Rutin targets AKT to inhibit ferroptosis in ventilator-induced lung injury.

Our previous research confirmed that rutin reduced ventilator-induced lung injury (VILI) in mice. Ferroptosis has been reported to participate in the pathogenic process of VILI. We will explore whether rutin inhibits ferroptosis to alleviate VILI. A mouse model of VILI was constructed with or without rutin pretreatment to perform a multiomics analysis. Hematoxylin-eosin (HE) staining and transmission electron microscopy were used to evaluate lung injury in VILI mice. Dihydroethidium (DHE) staining and the malondialdehyde (MDA) and superoxide dismutase (SOD) levels were detected. Molecular docking was performed to determine the binding affinity between rutin and ferroptosis-related proteins. Western blot analysis, real-time PCR (RT-PCR) and immunohistochemical (IHC) staining were conducted to detect the expression levels of GPX4, XCT, ACSL4, FTH1, AKT and p-AKT in lung tissues. Microscale thermophoresis (MST) was used to evaluate the binding between rutin and AKT1. Transcriptomic and proteomic analyses showed that ferroptosis may play a key role in VILI mice. Metabolomic analysis demonstrated that rutin may affect ferroptosis via the AKT pathway. Molecular docking analysis indicated that rutin may regulate the expression of ferroptosis-related proteins. Moreover, rutin upregulated GPX4 expression and downregulated the expression of XCT, ACSL4 and FTH1 in the lung tissues. Rutin also increased the ratio of p-AKT/AKT and p-AKT expression. MST analysis showed that rutin binds to AKT1. Rutin binds to AKT to activate the AKT signaling pathway, contributing to inhibit ferroptosis, thus preventing VILI in mice. Our study elucidated a possible novel strategy of involving the use of rutin for preventing VILI.

Simultaneous Pretreatment of Aspirin and Omega-3 Fatty Acid Attenuates Nuclear Factor-κB Activation in a Murine Model with Ventilator-Induced Lung Injury.

Ventilator-induced lung injury (VILI) is an important critical care complication. Nuclear factor-κB (NF-κB) activation, a critical signaling event in the inflammatory response, has been implicated in the tracking of the lung injury. The present study aimed to determine the effect of simultaneous pretreatment with enteral aspirin and omega-3 fatty acid on lung injury in a murine VILI model. We compared the lung inflammation after the sequential administration of lipopolysaccharides and mechanical ventilation between the pretreated simultaneous enteral aspirin and omega-3 fatty acid group and the non-pretreatment group, by quantifying NF-κB activation using an in vivo imaging system to detect bioluminescence signals. The pretreated group with enteral aspirin and omega-3 fatty acid exhibited a smaller elevation of bioluminescence signals than the non-pretreated group (p = 0.039). Compared to the non-pretreated group, the pretreatment group with simultaneous enteral aspirin and omega-3 fatty acid showed reduced expression of the pro-inflammatory cytokine, tumor necrosis factor-α, in bronchoalveolar lavage fluid (p = 0.038). Histopathological lung injury scores were also lower in the pretreatment groups compared to the only injury group. Simultaneous pretreatment with enteral administration of aspirin and omega-3 fatty acid could be a prevention method for VILI in patients with impending mechanical ventilation therapy.

Alpha 1-antitrypsin for treating ventilator-associated lung injury in acute respiratory distress syndrome rats.

Purpose: Mechanical ventilation (MV) is an essential life support tool for patients with acute respiratory distress syndrome (ARDS). However, MV for ARDS can result in ventilator-induced lung injury (VILI). This study aimed to assess whether alpha 1-antitrypsin (AAT) can reduce VILI in ARDS rats. Materials and Methods: Rats were randomly divided into five groups: the sham (S) group, MV (V) group, lipopolysaccharide (LPS) (L) group, MV/LPS (VL) group and MV/AAT (VA) group. Rats in the S group were anesthetized. The rats in the L group received LPS but not ventilation, the rats in the V group received only MV, and the rats in the VL and VA groups received LPS and MV. Additionally, the rats in the VA group were treated with AAT, and the other rats were injected with saline. The PaO2/FiO2 ratio and the wet/dry weight were assessed. The total protein and neutrophil elastase concentrations and the neutrophil and macrophage counts in bronchoalveolar lavage fluid (BALF) were evaluated. Proinflammatory factors in BALF and ICAM-1 and MIP-2 in serum were also tested. Furthermore, the oxidative stress response was detected, and histological injury and apoptosis were evaluated. Results: All the rats in the V, L and VL groups had significant lung injury, with the VL group exhibiting the most severe injury. Compared with the findings in the VL group, AAT significantly upregulated the PaO2/FiO2 ratio but decreased the wet/dry weight ratio and protein levels in BALF. AAT also reduced proinflammatory cytokine levels and inflammatory cell counts in BALF. Lung tissue injury and cell apoptosis were mitigated by AAT. Conclusions: AAT ameliorated VILI in ARDS rats. The protection conferred by AAT may be associated with the anti-inflammatory, antioxidative stress response and anti-apoptotic effects of AAT.

Epigallocatechin gallate attenuates mitochondrial DNA-induced inflammatory damage in the development of ventilator-induced lung injury.

OBJECTIVE: We aim to investigate the role of mitochondrial DNA (mtDNA), a novel endogenous pro-inflammatory cytokine, in the development of ventilator-induced lung injury (VILI). Moreover, the protective effect of epigallocatechin gallate (EGCG) on VILI through inhibiting local mtDNA release was examined. METHODS: From March 2015 to March 2016, bronchoalveolar lavage fluid (BALF) from 36 patients with VILI and well-matched 36 patients without VILI after major surgery were consecutively collected. The expression levels of mtDNA and inflammatory cytokines in BALF were tested. SD rats were divided into five groups: control, low tidal volume (7 ml/kg) group, high tidal volume (HTV, 40 ml/kg) group, HTV+low dose EGCG and HTV+high dose EGCG groups. BALF were collected to examine the expression levels of mtDNA and several inflammatory cytokines and the lung tissue was harvested for pathological examinations. In addition, cyclic stretch cell culture was used and culture media was collected to analyze expressions of inflammatory cytokines. Administration of mtDNA in a rat model and in vitro cell culturing were used to confirm its pro-inflammatory properties in the development of inflammatory lung injury. RESULTS: A Significant elevation of mtDNA was detected in BALF from patients with VILI (581 ±â€¯193 vs. 311 ±â€¯137, p < 0.05) and also in rats ventilated with HTV. EGCG could significantly inhibit HTV-induced local mtDNA release and attenuate the level of inflammatory lung injuries (reduced infiltration of local inflammatory cells, lower lung wet/dry ratio and expression levels of inflammatory cytokines). The beneficial effects of EGCG on preventing inflammatory lung injuries were in a concentration-dependent manner. Meanwhile, higher expression levels of mtDNA and inflammatory cytokines were observed in the media of cyclic stretched cell culture compared to those in the control group (p < 0.05). Furthermore, intra-tracheal administration of mtDNA in rats could lead to a marked increase of local inflammatory cytokines and subsequent inflammatory lung injuries (p < 0.05). And by adding mtDNA into the cell culture, higher level of inflammatory cytokines in the media was detected (p < 0.05). EGCG also showed preventive effects on inflammatory responses on a concentration-dependent manner (p < 0.05). CONCLUSION: The increased expression level of mtDNA and subsequent inflammatory cytokines overproduction may play an important role in the development of VILI. EGCG may be a potential novel therapeutic candidate for protection against VILI by inhibiting the local release of mtDNA.

Curcumin ameliorated ventilator-induced lung injury in rats.

BACKGROUND: Curcumin (CUR) is a Chinese medicine monomer with antioxidant and anti-inflammatory properties. The aim of this study was to investigate the effects and mechanisms of CUR treatment on ventilator-induced lung injury (VILI) in rats. METHODS: Total 50 SD rats were divided into five groups: sham, VILI, VILI+CUR-50 (CUR 50?mg/kg pretreated intraperitoneal), VILI+CUR-200 (CUR 200?mg/kg pretreated intraperitoneal) and VILI?+?DXM (5?mg/kg pretreated intraperitoneal). The morphology and ultrastructure were observed by microscope and transmission electron microscope. The wet to dry ratio, protein concentration in bronchoalveolar lavage fluid (BALF), evans blue dye (EBD) content, nuclear factor kappa B (NF-?B) activity, myeloperoxidase (MPO), malondialdehyde (MDA), xanthine oxidase (XO) and total antioxidative capacity (TAOC) levels were measured. RESULTS: Histological studies revealed that inflammatory cells infiltration and alveolar edema were significantly severe in VILI as compared to other groups. CUR-200 and DXM treatment reversed lung injury significantly. The wet to dry ratio, protein concentration in BALF, EBD content, MPO activity, tumor necrosis factor (TNF)-? level and NF-?B activity were significantly increased in VILI group as compared to other groups. CUR-200 and DXM treatment significantly suppressed permeability and inflammation induced by ventilation. Furthermore, the significantly higher MDA content in VILI could be markedly decreased by CUR-200 and DXM treatment while the levels of XO and TAOC were markedly recovered only by CUR (200?mg/kg) treatment after VILI. CONCLUSION: CUR could inhibit the inflammatory response and oxidative stress during VILI, which is partly through NF-?B pathway.

Differential and opposing effects of imatinib on LPS- and ventilator-induced lung injury.

Endothelial dysfunction underlies the pathophysiology of vascular disorders such as acute lung injury (ALI) syndromes. Recent work has identified the Abl family kinases (c-Abl and Arg) as important regulators of endothelial cell (EC) barrier function and suggests that their inhibition by currently available pharmaceutical agents such as imatinib may be EC protective. Here we describe novel and differential effects of imatinib in regulating lung pathophysiology in two clinically relevant experimental models of ALI. Imatinib attenuates endotoxin (LPS)-induced vascular leak and lung inflammation in mice but exacerbates these features in a mouse model of ventilator-induced lung injury (VILI). We next explored these discrepant observations in vitro through investigation of the roles for Abl kinases in cultured lung EC. Imatinib attenuates LPS-induced lung EC permeability, restores VE-cadherin junctions, and reduces inflammation by suppressing VCAM-1 expression and inflammatory cytokine (IL-8 and IL-6) secretion. Conversely, in EC exposed to pathological 18% cyclic stretch (CS) (in vitro model of VILI), imatinib decreases VE-cadherin expression, disrupts cell-cell junctions, and increases IL-8 levels. Downregulation of c-Abl expression with siRNA attenuates LPS-induced VCAM-1 expression, whereas specific reduction of Arg reduces VE-cadherin expression in 18% CS-challenged ECs to mimic the imatinib effects. In summary, imatinib exhibits pulmonary barrier-protective and anti-inflammatory effects in LPS-injured mice and lung EC; however, imatinib exacerbates VILI as well as dysfunction in 18% CS-EC. These findings identify the Abl family kinases as important modulators of EC function and potential therapeutic targets in lung injury syndromes.

Selective inhibition of intra-alveolar p55 TNF receptor attenuates ventilator-induced lung injury.

BACKGROUND: Tumour necrosis factor (TNF) is upregulated in the alveolar space early in the course of ventilator-induced lung injury (VILI). Studies in genetically modified mice indicate that the two TNF receptors play opposing roles during injurious high-stretch mechanical ventilation, with p55 promoting but p75 preventing pulmonary oedema. AIM: To investigate the effects of selective inhibition of intra-alveolar p55 TNF receptor on pulmonary oedema and inflammation during ventilator-induced lung injury using a newly developed domain antibody. METHODS: Anaesthetised mice were ventilated with high tidal volume and given an intratracheal bolus of p55-specific domain antibody or anti-TNF monoclonal antibody ('pure' VILI model). As a model of enhanced inflammation, a subclinical dose of lipopolysaccharide (LPS) was included in the intratracheal antibody bolus (LPS+VILI model). Development of lung injury was assessed by respiratory mechanics and blood gases and protein levels in lavage fluid. Flow cytometry was used to determine leucocyte recruitment and alveolar macrophage activation, while lavage fluid cytokines were assessed by ELISA. RESULTS: The ventilation protocol produced deteriorations in respiratory mechanics and gas exchange with increased lavage fluid protein levels in the two models. The p55-specific domain antibody substantially attenuated all of these changes in the 'pure' VILI model, while anti-TNF antibody was ineffective. In the LPS+VILI model, p55 blockade prevented deteriorations in respiratory mechanics and oxygenation and significantly decreased neutrophil recruitment, expression of intercellular adhesion molecule 1 on alveolar macrophages, and interleukin 6 and monocyte chemotactic protein 1 levels in lavage fluid. CONCLUSIONS: Selective inhibition of intra-alveolar p55 TNF receptor signalling by domain antibodies may open new therapeutic approaches for ventilated patients with acute lung injury.