract Background Volatile anesthetics (VA) may exert organ-protective effects in various experimental and clinical settings. Mechanical ventilation (MV) induces an inflammatory response and, depending on the ventilator settings chosen, injury in the lungs. It is unclear if prophylactic inhaled VA applied on healthy lungs prior to MV are protective regarding these effects.Methods Healthy, spontaneously breathing rats were exposed for 30 minutes to either isoflurane (1.8 Vol %), sevoflurane (3.0 Vol %) or no VA (controls). Animals were allowed to recover and then mechanically ventilated for 4 hours with either high (21 ml/kg body weight) or low (9 ml/kg body weight) tidal volume. Cardiorespiratory parameters and systemic inflammation were assessed at the beginning and during mechanical ventilation. Cellular, non-cellular and histologic markers of pulmonary injury and inflammation were determined.Results Irrespective of VA pretreatment, MV with high VT negatively affected markers of lung integrity such as arterial oxygenation and lung wet-to-dry ratio. Regarding the application of VA pretreatment protective effects on lung function were absent but there were changes in some markers of inflammation such as a decrease in blood lymphocyte counts and an increase in interleukin 6 concentration in plasma and in lung lavage fluid. These effects were heterogeneous regarding group allocation and time points.Conclusions In this in in vivo animal model, prophylactic administration of inhaled VA was not beneficial or protective regarding ventilation induced lung injury. However, there were effects suggestive of a modulation of inflammatory markers associated with VA prophylaxis. The clinical or biological relevance of these findings so far remain unclear and should be subject to further studies.

scholarly journals Mechanosensitive Cation Channel Piezo1 Contributes To Ventilator-Induced Lung Injury By Activating RhoA/ROCK1 In Rats

2021 ◽  
Author(s):  
Yang Zhang ◽  
Lulu Jiang ◽  
Tianfeng Huang ◽  
Dahao Lu ◽  
Yue Song ◽  
...  
Keyword(s):  
Mechanical Ventilation ◽  
Lung Injury ◽  
Tidal Volume ◽  
Cyclic Stretch ◽  
High Tidal Volume ◽  
Enzyme Linked Immunosorbent Assay ◽  
Sprague Dawley ◽  
Reverse Transcription Pcr ◽  
Ventilator Induced Lung Injury ◽  
Underlying Mechanisms

Abstract Background: Mechanical ventilation can induce or aggravate lung injury, which is termed ventilator‑induced lung injury. Piezo1 is a key element of the mechanotransduction process and can transduce mechanical signals into biological signals by mediating Ca2+ influx, which in turn regulates cytoskeletal remodeling and stress alterations. We hypothesized that it plays an important role in the occurrence of ventilator‑induced lung injury, and we investigated the underlying mechanisms. Methods: High tidal volume mechanical ventilation and high magnitude cyclic stretch were performed on Sprague Dawley rats, and A549 and human pulmonary microvascular endothelial cells, respectively, to establish ventilator‑induced lung injury models. Immunohistochemical staining, flow cytometry, histological examination, enzyme-linked immunosorbent assay, western blotting, quantitative real-time reverse transcription-PCR and survival curves were used to assess the effect of Piezo1 on induction of lung injury, as well as the signaling pathways involved.Results: We observed that Piezo1 expression increased in the lungs after high tidal volume mechanical ventilation and in cyclic stretch-treated cells. Mechanistically, we observed the enhanced expression of RhoA/ROCK1 in both cyclic stretch and Yoda1-treated cells, while the deficiency or inhibition of Piezo1 dramatically antagonized RhoA/ROCK1 expression. Furthermore, blockade of RhoA/ROCK1 signaling using an inhibitor did not affect Piezo1 expression. GSMTx4 was used to inhibit Piezo1, which alleviated ventilator‑induced lung injury-induced pathologic changes, water content and protein leakage in the lungs, and the induction of systemic inflammatory mediators, and improved the 7-day mortality rate in the model rats. Conclusions: These findings indicate that Piezo1 affects the development and progression of ventilator‑induced lung injury through promotion of RhoA/ROCK1 signaling.

Abstract 20538: Nrf2 Signaling Ameliorates Ventilator-Induced Lung Inflammation

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Ting Wang ◽  
Tao Jiang ◽  
Hector Quijada ◽  
Joseph B Mascarenhas ◽  
Venkateswaran Ramamoorthi Elangovan ◽  
...  
Keyword(s):  
Gene Expression ◽  
Endothelial Cell ◽  
Lung Injury ◽  
Tidal Volume ◽  
Mechanical Stress ◽  
Therapeutic Target ◽  
Lung Inflammation ◽  
Cyclic Stretch ◽  
Gene Expression Dysregulation ◽  
Novel Therapeutic

Ventilator-induced lung injury (VILI) is an acute lung injury that develops during mechanical ventilation with high tidal volume in patients in the intensive care unit (ICU). VILI is a common entity with an unacceptable mortality rate (~20%). As no current therapies exist for VILI, novel therapeutic targets for VILI and mechanical stress-induced tissue injury are needed. In this study, we defined Nrf2 signaling in VILI and mechanical stress-induced endothelial cell pathobiology. Nrf2 is a transcription factor that alters ROS balance and is a novel therapeutic target for VILI. First, we defined Nrf2 as a sensory marker for mechanical stress-induced endothelial cell injury. Antioxidant response element (ARE) luciferase plasmids were transfected to both endothelial cells and murine lung tissues. EC exposed to 18% cyclic stretch (18% CS) or mice exposed to excessive tidal volume ventilation (40 ml/kg) resulted in ARE activation, site of Nrf2 binding. We next confirmed Nrf2 signaling is essential for gene expression dysregulation by 18% CS via Nrf2 knockdown that significantly ameliorates 18% CS-induced gene expression pattern, including signaling pathways regulating endothelial barrier function including MYLK. Finally, we validated that Nrf2 is a novel and effective therapeutic target for VILI in pre-clinical models of VILI. Nrf2 activator sulforaphane (SF, 11.5 mg/kg) significantly attenuated VILI (40 ml/kg, 4 hrs)-induced lung inflammation, including BAL protein leakage and white blood cell infiltration. These novel findings characterized Nrf2 as a sensor for mechanical stress, a central mediator of mechanical stress induced gene expression dysregulation, and a novel therapeutic target for VILI.

Role of vasodilator-stimulated phosphoprotein in cGMP-mediated protection of human pulmonary artery endothelial barrier function

2008 ◽  
Vol 294 (4) ◽  
pp. L686-L697 ◽  
Author(s):  
Otgonchimeg Rentsendorj ◽  
Tamara Mirzapoiazova ◽  
Djanybek Adyshev ◽  
Laura E. Servinsky ◽  
Thomas Renné ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Pulmonary Artery ◽  
Actin Polymerization ◽  
Cell Junctions ◽  
Endothelial Barrier ◽  
Protective Effects ◽  
Barrier Dysfunction ◽  
Endothelial Barrier Dysfunction ◽  
Human Pulmonary Artery

Increased pulmonary endothelial cGMP was shown to prevent endothelial barrier dysfunction through activation of protein kinase G (PKGI). Vasodilator-stimulated phosphoprotein (VASP) has been hypothesized to mediate PKGI barrier protection because VASP is a cytoskeletal phosphorylation target of PKGI expressed in cell-cell junctions. Unphosphorylated VASP was proposed to increase paracellular permeability through actin polymerization and stress fiber bundling, a process inhibited by PKGI-mediated phosphorylation of Ser157 and Ser239. To test this hypothesis, we examined the role of VASP in the transient barrier dysfunction caused by H2O2 in human pulmonary artery endothelial cell (HPAEC) monolayers studied without and with PKGI expression introduced by adenoviral infection (Ad.PKG). In the absence of PKGI expression, H2O2 (100–250 μM) caused a transient increased permeability and pSer157-VASP formation that were both attenuated by protein kinase C inhibition. Potentiation of VASP Ser157 phosphorylation by either phosphatase 2B inhibition with cyclosporin or protein kinase A activation with forskolin prolonged, rather than inhibited, the increased permeability caused by H2O2. With Ad.PKG infection, inhibition of VASP expression with small interfering RNA exacerbated H2O2-induced barrier dysfunction but had no effect on cGMP-mediated barrier protection. In addition, expression of a Ser-double phosphomimetic mutant VASP failed to reproduce the protective effects of activated PKGI. Finally, expression of a Ser-double phosphorylation-resistant VASP failed to interfere with the ability of cGMP/PKGI to attenuate H2O2-induced disruption of VE-cadherin homotypic binding. Our results suggest that VASP phosphorylation does not explain the protective effect of cGMP/PKGI on H2O2-induced endothelial barrier dysfunction in HPAEC.

scholarly journals A single, 30 minutes pretreatment with Isoflurane or Sevoflurane is not protective against high tidal volume induced lung injury in rats

2020 ◽  
Author(s):  
Florian Setzer ◽  
Lars Hueter ◽  
Barbara Schmidt ◽  
Konrad Schwarzkopf ◽  
Torsten Schreiber
Keyword(s):  
Mechanical Ventilation ◽  
Body Weight ◽  
Lung Injury ◽  
Tidal Volume ◽  
Lavage Fluid ◽  
Lung Lavage ◽  
Protective Effects ◽  
Markers Of Inflammation ◽  
In Vivo Animal Model

Abstract Background: Volatile anesthetics (VA) may exert organ-protective effects in various experimental and clinical settings. Mechanical ventilation (MV) induces an inflammatory response and, depending on the ventilator settings chosen, injury in the lungs. It is unclear if prophylactic inhaled VA applied on healthy lungs prior to MV are protective regarding these effects. Methods: Healthy, spontaneously breathing rats were exposed for 30 minutes to either isoflurane (1.8 Vol %), sevoflurane (3.0 Vol %) or no VA. Animals were allowed to recover, intraperitoneally anesthetized and then mechanically ventilated for 4 hours with either high (21 ml/kg body weight) or low (9 ml/kg body weight) tidal volume (n = 12 per group). Cardiorespiratory parameters and systemic inflammation were assessed at the beginning and during mechanical ventilation. Cellular, non-cellular and histologic markers of pulmonary injury and inflammation were determined. Results: Irrespective of VA pretreatment, MV with high VT negatively affected markers of lung integrity such as arterial oxygenation and lung wet-to-dry ratio. Following VA pretreatment we found no protective effects on lung function but there were changes in some markers of inflammation such as a decrease in blood lymphocyte counts and an increase in interleukin 6 concentration in plasma and lung lavage fluid. These effects were heterogeneous regarding group allocation and time points. Conclusions: In this in in vivo animal model, prophylactic administration of inhaled VA was not beneficial or protective regarding ventilation induced lung injury. However, there were effects suggestive of a modulation of inflammatory markers associated with VA prophylaxis. The clinical or biological relevance of these findings so far remain unclear and should be subject to further studies.

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