Rap1 Mediates Protective Effects of Iloprost Against Lung Endothelial Barrier Dysfunction and Vascular Leak.

Lung inflammation and alterations in endothelial cell (EC) permeability are key events to development of acute lung injury (ALI). Protective effects of atrial natriuretic peptide (ANP) have been shown against inflammatory signaling and endothelial barrier dysfunction induced by gram-negative bacterial wall liposaccharide. We hypothesized that ANP may possess more general protective effects and attenuate lung inflammation and EC barrier dysfunction by suppressing inflammatory cascades and barrier-disruptive mechanisms shared by gram-negative and gram-positive pathogens. C57BL/6J wild-type or ANP knockout mice (Nppa−/−) were treated with gram-positive bacterial cell wall compounds, Staphylococcus aureus-derived peptidoglycan (PepG) and/or lipoteichoic acid (LTA) (intratracheal, 2.5 mg/kg each), with or without ANP (intravenous, 2 μg/kg). In vitro, human pulmonary EC barrier properties were assessed by morphological analysis of gap formation and measurements of transendothelial electrical resistance. LTA and PepG markedly increased pulmonary EC permeability and activated p38 and ERK1/2 MAP kinases, NF-κB, and Rho/Rho kinase signaling. EC barrier dysfunction was further elevated upon combined LTA and PepG treatment, but abolished by ANP pretreatment. In vivo, LTA and PepG-induced accumulation of protein and cells in the bronchoalveolar lavage fluid, tissue neutrophil infiltration, and increased Evans blue extravasation in the lungs was significantly attenuated by intravenous injection of ANP. Accumulation of bronchoalveolar lavage markers of LTA/PepG-induced lung inflammation and barrier dysfunction was further augmented in ANP−/− mice and attenuated by exogenous ANP injection. These results strongly suggest a protective role of ANP in the in vitro and in vivo models of ALI associated with gram-positive infection. Thus ANP may have important implications in therapeutic strategies aimed at the treatment of sepsis and ALI-induced gram-positive bacterial pathogens.

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Atrial natriuretic peptide attenuates LPS-induced lung vascular leak: role of PAK1

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00202.2009 ◽

2010 ◽

Vol 299 (5) ◽

pp. L652-L663 ◽

Cited By ~ 42

Author(s):

Anna A. Birukova ◽

Junjie Xing ◽

Panfeng Fu ◽

Bakhtiyor Yakubov ◽

Oleksii Dubrovskyi ◽

...

Keyword(s):

Acute Lung Injury ◽

Lung Injury ◽

Atrial Natriuretic Peptide ◽

Bronchoalveolar Lavage ◽

Knockout Mice ◽

Endothelial Barrier ◽

Protective Effects ◽

Barrier Dysfunction ◽

Vascular Leak ◽

Evans Blue Extravasation

Increased levels of atrial natriuretic peptide (ANP) in the models of sepsis, pulmonary edema, and acute respiratory distress syndrome (ARDS) suggest its potential role in the modulation of acute lung injury. We have recently described ANP-protective effects against thrombin-induced barrier dysfunction in pulmonary endothelial cells (EC). The current study examined involvement of the Rac effector p21-activated kinase (PAK1) in ANP-protective effects in the model of lung vascular permeability induced by bacterial wall LPS. C57BL/6J mice or ANP knockout mice (Nppa−/−) were treated with LPS (0.63 mg/kg intratracheal) with or without ANP (2 μg/kg iv). Lung injury was monitored by measurements of bronchoalveolar lavage protein content, cell count, Evans blue extravasation, and lung histology. Endothelial barrier properties were assessed by morphological analysis and measurements of transendothelial electrical resistance. ANP treatment stimulated Rac-dependent PAK1 phosphorylation, attenuated endothelial permeability caused by LPS, TNF-α, and IL-6, decreased LPS-induced cell and protein accumulation in bronchoalveolar lavage fluid, and suppressed Evans blue extravasation in the murine model of acute lung injury. More severe LPS-induced lung injury and vascular leak were observed in ANP knockout mice. In rescue experiments, ANP injection significantly reduced lung injury in Nppa−/− mice caused by LPS. Molecular inhibition of PAK1 suppressed the protective effects of ANP treatment against LPS-induced lung injury and endothelial barrier dysfunction. This study shows that the protective effects of ANP against LPS-induced vascular leak are mediated at least in part by PAK1-dependent signaling leading to EC barrier enhancement. Our data suggest a direct role for ANP in endothelial barrier regulation via modulation of small GTPase signaling.

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mTOR/STAT‐3 pathway mediates mesenchymal stem cell–secreted hepatocyte growth factor protective effects against lipopolysaccharide‐induced vascular endothelial barrier dysfunction and apoptosis

Journal of Cellular Biochemistry ◽

10.1002/jcb.27642 ◽

2018 ◽

Vol 120 (3) ◽

pp. 3637-3650 ◽

Cited By ~ 9

Author(s):

Shan‐Shan Meng ◽

Feng‐Mei Guo ◽

Xi‐Wen Zhang ◽

Wei Chang ◽

Fei Peng ◽

...

Keyword(s):

Stem Cell ◽

Growth Factor ◽

Mesenchymal Stem Cell ◽

Hepatocyte Growth Factor ◽

Endothelial Barrier ◽

Vascular Endothelial ◽

Protective Effects ◽

Barrier Dysfunction ◽

Endothelial Barrier Dysfunction ◽

Hepatocyte Growth

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

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00417.2007 ◽

2008 ◽

Vol 294 (4) ◽

pp. L686-L697 ◽

Cited By ~ 24

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.

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mTORC2 Activation Mediated by Mesenchymal Stem Cell-Secreted Hepatocyte Growth Factors for the Recovery of Lipopolysaccharide-Induced Vascular Endothelial Barrier

Stem Cells International ◽

10.1155/2021/9981589 ◽

2021 ◽

Vol 2021 ◽

pp. 1-12

Author(s):

Shan-Shan Meng ◽

Feng-Mei Guo ◽

Li-Li Huang ◽

Ying-Zi Huang ◽

Jian-Feng Xie ◽

...

Keyword(s):

Cell Proliferation ◽

Stem Cell ◽

Mesenchymal Stem Cell ◽

Endothelial Barrier ◽

Vascular Endothelial ◽

Protective Effects ◽

Barrier Dysfunction ◽

Endothelial Barrier Dysfunction ◽

Microvascular Endothelial ◽

Hepatocyte Growth

Acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) is characterized by pulmonary microvascular endothelial barrier dysfunction. Mesenchymal stem cell-secreted hepatocyte growth factor (HGF) has positive effects of lipopolysaccharide- (LPS-) induced pulmonary endothelial barrier. Studies have exhibited the mammalian TORC1 (mTORC1) signaling is of potent angiogenesis effects. The mTOR protein kinase has two distinct multiprotein complexes mTORC1 and mTORC2 that regulate different branches of the mTOR network. However, detailed mTORC2 mechanisms of HGF protective effects remain poorly defined. Therefore, the aim of this study was to determine whether mTORC2 mediated protective effects of MSC-secreted HGF against LPS-induced pulmonary microvascular endothelial barrier dysfunction activated like mTORC1 activation. We introduced MSC-PMVEC coculture transwell system and recombinant murine HGF on LPS-induced endothelial cell barrier dysfunction in vitro and then explored potential mechanisms by lentivirus vector-mediated HGF, mTORC1 (raptor), and mTORC2 (rictor) gene knockdown modification. Endothelial paracellular and transcellular permeability, adherent junction protein (VE-Cadherin), cell proliferation, apoptosis, and mTOR-associated proteins were tested. These revealed that HGF could promote quick reestablishment of adherent junction VE-cadherin and decrease endothelial paracellular and transcellular permeability during LSP-induced endothelial dysfunction with the involvement of mTORC2 (rictor) and mTORC1 (raptor) pathways. Raptor and rictor knockdown in LPS-induced PMEVECs with stimulation of HGF increased apoptosis ratio, activated Cleaved-Caspase-3 expression, and downregulated cell proliferation. Moreover, mTORC2/Akt but not mTORC2/PKC had significance on HGF endothelial protective effects. Taken together, these highlight activation mTORC2 pathway could also contribute to vascular endothelial barrier recovery by MSC-secreted HGF in LPS stimulation.

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PKA inhibits RhoA activation: a protection mechanism against endothelial barrier dysfunction

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00429.2002 ◽

2003 ◽

Vol 284 (6) ◽

pp. L972-L980 ◽

Cited By ~ 120

Author(s):

Jing Qiao ◽

Fei Huang ◽

Hazel Lum

Keyword(s):

Endothelial Cells ◽

Protein Kinase ◽

Rho Gtpases ◽

Endothelial Barrier ◽

Intracellular Camp ◽

Protective Effects ◽

Barrier Dysfunction ◽

Similar Time ◽

Endothelial Barrier Dysfunction ◽

Rhoa Activation

Much evidence indicates that cAMP-dependent protein kinase (PKA) prevents increased endothelial permeability induced by inflammatory mediators. We investigated the hypothesis that PKA inhibits Rho GTPases, which are regulator proteins believed to mediate endothelial barrier dysfunction. Stimulation of human microvascular endothelial cells (HMEC) with thrombin (10 nM) increased activated RhoA (RhoA-GTP) within 1 min, which remained elevated approximately fourfold over control for 15 min. The activation was accompanied by RhoA translocation to the cell membrane. However, thrombin did not activate Cdc42 or Rac1 within similar time points, indicating selectivity of activation responses by Rho GTPases. Pretreatment of HMEC with 10 μM forskolin plus 1 μM IBMX (FI) to elevate intracellular cAMP levels inhibited both thrombin-induced RhoA activation and translocation responses. FI additionally inhibited thrombin-mediated dissociation of RhoA from guanine nucleotide dissociation inhibitor (GDI) and enhanced in vivo incorporation of32P by GDI. HMEC pretreated in parallel with FI showed >50% reduction in time for the thrombin-mediated resistance drop to return to near baseline and inhibition of ∼23% of the extent of resistance drop. Infection of HMEC with replication-deficient adenovirus containing the protein kinase A inhibitor gene (PKA inhibitor) blocked both the FI-mediated protective effects on RhoA activation and resistance changes. In conclusion, the results provide evidence that PKA inhibited RhoA activation in endothelial cells, supporting a signaling mechanism of protection against vascular endothelial barrier dysfunction.

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