Mechanical ventilation of lungs is capable of activating the innate immune system and inducing sterile inflammatory response. The proinflammatory cytokine IL-1β is among the definitive markers for accurately identifying ventilator-induced lung inflammation. However, mechanisms of IL-1β release during mechanical ventilation are unknown. Here we show that cyclic stretch activates the NLRP3 inflammasomes and induces the release of IL-1β in mouse alveolar macrophages via caspase-1- and TLR4-dependent mechanisms. We also observed that NADPH oxidase subunit gp91phox was dispensable for stretch-induced cytokine production whereas mitochondrial generation of reactive oxygen species was required for stretch-induced NLRP3 inflammasome activation and IL-1β release. Further, mechanical ventilation activated the NLRP3 inflammasomes in mouse alveolar macrophages and increased the production of IL-1β in vivo. IL-1β neutralization significantly reduced mechanical ventilation-induced inflammatory lung injury. These findings suggest that the alveolar macrophage NLRP3 inflammasome may sense lung alveolar stretch to induce the release of IL-1β, and hence may contribute to the mechanism of lung inflammatory injuryduring mechanical ventilation.
Activation of TLR4 by the endotoxin LPS is a critical event in the pathogenesis of Gram-negative sepsis. Caveolin-1, the signaling protein associated with caveolae, is implicated in regulating the lung inflammatory response to LPS; however, the mechanism is not understood. Here we investigated the role of caveolin-1 in regulating TLR4 signaling in endothelial cells. We observed that LPS interaction with CD14 in endothelial cells induced Src-dependent caveolin-1 phosphorylation at Tyr 14. Using TLR4-MD2-CD14 transfected HEK-293 cell line and caveolin-1-deficient (cav-1−/−) mouse lung microvascular endothelial cells, we demonstrated that caveolin-1 phosphorylation at Tyr14 following LPS exposure induced caveolin-1 and TLR4 interaction and thereby TLR4 activation of MyD88, leading to NF-κB activation and generation of proinflammatory cytokines. Exogenous expression of phosphorylation-deficient Y14F caveolin-1 mutant in cav-1−/− mouse pulmonary vasculature rendered the mice resistant to LPS compared to reintroduction of wild type caveolin-1. Thus, caveolin-1 Y14 phosphorylation was required for the interaction with TLR4 and activation of TLR4-MyD88 signaling and sepsis-induced lung inflammation. Inhibiting caveolin-1 Tyr14 phosphorylation and resultant inactivation of TLR4 signaling in pulmonary vascular endothelial cells represents a novel strategy for preventing sepsis-induced lung inflammation and injury.
Objective Vascular endothelial (VE)-cadherin is the predominant component of endothelial adherens junctions essential for cell-cell adhesion and formation of the vascular barrier. Endocytic recycling is an important mechanism for maintaining the expression of cell surface membrane proteins. However, little is known about the molecular mechanism of VE-cadherin recycling and its role in maintenance of vascular integrity. Approach and Results Using calcium-switch assay, confocal imaging, cell surface biotinylation and flow cytometry, we showed that VE-cadherin recycling required Rab11a and Rab11-family interacting protein 2 (FIP2). Yeast 2-hybrid assay and co-immunoprecipitation demonstrated that direct interaction of VE-cadherin with FIP2 (at aa 453-484) formed a ternary complex with Rab11a in human endothelial cells. Silencing of Rab11a or FIP2 in endothelial cells prevented VE-cadherin recycling and VE-cadherin expression at endothelial plasma membrane. Further, inactivation of Rab11a signaling blocked junctional re-annealing following vascular inflammation. Selective knockdown of Rab11a in pulmonary microvessels markedly increased vascular leakage in mice challenged with lipopolysaccharide or polymicrobial sepsis. Conclusions Rab11a/FIP2-mediated VE-cadherin recycling is required for formation of adherens junctions and restoration of vascular endothelial barrier integrity and hence a potential target for clinical intervention in inflammatory disease.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.