Mediators In Bronchoalveolar Lavage Fluid Research Articles

Protective effects of an aqueous extract of Protaetia brevitarsis seulensis larvae in an ovalbumin-induced murine model of asthma

ABSTRACT Asthma is a disease-related allergic response characterized by the accumulation of inflammatory cells in airway tissue. Protaetia brevitarsis seulensis larvae are known for their nutritional value and have been used in food; however, it is unknown whether they exert protective effects in a murine ovalbumin (OVA)-induced asthmatic model. We investigated the protective effects of an aqueous extract of P. brevitarsis seulensis larvae (PBE) against OVA-induced asthma by performing histopathological analysis and measuring inflammatory mediators in bronchoalveolar lavage fluid using enzyme-linked immunosorbent assay and flow cytometry. Asthmatic symptoms were attenuated by PBE, as indicated by decreased inflammation in lung tissue and decreased levels of inflammatory cells, inflammatory cytokines, and Th2 cell activation. Additionally, four out of six major metabolites isolated from P. brevitarsis seulensis attenuated Th2 cell activation, indicating that PBE attenuates asthmatic inflammation by regulating Th2 cell activation. Therefore, our findings support PBE as a pharmacological candidate for asthma treatment.

Bronchial epithelial cell transcriptome shows endotype heterogeneity of asthma in patients with NSAID-exacerbated respiratory disease

Nonsteroidal anti-inflammatory drugs-exacerbated respiratory disease (N-ERD) is currently classified as a type-2 (T2) immune-mediated disease characterized by asthma, chronic rhinosinusitis, and hypersensitivity to cyclooxygenase-1 inhibitors. The aim of this study was to characterize immunological endotypes of N-ERD based on the gene expression profile in the bronchial epithelium. mRNA transcriptome (mRNA-sequencing) was analyzed in bronchial brushings from patients with N-ERD (n=22), those with nonsteroidal anti-inflammatory drug-tolerant asthma (NTA, n= 21), and control subjects (n=11). Additionally, lipid and protein mediators were measured in bronchoalveolar lavage fluid (BALF). Initial analysis of the entire asthma group revealed 2 distinct gene expression signatures: "T2-high" with increased expression of T2-related genes (eg, CLCA1, CST1), and "proinflammatory" characterized by the expression of innate immunity (eg, FOSB, EGR3) and IL-17A response genes. These endotypes showed similar prevalence in N-ERD and NTA (eg, T2-high: 33% and 32%, respectively). T2-high asthma was characterized by increased expression of mast cell and eosinophil markers, goblet cell hyperplasia, and elevated LTE4 and PGD2 in BALF. Patients with a proinflammatory endotype showed mainly neutrophilic inflammation and increased innate immunity mediators in BALF. Furthermore, the proinflammatory signature was associated with a more severe course of asthma and marked airway obstruction. These signatures could be recreated invitro by exposure of bronchial epithelial cells to IL-13 (T2-high) and IL-17A (proinflammatory). T2-high signature was found only in one-third of patients with N-ERD, which was similar to what was found in patients with NTA. The proinflammatory endotype, which also occurred in N-ERD, suggests a novel mechanism of severe disease developing on a non-T2 background.

A High-Fiber Diet or Dietary Supplementation of Acetate Attenuate Hyperoxia-Induced Acute Lung Injury.

A high fiber diet (HFD) and dietary supplementation with acetate have been reported to have beneficial effects in a variety of diseases. We investigated the effects of a HFD and acetate supplementation on the gut microbiota and hyperoxia-induced acute lung injury (HALI) in mice. Mice were fed a control diet, HFD, or acetate supplementation for three weeks, and their gut microbiome composition, lung tissues, and bronchoalveolar lavage fluid (BALF) were examined after exposure to ambient air or hyperoxia. Both the HFD and acetate supplementation modified the gut microbiota community and increased the proportion of acetate-producing bacteria in mice exposed to hyperoxia. The HFD and acetate supplementation also increased the abundance of Bacteroides acidifaciens and reduced gut dysbiosis according to the ratio of Firmicutes to Bacteroidetes. Compared with hyperoxia-exposed mice fed a control diet, both the HFD and acetate supplementation significantly increased the survival time while reducing the severity of pulmonary edema and the concentrations of protein and inflammatory mediators in BALF. Moreover, the HFD and acetate supplementation reduced the production of free radicals, attenuated NF-κB signaling activation, and decreased apoptosis in the lung tissues. Overall, this study indicates that a HFD or acetate supplementation reduces the severity of HALI through alterations in the gut microbiota to exert anti-inflammatory effects.

Pro‐Inflammatory Profiling Of RAGE Targeted Mice Following Short‐Term Tobacco Exposure

Inflammation underpins pulmonary disease progression during tobacco smoke exposure that may culminate in irreversdible pulmonary remodeling. While primary smoke poses notable risk, nearly half of the US population is also at risk due to exposure to secondhand smoke (SHS). In the present study, we assessed a potential role for RAGE, a cell‐surface pattern recognition receptor implicated in pro‐inflammatory signaling, following exposure to SHS. Specifically, we used wild type, RAGE null, and lung‐specific RAGE overexpressing transgenic (TG) mice and evaluated the elaboration of inflammatory mediators in bronchoalveolar lavage fluid (BALF). Select mice were administered semi‐synthetic glycosaminoglycan ethers (SAGEs), a family of anionic, partially lipophilic sulfated polysaccharide derivatives known to inhibit RAGE signaling. Mice were exposed to room air (RA) or SHS from three Kentucky 3R4F research cigarettes via a nose‐only delivery system (Sireq Scientific, Montreal, Canada) five days a week and ip injections of PBS or SAGE (a 30mg/kg body weight) occurred three times per week from PN40 until sacrifice date on PN70. RAGE mRNA and protein expression was elevated in wild type and TG mice following SHS exposure and no expression was detected in RAGE nulls. BALF analyses revealed RAGE‐mediated control of leukocyte extravasation and a multiplex cytokine array confirmed a role for RAGE in the coordination of pro‐inflammatory chemokine/cytokine secretion. Among other mediators, TNF‐a, MIP‐2, and IL‐1b were each differentially secreted by lung tissue following SHS exposure and concentrations were significantly decreased in BALF from exposed RAGE null or wild type mice concomitantly administered SAGEs. In summary, inflammatory responses induced by SHS exposure were influenced by the availability of RAGE, as evidenced by RAGE nulls and SAGE treatment. These data reveal fascinating data suggesting the utility of RAGE abrogation in lessening smoke‐induced pulmonary exacerbations.

The Activation of AMPK/NRF2 Pathway in Lung Epithelial Cells Is Involved in the Protective Effects of Kinsenoside on Lipopolysaccharide-Induced Acute Lung Injury.

The disorder of mitochondrial dynamic equilibrium of lung epithelial cell is one of the critical causes of acute lung injury (ALI). Kinsenoside (Kin) serves as an active small-molecule component derived from traditional medicinal herb displaying multiple pharmacological actions in cancers, hyperglycemia, and liver disease. The objective of this study was to investigate the effects of Kin on lipopolysaccharide- (LPS-) induced ALI and further explore possible molecular mechanisms. Kin was administered orally (100 mg/kg/day) for 7 consecutive days before LPS instillation (5 mg/kg). After 12 hours, pathological injury, inflammatory response, and oxidative stress were detected. The results demonstrated that Kin significantly alleviated lung pathological injury and decreased the infiltration of inflammatory cells and the release of inflammatory mediators in bronchoalveolar lavage fluid (BALF), apart from inhibiting the production of reactive oxygen species (ROS) and lipid peroxidation. Meanwhile, Kin also promoted mitochondrial fusion and restrained mitochondrial fission in mice with ALI. In terms of mechanism, Kin pretreatment increased the phosphorylation of AMP-activated protein kinase (AMPK) and the protein level of nuclear factor erythroid 2-related factor 2 (NRF2). In Ampk-α knockout mice challenged with LPS, Kin lost its pulmonary protective effects, accompanied by lower NRF2 level. In vitro experiments further unveiled that either AMPK inhibition by Compound C or NRF2 knockdown by siRNA abolished the protective roles of Kin in LPS-treated A549 lung epithelial cells. And NRF2 activator TAT-14 could reverse the effects of Ampk-α deficiency. In conclusion, Kin possesses the ability to prevent LPS-induced ALI by modulating mitochondrial dynamic equilibrium in lung epithelial cell in an AMPK/NRF2-dependent manner.

Abietic acid attenuates sepsis-induced lung injury by inhibiting nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway to inhibit M1 macrophage polarization

Lung injury is one of the leading causes of death in sepsis. Abietic acid (AA) has demonstrated anti-inflammatory and bacteriostatic properties. Herein, we established a mouse model of sepsis by cecal ligation and puncture, and intraperitoneally injected AA to treat. Lung injury was assessed by H&E staining and the inflammation in bronchoalveolar lavage fluid (BALF) were assessed by counting the number of inflammatory cells and detecting the content of inflammatory factors. Meanwhile, we also designed to study the effect of AA on lipopolysaccharide (LPS)-induced inflammatory response and macrophage marker gene expression in RAW264.7 cells in vitro. In this study, we found that AA inhibited LPS-induced secretion of inflammatory mediators (IL-1β, TNF-α, IL-6 and MIP-2), and decreased the expression of M1 macrophage e markers (CD16 and iNOS) and p-p65 protein, while increased the expression of M2 macrophage markers (CD206 and Arg-1) in RAW264.7 cells in vitro. In vivo, the therapy of AA not only rescued septic animals, but also attenuated lung injury in sepsis mice. Moreover, AA decreased the number of total cells, neutrophils and macrophages, the conceration of total protein, and the levels of inflammatory mediators in BALF of sepsis mice. Further, we found that AA inhibited M1 macrophage polarization and blocked nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway in BALF of sepsis mice. In conclusion, Abietic acid attenuates sepsis-induced lung injury, and its mechanism may be related to reducing inflammation by inhibiting NF-κB signaling to inhibit M1 macrophage polarization.

Autoantibody of interleukin-17A induced by recombinant Mycobacterium smegmatis attenuates airway inflammation in mice with neutrophilic asthma

Objective Previous studies have shown Interleukin (IL)-17A as an important contributor to the development of severe asthma, which is mainly characterized by neutrophilic inflammation and less response to corticosteroids. Consequently, the IL-17A-neutrophil axis could be a potential therapeutic target. Previously, we constructed a recombinant Mycobacterium smegmatis (rMS) expressing fusion protein Ag85A-IL-17A, and confirmed it could induce production of IL-17A autoantibody in vivo. This study uses a murine model of neutrophilic asthma to further investigate the effects of rMS on airway inflammation. Methods DO11.10 mice were divided into four groups: phosphate buffered saline (PBS), asthma, rMS and MS. This murine model of neutrophilic asthma was established with ovalbumin (OVA) challenge, whereby PBS, rMS and MS were administered intranasally. Anti-inflammatory effects on inflammatory cell infiltration and expression of inflammatory mediators in bronchoalveolar lavage fluid (BALF) were evaluated, along with histopathological changes in lung tissues. Results A sustained high-titer IL-17A autoantibody was detected in sera of the rMS group. Compared to the asthma group, the number of neutrophils, IL-17A, CXCL-1 levels and MPO activity in the rMS group were all significantly reduced (p < 0.01). Histological analysis showed rMS remarkably suppressed inflammatory infiltration around bronchia. The inflammation score and the mucus score in the rMS group were both significantly lower than those in the asthma group (p < 0.001). Conclusion rMS ameliorated airway inflammation in mice with neutrophilic asthma caused by inducing IL-17A autoantibody and regulating the IL-17A-neutrophil axis, thus offering a possible novel treatment for neutrophilic asthma.

Biotrauma during ultra-low tidal volume ventilation and venoarterial extracorporeal membrane oxygenation in cardiogenic shock: a randomized crossover clinical trial

BackgroundCardiogenic pulmonary oedema (CPE) may contribute to ventilator-associated lung injury (VALI) in patients with cardiogenic shock. The appropriate ventilatory strategy remains unclear. We aimed to evaluate the impact of ultra-low tidal volume ventilation with tidal volume of 3 ml/kg predicted body weight (PBW) in patients with CPE and veno–arterial extracorporeal membrane oxygenation (V–A ECMO) on lung inflammation compared to conventional ventilation.MethodsA single-centre randomized crossover trial was performed in the Cardiac Intensive Care Unit (ICU) at a tertiary university hospital. Seventeen adults requiring V–A ECMO and mechanical ventilation due to cardiogenic shock were included from February 2017 to December 2018. Patients were ventilated for two consecutive periods of 24 h with tidal volumes of 6 and 3 ml/kg of PBW, respectively, applied in random order. Primary outcome was the change in proinflammatory mediators in bronchoalveolar lavage fluid (BALF) between both ventilatory strategies.ResultsVentilation with 3 ml/kg PBW yielded lower driving pressures and end-expiratory lung volumes. Overall, there were no differences in BALF cytokines. Post hoc analyses revealed that patients with high baseline levels of IL-6 showed statistically significant lower levels of IL-6 and IL-8 during ultra-low tidal volume ventilation. This reduction was significantly proportional to the decrease in driving pressure. In contrast, those with lower IL-6 baseline levels showed a significant increase in these biomarkers.ConclusionsUltra-low tidal volume ventilation in patients with CPE and V–A ECMO may attenuate inflammation in selected cases. VALI may be driven by an interaction between the individual proinflammatory profile and the mechanical load overimposed by the ventilator.Trial registration The trial was registered in ClinicalTrials.gov (identifier NCT03041428, Registration date: 2nd February 2017).

Chemokines and eicosanoids fuel the hyperinflammation within the lungs of patients with severe COVID-19

BackgroundSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can lead to a variety of clinical outcomes, ranging from the absence of symptoms to severe acute respiratory disease and ultimately death. A feature of patients with severe coronavirus disease 2019 (COVID-19) is the abundance of inflammatory cytokines in the blood. Elevated levels of cytokines are predictive of infection severity and clinical outcome. In contrast, studies aimed at defining the driving forces behind the inflammation in lungs of subjects with severe COVID-19 remain scarce.ObjectiveOur aim was to analyze and compare the plasma and bronchoalveolar lavage (BAL) fluids of patients with severe COVID-19 (n = 45) for the presence of cytokines and lipid mediators of inflammation (LMIs).MethodsCytokines were measured by using Luminex multiplex assay, and LMIs were measured by using liquid chromatography–tandem mass spectrometry.ResultsWe revealed high concentrations of numerous cytokines, chemokines, and LMIs in the BAL fluid of patients with severe COVID-19. Of the 13 most abundant mediators in BAL fluid, 11 were chemokines, with CXCL1 and CXCL8 being 200 times more abundant than IL-6 and TNF-α. Eicosanoid levels were also elevated in the lungs of subjects with severe COVID-19. Consistent with the presence chemotactic molecules, BAL fluid samples were enriched for neutrophils, lymphocytes, and eosinophils. Inflammatory cytokines and LMIs in plasma showed limited correlations with those present in BAL fluid, arguing that circulating inflammatory molecules may not be a reliable proxy of the inflammation occurring in the lungs of patients with severe COVID-19.ConclusionsOur findings indicate that hyperinflammation of the lungs of patients with severe COVID-19 is fueled by excessive production of chemokines and eicosanoids. Therapeutic strategies to dampen inflammation in patients with COVID-19 should be tailored accordingly.

Lactic Acid Bacteria and Natural Product Complex Ameliorates Ovalbumin-Induced Airway Hyperresponsiveness in Mice.

The incidence of respiratory diseases, such as asthma, has substantially increased in recent times owing to environmental changes, such as air pollution. Induction of a chronic inflammatory response begins with production of biologically active mediators from the airway epithelium, which attracts and recruits inflammatory cells into the lung airway. In our previous study, we confirmed that Lactobacillus casei HY2782 and Bifidobacterium animalis spp. lactis HY8002 could improve lung inflammation in the COPD animal model. In this study, we investigated the effect of the HY2782 complex against airway hyperresponsiveness by using an ovalbumin (OVA)-induced animal model. An orally administered HY2782 complex on OVA-induced allergic asthma in a BALB/c mouse model was used. The present results showed that the HY2782 complex suppressed total immunoglobulin E in serum and bronchoalveolar lavage fluid (BALF). The cytokine production profile in BALF and serum revealed that the HY2782 complex showed reduced levels of Th2 cytokines among immune factors released due to the elevated allergic response. Levels of inflammatory mediators in BALF, MCP-1, MIP-2, and CXCL-9 were decreased by oral administration of the HY2782 complex. Lower numbers of eosinophils and neutrophils in BALF suggested that inflammation was ameliorated by the HY2782 complex. Histological observation of lung sections also showed infiltration of fewer cells. From results, we suggested that the HY2782 complex effectively responds to improvement of the immune response and airway hypersensitivity reaction because of the anti-inflammatory effect of the Pueraria lobata root extract and antioxidant effect of HY2782.

Melatonin Attenuates Sepsis-Induced Acute Lung Injury Through Improvement of Epithelial Sodium Channel-Mediated Alveolar Fluid Clearance Via Activation of SIRT1/SGK1/Nedd4-2 Signaling Pathway.

Acute lung injury is characterized by alveolar vascular barrier injury, and protein-rich pulmonary oedema. Alveolar fluid clearance is closely related to the prognosis of patients with acute lung injury. Melatonin has been shown to have a protective effect on multiple organ injury induced by sepsis. In this study we investigated the effect of melatonin on alveolar fluid clearance (AFC) and explored its potential mechanisms in sepsis-induced acute lung injury. The cecal ligation and puncture was adopted to establish mouse sepsis model. Morphological changes of lung tissues with the hematoxylin staining were observed. AFC and lung wet/dry weight ratio were measured to assess pulmonary edema. Inflammatory mediators in bronchoalveolar lavage fluid were analyzed via enzyme-linked immunosorbent assay. NAD+/NADH and SIRT1 activity were measured by colorimetric assay kit. The protein expressions of epithelial sodium channel (ENaC), silent information regulator1 (SIRT1), SGK1 and Nedd4-2 were immunoblotted by western blot in vivo and in vitro. The distribution of α-ENaC and SIRT1 was detected by immunofluorescence. We found that melatonin attenuated sepsis induced lung injury, improved survival rate, enhanced alveolar fluid clearance, improved SIRT1 activity, increased protein expressions of SIRT1 and ENaC, and activated SGK1/Nedd4-2 pathway. Furthermore, SIRT1 inhibitor EX527 counteracted the effects of melatonin on alveolar fluid clearance and ENaC. These results revealed that melatonin enhanced ENaC-mediated AFC via the SIRT1/SGK1/Nedd4-2 signaling pathway. Our study demonstrated that melatonin might provide a novel therapeutic strategy for sepsis-induced acute lung injury.

Alkaline phosphatase in pulmonary inflammation\u2014a translational study in ventilated critically ill patients and rats

BackgroundAlkaline phosphatase (AP), a dephosphorylating enzyme, is involved in various physiological processes and has been shown to have anti-inflammatory effects.AimTo determine the correlation between pulmonary AP activity and markers of inflammation in invasively ventilated critically ill patients with or without acute respiratory distress syndrome (ARDS), and to investigate the effect of administration of recombinant AP on pulmonary inflammation in a well-established lung injury model in ratsMethodsAP activity was determined and compared with levels of various inflammatory mediators in bronchoalveolar lavage fluid (BALF) samples obtained from critically ill patients within 2 days of start of invasive ventilation. The endpoints of this part of the study were the correlations between AP activity and markers of inflammation, i.e., interleukin (IL)-6 levels in BALF. In RccHan Wistar rats, lung injury was induced by intravenous administration of 10 mg/kg lipopolysaccharide, followed by ventilation with a high tidal volume for 4 h. Rats received either an intravenous bolus of 1500 IU/kg recombinant AP or normal saline 2 h after intravenous LPS administration, right before start of ventilation. Endpoints of this part of the study were pulmonary levels of markers of inflammation, including IL-6, and markers of endothelial and epithelial dysfunction.ResultsBALF was collected from 83 patients; 10 patients had mild ARDS, and 15 had moderate to severe ARDS. AP activity correlated well with levels of IL-6 (r = 0.70), as well as with levels of other inflammatory mediators. Pulmonary AP activity between patients with and without ARDS was comparable (0.33 [0.14–1.20] vs 0.55 [0.21–1.42] U/L; p = 0.37). Animals with acute lung injury had markedly elevated pulmonary AP activity compared to healthy controls (2.58 [2.18–3.59] vs 1.01 [0.80–1.46] U/L; p < 0.01). Intravenous administration of recombinant AP did neither affect pulmonary inflammation nor endothelial and epithelial dysfunction.ConclusionsIn ventilated critically ill patients, pulmonary AP activity correlates well with markers of pulmonary inflammation, such as IL-6 and IL-8. In animals with lung injury, pulmonary AP activity is elevated. Administration of recombinant AP does not alter pulmonary inflammation and endothelial or epithelial dysfunction in the acute phase of a murine lung injury model.

The protective effect of beclomethasone dipropionate in the modeling of chronic obstructive pulmonary disease

The purpose of the study was to evaluate the protective effect of the inhaled glucocorticosteroids of beclomethasone dipropionate (BDP) using the experimental model of chronic obstructive pulmonary disease (COPD). Materials and Methods . The COPD model was reproduced in rats ( n = 30) during a discrete 60-day exposure to nitrogen dioxide, 9 individuals formed an intact group. Inhalation of BDP (50 micrograms per day) and control ( n = 15) placebo inhalation were performed for 60 days in the rats in the experimental group ( n = 15). The cell composition and cytokine enzymatic profile of bronchoalveolar lavage fluid (BALF), its content of secretory immunoglobulin A (sIgA) and surfactant protein D (SP-D) were determined by ELISA. Histological and morphometric lungs examination was performed. Results . Typical for COPD pathomorphological changes in the lungs were determined in the placebo group, the content of inflammatory cells and pro-inflammatory mediators in BALF increased, the functional activity of bronchoalveolar epithelium (SP-D, sIgA) deteriorated. There was a clear trend in the activity of neutrophilic-lymphocytic inflammation decrease with normalization of the profile of pro-inflammatory cytokines and enzymes in BALF In the group of animals receiving BDP; there was also a restoration of the morphological structure of the bronchoalveolar epithelium and improvement of its functional activity. Conclusion . The using of BDP in rats with the experimental COPD model had an expressed anti-inflammatory effect associated with the activation of physiological repair processes in the lungs, manifested by the restoration of structural, immune barrier integrity and functional activity of the bronchoalveolar epithelium.

Early protein expression profile in bronchoalveolar lavage fluid and clinical outcomes in primary graft dysfunction after lung transplantation.

Primary graft dysfunction (PGD) remains a major post-transplant complication and is associated with increased morbidity and mortality. Mechanisms evoking PGD are not completely clear, but inflammation plays a central role. We investigated the association between PGD and inflammatory proteins present in immediate postoperative bronchoalveolar lavage. All double-lung recipients transplanted at our institution from 2002 to 2018 were included in our study. We retrospectively selected 80 consecutive lung transplant recipients with different PGD grades (n = 20 for each PGD grades 0-1 to 2-3). In bronchoalveolar lavage performed within the first 24 h after donor aortic cross-clamping following lung transplantation, concentrations of 30 cytokines, chemokines and growth factors were assessed by enzyme-linked immunosorbent assay (ELISA) and correlated with donor and recipient demographics and outcomes. For analysis, 2 groups were defined: 'mild' PGD (grade 0-1) and 'severe' PGD (grades 2-3). Significant differences between mild and severe PGD were found in 8 biomarkers [interleukin (IL)-6, IL-10, IL-13, eotaxin, granulocyte colony-stimulating factor, interferon γ, macrophage inflammatory protein 1α, surfactant protein D (SP-D); P < 0.05]. Increased IL-10 and IL-13, but none of the other proteins, were associated with short-term outcome (longer time to extubation; P = 0.005 and P < 0.0001; increased intensive care unit stay; P = 0.012 and P < 0.0001; and hospital stay; P = 0.041 and P = 0.002). There were no significant differences in donor and recipient characteristics between the groups. Expression profiles of key inflammatory mediators in bronchoalveolar lavage fluid differed significantly between lung transplant recipients with severe versus mild PGD and correlated with clinical outcome variables. Further research should focus on the early mechanisms leading to PGD.

Propofol reduces acute lung injury by up-regulating gamma-aminobutyric acid type a receptors

BackgroundWe used a two-hit lung injury rat model that involves mechanical ventilation (MV) following lipopolysaccharide exposure to investigate the effects of propofol on the expression of GABAA receptors (GABAAR) and cytokine responses, and we then determined the specific effects of GABA on cytokine responses in vitro in alveolar epithelial cells (AECs). MethodsForty-eight adult male Wister rats were equally and randomly divided into the following 4 groups (n = 12) using a random number table: sham group, sham+propofol group, lipopolysaccharide (LPS) + VILI group, and LPS + VILI + propofol group. All animals were anesthetized, and the animals received a 3.75 mg/kg intratracheal instillation of endotoxins or phosphate-buffered saline (PBS) as the control, as described previously. After 30 min, rats were ventilated for 5 h in a volume-controlled ventilation mode. In the LPS + VILI group, animals were ventilated with a tidal volume (Vt) of 22 ml/kg and zero positive end-expiratory pressure (PEEP) at a respiratory rate of 16–18 breaths/min, whereas control (sham) rats were ventilated with a Vt of 6 ml/kg and PEEP of 5 cmH2O at a rate of 45–55 breaths/min. The FiO2 remained constant as 0.4, propofol was administered intravenously in the LPS + VILI + propofol and sham + propofol groups at a rate of 10 mg·kg−1·h−1 while normal saline at the same rate was intravenously administered in the LPS + VILI and sham groups during the entire mechanical ventilation period. Five hours after mechanical ventilation, the rats were killed. Survival rates, histopathology, concentrations of inflammatory mediators in bronchoalveolar lavage fluid (BALF), wet weight/dry weight (W/D) ratio of the lung, myeloperoxidase (MPO) activity in lung tissues, and expression of GAD and GABAAR by immunohistochemical detection and Western blotting were assessed. Then, human type II-like alveolar epithelial cells (A549 cells) were cultured to full confluence and incubated with GABA (100 nM) alone, picrotoxin alone, a GABAAR antagonist (PTX, 50 nM), or GABA + PTX for 10 min, followed by stimulation with LPS (control) at 100 ng/ml for 4 h. The concentrations of IL-1β, IL-2, IL-8, and IL-10 were then measured. ResultsAdministration of propofol in a two-hit lung injury rat model can increase survival rates and the expression of GAD and GABAAR (P < .05). The administration of propofol can attenuate the release of pro-inflammatory cytokines both in vivo and in vitro, and the administration of propofol can attenuate histopathological changes, the W/D ratio, and MPO activity (P < .05). ConclusionsIn this study, we found that the administration of propofol improved lung function, alleviated lung injury, and up-regulated the GAD and GABAAR expressions in a two-hit model of acute lung injury (ALI) characterized by intratracheal instillation of an endotoxin and prolonged MV. Therefore, the protective effects of propofol may be associated with the up-regulation of GABAA receptors in AECs.

PM2.5-induced lung inflammation in mice: Differences of inflammatory response in macrophages and type II alveolar cells.

Particulate matter 2.5 (<PM2.5 μm) leads to chronic obstructive pulmonary disease. In this study, biomarkers related to inflammation and oxidative stress in vitro and in vivo experiments were investigated to clarify the PM2.5-induced lung inflammation mechanisms. In an in vitro study using RAW264.7 cells, PM2.5 caused phosphorylation of nuclear factor-κB, p38 mitogen-activated protein kinase and extracellular response kinases, an increase of proinflammatory gene and protein expressions (e.g. monocyte chemotactic protein-1, tumor necrosis factor-α). These biomarkers were substantially attenuated by polymyxin B (PMB). PM2.5 induced heme oxygenase-1 (HO-1) gene, which was attenuated by N-acetylcysteine (NAC). However, the suppressive effects of NAC on inflammatory biomarkers were very weak. In bone marrow-derived macrophages (BMDMs) of wild-type BALB/c mice, the effects of PMB and NAC on PM2.5-induced inflammatory responses were similar to RAW264.7 cells. In BMDMs of MyD88-/- mice, PM2.5-induced proinflammatory mediators were substantially more attenuated. PM2.5 caused an increase of proinflammatory gene expressions (interleukin-6, cyclooxygenase 2) and HO-1 gene in MLE-12 cells (mouse alveolar cell line). These biomarkers were substantially attenuated by NAC, but not by PMB. When BALB/c mice were exposed intratracheally to 0.2 mg PM2.5, PM2.5 caused severe lung inflammation, an increase of neutrophils along with proinflammatory mediators in bronchoalveolar lavage fluid. The inflammation was attenuated by NAC, particularly by NAC + PMB, but not by PMB alone. These results indicate that macrophages may act sensitively to lipopolysaccharide (LPS) present in PM2.5 and release proinflammatory mediators via the LPS/MyD88 pathway. However, type II alveolar cells may react sensitively to oxidative stress induced by PM2.5 and cause inflammatory response. Therefore, overall, PM2.5 may cause predominantly oxidative stress-dependent inflammation rather than LPS/MyD88-dependent inflammation in type II alveolar cell-rich lungs. Copyright © 2017 John Wiley & Sons, Ltd.

Emodin Combined with Nanosilver Inhibited Sepsis by Anti-inflammatory Protection.

Background: Emodin is the main active component of rhubarb, which has demonstrated many beneficial effects against inflammation. Nanosilver is an effective antimicrobial agent. The present study was designed to observe the effects of Emodin combined with silver nanoparticles (E/S) on sepsis protection and related mechanism.Methods: E/S was prepared by loading different concentrations of Emodin on nanosilver and cytotoxicity of E/S were determined by suphorhodamine B assays. Anti-microbial activities of E/S were assayed by direct interaction with various common pathogens and anti-adhesive activites of E/S on leukocytes with endothelial cells were assayed by biochemical analysis. Next, inflammatory cell enumeration, inflammatory mediators in bronchoalveolar lavage fluid (BALF) and endothelial cell function were analyzed on a clinically relevant model of sepsis induced by cecal ligation and puncture (CLP) after E/S administration. The effects of E/S on NF-κB and p38 were also examined by western blot.Results: E/S exhibited little cytotoxicity action on endothelial cells and significant inhibitory activities against all tested common microorganisms and adherence between leukocyte and endothelial cells. E/S induced anti-sepsis protection mainly mediated by inhibition of inflammatory cells infiltration, down-regulation of TNF-alpha, IL-8 and lactic dehydrogenase (LDH), and inhibition of NF-κB and p38 pathways in mice 24 h post-CLP.Conclusion: Our data suggest that E/S has strong anti-sepsis effects, which was related with anti-inflammatory protection and thereby promote survival following sepsis challenge.

Intratracheal Instillation of Perfluorohexane Modulates the Pulmonary Immune Microenvironment by Attenuating Early Inflammatory Factors in Patients With Smoke Inhalation Injury: A Randomized Controlled Clinical Trial.

Smoke inhalation injury (SII) is associated with significant morbidity and mortality in burn patients, and effective treatments are lacking. Perfluorocarbons (PFCs) have a protective effect against acute lung injury. We aimed to assess the therapeutic effects of perfluorohexane on burn patients with SII. Patients with burns complicated by moderately severe SII were randomly divided into control (n = 11) and PFC groups (n = 12). The control group received conventional treatment (anti-infection, nutritional support, antishock measures, and supportive treatment). The PFC group received endotracheal perfluorohexane instillation in addition to conventional treatment. On admission and 3 days later, therapeutic effects were evaluated and inflammatory mediators in bronchoalveolar lavage fluid and plasma were analyzed. There was no significant difference between the control and PFC group on admission. After 3 days, perfluorohexane treatment significantly (P < .05) increased lung dynamic compliance, and reduced alveolar-arterial oxygen gradient, Acute Physiology and Chronic Health Evaluation II score, percentage of neutrophils, and levels of interleukin-6, interleukin-8, and tumor necrosis factor alpha in bronchoalveolar lavage fluid; there was no significant change in the control group before and after treatment. Intratracheal instillation of perfluorohexane modulates the pulmonary immune microenvironment and supplements current conventional treatments for burn patients with SII.

Protective effect of suppressing STAT3 activity in LPS-induced acute lung injury.

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are diseases with high mortality. Macrophages and neutrophils are responsible for inflammatory responses in ALI and ARDS, which are characterized by excessive production of proinflammatory mediators in bronchoalveolar lavage fluid (BALF) and plasma. Aberrant activation of the JAK/STAT pathway is critical for persistent inflammation in many conditions such as infection and autoimmunity. Given the importance of the STAT3 transcription factor in activating macrophages and neutrophils and augmenting inflammation, we investigated the therapeutic potential of inhibiting STAT3 activity using the small-molecule STAT3 inhibitor, LLL12. Our results demonstrate that LPS induces STAT3 activation in macrophages in vitro and in CD45+CD11b+ cells from BALF in the LPS-induced ALI model in vivo. LLL12 treatment inhibits LPS-induced lung inflammation in the ALI model, which is accompanied by suppression of LPS-induced STAT3 activation and an inhibition of macrophage and inflammatory cell infiltration in lung and BALF. LLL12 treatment also suppresses expression of proinflammatory genes including IL-1β, IL-6, TNF-α, iNOS, CCL2, and MHC class II in macrophages and inflammatory cells from BALF and serum as determined by ELISA. Furthermore, hyperactivation of STAT3 in LysMCre-SOCS3fl/fl mice accelerates the severity of inflammation in the ALI model. Both pre- and post-LPS treatment with LLL12 decrease LPS-induced inflammatory responses in mice with ALI. Importantly, LLL12 treatment attenuates STAT3 phosphorylation in human peripheral blood mononuclear cells induced by plasma from patients with ARDS, which suggests the feasibility of targeting the STAT3 pathway therapeutically for patients with ALI and ARDS.

The Role of Toll-Like Receptors and Myeloid Differentiation Factor 88 in Bjerkandera adusta-Induced Lung Inflammation

Background: Recently, a cluster of patients with an intractable allergic fungal cough who were characterized by sensitization to Bjerkandera adusta was reported. In the present study, the role of Toll-like receptors and myeloid differentiation factor 88 (MyD88) in B. adusta-induced lung inflammation was investigated. Methods: Wild-type (WT), TLR2^-/-,TLR4^-/-, and MyD88^-/- BALB/c mice were intratracheally challenged with B. adusta 4 times at 2-week intervals. Lung pathology, bronchoalveolar lavage fluid (BALF) cytological profiles, and inflammatory mediators in BALF were investigated. Bone marrow-derived macrophages (BMDM) from TLR2^-/-,TLR4^-/-, TLR2/4^-/-, TLR7/9^-/-,MyD88^-/-, and WT C57BL/6J mice were stimulated with B. adusta for 12 h, and inflammatory mediators in the culture medium were measured. Results:B. adusta caused lung inflammation along with Th2 cytokine [interleukin (IL)-5 and IL-13] and eosinophil-related chemokine [eotaxin and monocyte chemotactic protein (MCP-3)] production, an increase in eosinophils in BALF, and eosinophil infiltration in the airways in WT and TLR4^-/- mice. However, Th2 and eosinophil-related responses in TLR2^-/- and MyD88^-/- mice were low or undetectable. The induction of neutrophils and IL-6, IL-12, IL-17A, and MCP-1 in the BALF of MyD88^-/- mice was attenuated compared to that in WT mice. The induction of IL-6, TNF-α, MCP-1, and macrophage inflammatory protein-1α was reduced or undetectable in B. adusta-stimulated BMDM from TLR7/9^-/- and MyD88^-/- mice compared to WT mice. Conclusions: These results suggest that TLR2 and the adapter protein MyD88 may play an important role in the induction of eosinophils by B. adusta. However, TLR7/9-MyD88 might be important in the induction of neutrophils and the relevant inflammatory mediators, especially IL-17A.