Bronchoalveolar Lavage Fluid Protein Research Articles

Effects of Dexmedetomidine on Inflammatory Factors and Mitogen Activated Protein Kinase-Related Signaling Pathways of Alveolar Lavage Fluid in Mice with Lipopolysaccharide-Induced Acute Lung Injury

To investigate the effects and mechanisms of Dexmedetomidine on inflammatory factors and mitogenactivated protein kinase related signaling pathways in alveolar lavage fluid of mice with acute lung injury induced by lipopolysaccharide. 36 clean-grade male Kunming mice were randomly divided into blank control group (normal control group), lipopolysaccharide-induced acute lung injury model group, and lipopolysaccharide+dexmedetomidine group, with 12 mice in each group. The lipopolysaccharide group mice were intraperitoneally injected with 0.1 ml 5 mg kg-1 Lipopolysaccharide, lipopolysaccharide+Dexmedetomidine mice were injected with 25 μg kg-1. Dexmedetomidine intraperitoneally with 0.1 ml and 1 h later with 5 mg kg-1. Lipopolysaccharide injected with 0.1 ml intraperitoneally and the blank control group mice were intraperitoneally injected with normal saline of the same volume as the lipopolysaccharide group. All mice in each group were sacrificed after intraperitoneal injection for 6 h, and their lung tissues were separated by thoracotomy and their bronchoalveolar lavage fluid was recovered for subsequent detection. Analysis groups of mice lung tissue pathology, compare groups of mice lung tissue pathology score, lung wet weight/dry weight (W)/(D), mitogen-activated protein kinase pathways (p38 lightning, extracellular signal regulating kinase, c-jun amino terminal kinase) and bronchoalveolar lavage fluid protein expression level of tumor necrosis factor alpha beta, interleukin-1β, myeloperoxidase, protein concentration. In the blank control group, lung tissue structure was basically normal and alveolar structure was intact. lipopolysaccharide group mice had obvious lung tissue damage, and inflammatory cells appeared in the alveoli, with significantly increased exudation of red blood cells and proteins. Lipopolysaccharide+Dexmedetomidine group mice lung tissue damage significantly improved, inflammatory cell infiltration decreased, and red blood cell, protein exudation significantly decreased. Lung histopathological scores and bronchoalveolar lavage fluid levels of tumor necrosis factor alpha, interleukin- 1β and myeloperoxidase in lipopolysaccharide group were significantly higher than those in blank control group (p<0.05). The lung histopathological scores and the levels of tumor necrosis factor alpha, interleukin- 1β and myeloperoxidase in bronchoalveolar lavage fluid of lipopolysaccharide+Dexmedetomidine group were significantly lower than those of lipopolysaccharide group (p<0.05). The protein concentration in W/D and bronchoalveolar lavage fluid in lipopolysaccharide group was significantly higher than that in blank control group (p<0.05). Protein concentration in W/D and bronchoalveolar lavage fluid of lipopolysaccharide+Dexmedetomidine group was significantly lower than that of lipopolysaccharide group (p<0.05). The protein expression levels of extracellular signal regulating kinase, c-jun amino terminal kinase and P-P38 in THE lipopolysaccharide group were significantly higher than those in the blank control group (p<0.05). The protein expression levels of extracellular signal regulating kinase, c-jun amino terminal kinase and P-P38 in the lipopolysaccharide+Dexmedetomidine group were significantly lower than those in the lipopolysaccharide group (p<0.05). There was no significant difference in extracellular signal regulating kinase, c-jun amino terminal kinase and p38 protein expression levels in each group (p>0.05). Dexmedetomidine can significantly alleviate acute lung injury induced by lipopolysaccharide in mice, and its mechanism may be realized by blocking the activation of mitogen-activated protein kinaserelated signaling pathway and thereby reducing inflammatory response, providing a new target for the treatment of sepsis induced acute lung injury in the futurey.

Astragaloside IV alleviates PM2.5-induced lung injury in rats by modulating TLR4/MyD88/NF-κB signalling pathway

ObjectiveAstragaloside IV (AS IV) is antioxidant and anti-inflammatory product, which is extracted from the Chinese herb Astragalus membranaceus. It is widely used in a variety of inflammatory diseases. The research was to explored the protective effects of AS IV against lung injury induced by particulate matter 2.5 (PM2.5) in vivo. Subjects and methodsThirty-five male Sprague-Dawley rats were randomly divided into five groups (n=7 per group). (1) Normal saline group (NS), (2) AS IV group (AS) (100 mg/kg), (3) PM2.5 group (PM2.5), (4) PM2.5 + AS IV group (ASL) (50 mg/kg), and (5) PM2.5 + AS IVgroup (ASH) (100 mg/kg). Rats were pre-treated with AS IV intraperitoneally (50 and 100 mg/kg/day) for three days. Then, PM2.5 (7.5 mg/kg) was given by intratracheal instillation to induce lung injury. Six hours after PM2.5 stimulation, the rats were euthanized. Bronchoalveolar lavage fluid (BALF) was collected for assay of cytokines. Lung tissue was collected for oxidative stress, histology, immunohistochemistry, transmission electron microscope, and western blot analyses. ResultsAS IV alleviated PM2.5-induced lung injury by decreasing lung dry-wet ratio, reducing the level of interleukin-6 (IL-6), tumour necrosis factor-alpha (TNF-α), and C-reactive protein (CRP) in BALF, and reduced oxidative stress response in lung tissue. Western blot results revealed that AS-IV regulated the expression of TLR4/MyD88/NF-κB pathway proteins in lung tissues. ConclusionAS IV mitigated PM2.5 induced lung injury by regulating the activity of TLR4/MyD88/NF-κB signalling pathway, reducing inflammatory and oxidative stress responses.

JNK‑IN‑8 treatment alleviates lipopolysaccharide‑induced acute lung injury via suppression of inflammation and oxidative stress regulated by JNK/NF‑κB signaling.

JNK serves critical roles in numerous types of inflammation- and oxidative stress-induced disease, including acute lung injury (ALI). JNK-IN-8 is the first irreversible JNK inhibitor that has been described. However, whether JNK-IN-8 can prevent lipopolysaccharide (LPS)-induced ALI by inhibiting JNK activation and its downstream signaling is poorly understood. The objective of the present study was to investigate the specific therapeutic effects of JNK-IN-8 on LPS-induced ALI and the molecular mechanisms involved. JNK-IN-8 attenuated myeloperoxidase activity, malondialdehyde and superoxide dismutase content and the lung wet/dry ratio, and improved the survival rate following lethal injection of LPS. Additionally, JNK-IN-8 decreased bronchoalveolar lavage fluid protein levels, lactate dehydrogenase activity, neutrophil infiltration and the number of macrophages (as demonstrated by flow cytometry), as well as the production of TNF-α, IL-6 and IL-1β (as evaluated via ELISA). In addition, reverse transcription-quantitative PCR and ELISA showed that JNK-IN-8 attenuated LPS-induced inflammatory cytokine production and oxidative stress in primary murine peritoneal macrophages and RAW264.7 cells in vitro. Furthermore, the present study demonstrated that the JNK/NF-κB signaling pathway was involved in the therapeutic effect of JNK-IN-8 against LPS-induced injury both in vivo and in vitro. In conclusion, these findings indicated that JNK-IN-8 had a therapeutic effect on LPS-induced ALI in mice. The mechanism may be associated with inhibition of the JNK/NF-κB signaling pathway. JNK-IN-8 may be a potential therapeutic agent for the treatment of ALI.

Protective effect of FOXP3-mediated miR-146b-5p/Robo1/NF-κB system on lipopolysaccharide-induced acute lung injury in mice.

BackgroundAs a key transcription factor, forkhead box protein 3 (FOXP3) plays an important role in the development and function of natural cluster of differentiation 4 [CD4 (+)] regulatory T cells (Treg cells). However, the function of FOXP3 in Lipopolysaccharide (LPS)-induced acute lung injury (ALI) through regulating miR-146b-5p is unclear. This research aimed to disclose the regulatory effect of the FOXP3-mediated miR-146b-5p/Roundabout 1 (Robo1)/NF-κB system on LPS-induced ALI in mice.MethodsThe mice were subjected to 5 mg/kg of LPS via intratracheal instillation to induce ALI and generate the ALI model. Mice was divided into five group, including control group, ALI group, ALI + FOXP3 group, the ALI + miR antagomir group and ALI + miR antagomir+ FOXP3 group. Lung tissue injury were detected by hematoxylin and eosin (HE) staining. Lung wet/dry weight ratio, total cells in bronchoalveolar lavage fluid (BALF), total protein in BALF and the polymorphonuclear leukocyte (PMN) in BALF were detected. The levels of tumor necrosis factor-α (TNF-α), Interleukin 6 (IL-6) and IL-1β were detected by enzyme-linked immunosorbent assay (ELISA) kit. The dual-luciferase reporter assay were used to detect the target relationship between FOXP3 and Robo1. Mice was divided into five group, including control group, ALI group, ALI + FOXP3 group, ALI + Robo1 group and ALI + FOXP3+ Robo1 group. The protein levels of FOXP3, Robo1 and p-p65 were detected by western bolt. The mRNA levels of miR-146b-5p and Robo1 were detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR).ResultsAlthough protein expression levels of FOXP3 were significantly down-regulated in the ALI model, the increased FOXP3 levels promoted an increase in miR-146b-5p. Compared with the control group, the ALI model group exhibited severe histopathologic injury, such as thickening of the alveolar wall, pulmonary congestion, and decreased alveolar numbers. By mediating the overexpression of miR-146b-5p, FOXP3 also increased alveolar clearance and inhibited inflammatory responses in the ALI model. Importantly, Robo1 is a potential target of miR-146b-5p.ConclusionsFOXP3 could inhibit NF-κB activation, reduce lung pathological damage, and inhibit inflammatory responses by mediating the miR-146b-5p/Robo1/NF-κB system in the ALI model. These results may provide a new potential target for the treatment of ALI disease.

The pulmonary toxicity of carboxylated or aminated multi-walled carbon nanotubes in mice is determined by the prior purification method

BackgroundInhalation of multi-walled carbon nanotubes (MWCNTs) poses a potential risk to human health. In order to safeguard workers and consumers, the toxic properties of MWCNTs need to be identified. Functionalization has been shown to either decrease or increase MWCNT-related pulmonary injury, depending on the type of modification. We, therefore, investigated both acute and chronic pulmonary toxicity of a library of MWCNTs derived from a common pristine parent compound (NC7000).MethodsMWCNTs were thermally or chemically purified and subsequently surface functionalized by carboxylation or amination. To evaluate pulmonary toxicity, male C57BL6 mice were dosed via oropharyngeal aspiration with either 1.6 or 4 mg/kg of each MWCNT type. Mitsui-7 MWCNT was used as a positive control. Necropsy was performed at days 3 and 60 post-exposure to collect bronchoalveolar lavage fluid (BALF) and lungs.ResultsAt day 3 all MWCNTs increased the number of neutrophils in BALF. Chemical purification had a greater effect on pro-inflammatory cytokines (IL-1β, IL-6, CXCL1) in BALF, while thermal purification had a greater effect on pro-fibrotic cytokines (CCL2, OPN, TGF-β1). At day 60, thermally purified, carboxylated MWCNTs had the strongest effect on lymphocyte numbers in BALF. Thermally purified MWCNTs caused the greatest increase in LDH and total protein in BALF. Furthermore, the thermally purified and carboxyl- or amine-functionalized MWCNTs caused the greatest number of granulomatous lesions in the lungs. The physicochemical characteristics mainly associated with increased toxicity of the thermally purified derivatives were decreased surface defects and decreased amorphous content as indicated by Raman spectroscopy.ConclusionsThese data demonstrate that the purification method is an important determinant of lung toxicity induced by carboxyl- and amine-functionalized MWCNTs.

Pulmonary surfactant and drug delivery: Vehiculization, release and targeting of surfactant/tacrolimus formulations

This work explores the potential for strategizing pulmonary surfactant (PS) for drug delivery over the respiratory air-liquid interface: the interfacial delivery. The efficacy of PS- and interface-assisted drug vehiculization was determined both in vitro and in vivo using a native purified porcine PS combined with the hydrophobic anti-inflammatory drug Tacrolimus (TAC), a calcineurin inhibitor. In vitro assays were conducted in a novel double surface balance setup designed to emulate compression-expansion dynamics applied to interfacially connected drug donor and recipient compartments. In this setup, PS transported TAC efficiently over air-liquid interfaces, with compression/expansion breathing-like dynamics enhancing rapid interface-assisted diffusion and drug release. The efficacy of PS-assisted TAC vehiculization was also evaluated in vivo in a mouse model of lipopolysaccharide (LPS)-induced acute lung injury (ALI). In anesthetized mice, TAC combined with PS was intra-nasally (i.n) instilled prior administering i.n. LPS. PS/TAC pre-treatment caused greater TAC internalization into a higher number of lung cells obtained from bronchoalveolar lavages (BAL) than TAC pre-treatment alone. Additionally, the PS/TAC combination but not TAC or PS alone attenuated the LPS-induced pro-inflammatory effects reducing cells and proteins in BAL fluid. These findings indicated that PS-mediated increase in TAC uptake blunted the pro-injurious effects of LPS, suggesting a synergistic anti-inflammatory effect of PS/drug formulations. These in vitro and in vivo results establish the potential utility of PS to open novel effective delivery strategies for inhaled drugs.

Echinacoside attenuates lipopolysaccharide-induced acute lung injury in newborn mice via inactivation of NF- κB/NLRP3 signaling pathway

Purpose: To investigate the effect of echinacoside (ECH) on acute lung injury (ALI) and the underlying mechanism of action.Methods: The ALI model was established through intranasal instillation of lipopolysaccharide (LPS). Lung tissue damage was determined using hematoxylin and eosin (H&amp;E) staining and lung wet-to-dry–weight ratio. Bronchoalveolar lavage fluid (BALF) protein concentration, cell count, and cytokine level were evaluated. Western blotting was used to determine protein expression level.Results: ECH attenuated lung tissue injury and lung wet-to-dry–weight ratio in the ALI model (p &lt; 0.01). The total protein content and number of total cells, neutrophils, and macrophages increased in BALF of mice treated with LPS, but these increases were reversed by ECH treatment (p &lt; 0.01). The levels of TNF-α and IL-1β increased in BALF and lung tissue of LPS-treated mice; however, ECH treatment decreased these changes (p &lt; 0.01). In addition, ECH inhibited the activation of the nuclear factor-κB (NF-κB)/NLR family pyrin domain containing 3 (NLRP3) pathway in LPS-treated mice (p &lt; 0.01).Conclusion: Echinacoside attenuates LPS-induced ALI via inactivation of the NF-κB/NLRP3 pathway, making echinacoside a potential drug for the treatment of ALI.&#x0D; Keywords: Echinacoside, Acute lung injury, Lipopolysaccharide, Nuclear factor-κB, NLR family pyrin domain containing 3

Proteomic characteristics of bronchoalveolar lavage fluid in critical COVID-19 patients.

Up to 10-20% of patients with coronavirus disease 2019 (COVID-19) develop a severe pulmonary disease due to immune dysfunction and cytokine dysregulation. However, the extracellular proteomic characteristics in respiratory tract of these critical COVID-19 patients still remain to be investigated. In the present study, we performed a quantitative proteomic analysis of the bronchoalveolar lavage fluid (BALF) from patients with critical COVID-19 and from non-COVID-19 controls. Our study identified 358 differentially expressed BALF proteins (P<0.05), among which 41 were significantly changed after using the Benjamini-Hochberg correction (q<0.05). The up-regulated signaling was found to be mainly involved in inflammatory signaling and response to oxidative stress. A series of increased extracellular factors including Tenascin-C (TNC), Mucin-1 (KL-6 or MUC1), Lipocalin-2 (LCN2), periostin (POSTN), Chitinase 3-like 1 (CHI3L1 or YKL40), and S100A12, and the antigens including lymphocyte antigen 6D/E48 antigen (LY6D), CD9 antigen, CD177 antigen, and prostate stem cell antigen (PSCA) were identified, among which the proinflammatory factors TNC and KL-6 were further validated in serum of another thirty-nine COVID-19 patients and healthy controls, showing high potentials of being biomarkers or therapeutic candidates for COVID-19. This BALF proteome associated with COVID-19 would also be a valuable resource for researches on anti-inflammatory medication and understanding the molecular mechanisms of host response. DATABASE: Proteomic raw data are available in ProteomeXchange (http://proteomecentral.proteomexchange.org) under the accession number PXD022085, and in iProX (www.iprox.org) under the accession number IPX0002429000.

Baicalin Liposome Alleviates Lipopolysaccharide-Induced Acute Lung Injury in Mice via Inhibiting TLR4/JNK/ERK/NF-κB Pathway.

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are challenging diseases with the high mortality in a clinical setting. Baicalin (BA) is the main effective constituent isolated from the Chinese medical herb Scutellaria baicalensis Georgi, and studies have proved that it has a protective effect on ALI induced by lipopolysaccharide (LPS) due to the anti-inflammatory efficacy. However, BA has low solubility which may limit its clinical application. Hence, we prepared a novel drug delivery system—Baicalin liposome (BA-LP) in previous research—which can improve some physical properties of BA. Therefore, we aimed to explore the effect of BA-LP on ALI mice induced by LPS. In pharmacokinetics study, the values of t1/2 and AUC0-t in the BA-LP group were significantly higher than that of the BA group in normal mice, indicating that BA-LP could prolong the duration time in vivo of BA. The BA-LP group also showed a higher concentration in lung tissues than the BA group. Pharmacodynamics studies showed that BA-LP had a better effect than the BA group at the same dosage on reducing the W/D ratio, alleviating the lung injury score, and decreasing the proinflammatory factors (TNF-α, IL-1β) and total proteins in bronchoalveolar lavage fluids (BALF). In addition, the therapeutic effects of BA-LP showed a dose-dependent manner. Western blot analysis indicated that the anti-inflammatory action of BA could be attributed to the inhibition of the TLR4-NFκBp65 and JNK-ERK signaling pathways. These results suggest that BA-LP could be a valuable therapeutic candidate in the treatment of ALI.

Clinical significance of CC16 and IL-12 in bronchoalveolar lavage fluid of various stages of silicosis.

Identification of novel biomarkers for silicosis could be helpful for disease diagnosis and pathophysiological mechanism exploration. Our study aims to investigate the Clara cell secretory 16-kd protein (CC16) and interleukin-12 (IL-12) levels in bronchoalveolar lavage fluid (BALF) in patients with silicosis at various stages. The enzyme-linked immunosorbent assay (ELISA) double antibody sandwich method was used to determine the CC16 and IL-12 in BALF levels from 79 patients with silicosis of various stages. Correlation analyses were performed between CC16 and IL-12 levels, and lung function and cytological counts in patients with silicosis at various stages. There were no significant differences in the BALF recovery volume, the number of cells, percentages of macrophages and lymphocytes in the alveolar lavage fluid of patients with silicosis in different stages (P>0.05); the percentage of neutrophils in stage I and stage II were higher than the control group (P<0.05) with statistically significant differences. The CC16 in BALF levels in stage I and II silicosis groups lower than the control group and stage III silicosis group with statistically significant differences (P<0.05), whereas CC16 levels in stage II silicosis group are higher than the stage I group (P<0.01). The IL-12 levels were higher than the control group (P<0.01), and the IL-12 levels in stage II and III silicosis group was higher than the stage I silicosis group (P<0.01). With the increase of the length of dust service, the CC16 and IL-12 levels decreased and showed a positive correlation between these indexes (correlation coefficient r=0.559, P<0.01). In addition, CC16 silicosis patient levels were positively correlated with FEV1/FVC and VCmax (r=0.242, 0.257; both P<0.05); IL-12 levels were negatively correlated with FEV1 and VC max (r=-0.250, -0.483; both P<0.05). The CC16 and IL-12 levels may have a specific reference value for the early diagnosis of silicosis and the assessment of lung function.

The anti-asthmatic potential of flavonol kaempferol in an experimental model of allergic airway inflammation

Flavonol kaempferol possesses a broad spectrum of potent pharmacological activities that seem to be effective in the modulation of allergic respiratory diseases. In our study, an experimental animal model of ovalbumin (OVA)-induced allergic airway inflammation in guinea pigs was used to determine the anti-asthmatic potential of kaempferol. The parameters of specific airway resistance (sRaw) and cough reflex response were evaluated in vivo. In vitro, an assessment of tracheal smooth muscle (TSM) contractility and analyses of inflammatory cytokines (IL-4, IL-5, IL-13, GM-CSF, IFN-γ), transforming growth factor (TGF-β1), immune cells count and ciliary beating frequency (CBF) were performed. Both single (6, 20 mg/kg b. w. p. o.) and long-term administered doses of kaempferol (20 mg/kg b. w. p. o., 21 days) suppressed sRaw provoked by histamine in conscious animals. The administration of kaempferol for 21 days attenuated histamine-induced TSM contractility in vitro and ameliorated the progression of chronic airway inflammation by decreasing the levels of IL-5, IL-13, GM-CSF, eosinophil count in bronchoalveolar lavage (BAL) fluid and TGF-β1 protein level in lung tissue. Kaempferol also eliminated the alterations in cough reflex sensitivity invoked by OVA-sensitization, but it did not affect CBF. The results demonstrate that flavonol kaempferol can modulate allergic airway inflammation and associated asthma features (AHR, aberrant stimulation of cough reflex).

Thalictrum minus L. ameliorates particulate matter-induced acute lung injury in mice

Ethnopharmacological relevanceThalictrum minus L., which is widespread across Eurasia, is utilized as a folk medicine for treating dysentery, bedsore, fungal infection and lung inflammation in China, Mongolia and Iran. Aim of the studyA Mongolian folk medicinal plant named Thalictrum minus L. (TML) has been extensively used for the treatment of lung inflammation, bacterial and fungal infection and tuberculosis. Our present study aims to investigate the effectiveness of TML against particulate matter (PM)-induced acute lung injury (ALI) and the potential underlying mechanisms. Materials and methodsInitially, HPLC-Q-TOF was applied for the qualitative analysis and HPLC was used for quantitative analysis of main components in TML. Then, the mice model of ALI was induced by PM via intratracheally instilled with 50 mg/kg body weight of Standard Reference Material1648a (SRM1648a), and TML (10, 20, 40 mg/kg) were administered orally 1 h prior to PM. The efficacy and molecular mechanisms in the presence or absence of TML were elucidated. ResultsEleven main ingredients were detected in TML and the contents of homoorientin and berberine were quantified. Additionally, the results demonstrated that TML profoundly inhibited weight loss in mice and ameliorated lung pathological injury induced by PM. Furthermore, we also found that TML significantly decreased the lung wet to dry weight (W/D) ratios, reduced total protein in bronchoalveolar lavage fluid (BALF), and effectively attenuated PM-induced increased leukocyte and macrophages in BALF. Meanwhile, TML could pronouncedly inhibited myeloperoxidase (MPO) activity in lung tissues, decreased the PM-induced inflammatory cytokines including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β), reduced nitric oxide (NO) and increased superoxide dismutase (SOD) in BALF. In addition, TML markedly facilitated the expression of p-AMPK-Nrf2 and suppressed the expression of KEAP, prohibited the activation of the MAPKs-NLRP3/caspase-1 and cyclooxygenase-2 (COX2), and inhibited apoptotic pathways. ConclusionThese findings indicated that TML attenuated PM-induced ALI through suppressing the release of inflammatory cytokines and alleviating oxidative damage correlated with the AMPK-Nrf2/KEAP signaling pathways, MAPKs-NLRP3/caspase-1 signaling pathways, as well as apoptotic pathways.

Reduced allergic lung inflammation and airway responsiveness in mice lacking the cytoskeletal protein gelsolin.

Airway smooth muscle hyperresponsiveness associated with chronic airway inflammation leads to the typical symptoms of asthma including bronchoconstriction and wheezing. Asthma severity is associated with airway inflammation; therefore, reducing airway inflammation is an important therapeutic target. Gelsolin is an actin capping and severing protein that has been reported to be involved in modulation of the inflammatory response. Using mice genetically lacking gelsolin, we evaluated the role of gelsolin in the establishment of house dust mite (HDM) antigen-induced allergic lung inflammation. The genetic absence of gelsolin was found to be protective against HDM sensitization, resulting in reduced lung inflammation, inflammatory cytokines, and Muc5AC protein in bronchoalveolar lavage (BAL) fluid. The number of eosinophils, lymphocytes, and interstitial macrophages in the BAL were increased after HDM sensitization in wild-type mice but were attenuated in gelsolin-null mice. The observed attenuation of inflammation may be partly due to delayed migration of immune cells, because the reduced eosinophils in the BALs from gelsolin-null mice compared with controls occurred despite similar amounts of the chemoattractant eotaxin. Splenic T cells demonstrated similar proliferation rates, but ex vivo alveolar macrophage migration was delayed in gelsolin-null mice. In vivo, the reduced lung inflammation after HDM sensitization in gelsolin-null mice was associated with significantly diminished airway resistance to inhaled methacholine compared with HDM-treated wild-type mice. Our results suggest that modulation of gelsolin expression or function in selective inflammatory cell types that modulate allergic lung inflammation could be a therapeutic approach for asthma.

Novel protein pathways in development and progression of pulmonary sarcoidosis

Pulmonary involvement occurs in up to 95% of sarcoidosis cases. In this pilot study, we examine lung compartment-specific protein expression to identify pathways linked to development and progression of pulmonary sarcoidosis. We characterized bronchoalveolar lavage (BAL) cells and fluid (BALF) proteins in recently diagnosed sarcoidosis cases. We identified 4,306 proteins in BAL cells, of which 272 proteins were differentially expressed in sarcoidosis compared to controls. These proteins map to novel pathways such as integrin-linked kinase and IL-8 signaling and previously implicated pathways in sarcoidosis, including phagosome maturation, clathrin-mediated endocytic signaling and redox balance. In the BALF, the differentially expressed proteins map to several pathways identified in the BAL cells. The differentially expressed BALF proteins also map to aryl hydrocarbon signaling, communication between innate and adaptive immune response, integrin, PTEN and phospholipase C signaling, serotonin and tryptophan metabolism, autophagy, and B cell receptor signaling. Additional pathways that were different between progressive and non-progressive sarcoidosis in the BALF included CD28 signaling and PFKFB4 signaling. Our studies demonstrate the power of contemporary proteomics to reveal novel mechanisms operational in sarcoidosis. Application of our workflows in well-phenotyped large cohorts maybe beneficial to identify biomarkers for diagnosis and prognosis and therapeutically tenable molecular mechanisms.

MCTR1 enhances the resolution of lipopolysaccharide-induced lung injury through STAT6-mediated resident M2 alveolar macrophage polarization in mice.

Acute respiratory distress syndrome (ARDS) is a fatal disease characterized by excessive infiltration of inflammatory cells. MCTR1 is an endogenously pro‐resolution lipid mediator. We tested the hypothesis that MCTR1 accelerates inflammation resolution through resident M2 alveolar macrophage polarization. The mice received MCTR1 via intraperitoneal administration 3 days after LPS stimulation, and then, the bronchoalveolar lavage (BAL) fluid was collected 24 hours later to measure the neutrophil numbers. Flow cytometry was used to sort the resident and recruited macrophages. Post‐treatment with MCTR1 offered dramatic benefits in the resolution phase of LPS‐induced lung injury, including decreased neutrophil numbers, reduced BAL fluid protein and albumin concentrations and reduced histological injury. In addition, the expression of the M2 markers Arg1, FIZZ1, Remlα, CD206 and Dectin‐1 was increased on resident macrophages in the LPS + MCTR1 group. Resident macrophage depletion abrogated the therapeutic effects of MCTR1, and reinjection of the sorted resident macrophages into the lung decreased neutrophil numbers. Finally, treatment with MCTR1 increased STAT6 phosphorylation. The STAT6 inhibitor AS1517499 abolished the beneficial effects of MCTR1. In conclusion, MCTR1 promotes resident M2 alveolar macrophage polarization via the STAT6 pathway to accelerate resolution of LPS‐induced lung injury.

Protective effect of S-nitrosoglutathione pretreatment on acute lung injury in septic rats.

Objective(s):To investigate the protective effect of S-nitrosoglutathione (SNG) pretreatment on acute lung injury (ALI) in septic rats. Materials and Methods:We constructed a rat model of sepsis by cecal ligation and perforation (CLP), and randomly divided into Sham, CLP, and CLP+SNG (0.25 and 0.5 mg/kg) groups. We used H&E staining and lung wet/dry ratio to assess the severity of lung injury, detected the levels of protein and cells in bronchoalveolar lavage fluid (BALF) and the levels of TNF-α, IL-1β, TLR4 mRNA, and NF-κB p65 mRNA in the lung tissue, and assessed the levels of glutathione (GSH), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase in the lung tissue.Results:A rat model of sepsis was successfully constructed by CLP, and pretreatment of SNG significantly increased the survival of septic rats (P<0.001) and decreased the lung tissue injury scores (P<0.001) and lung wet/dry ratio (P<0.01) in a dose-dependent manner. Furtherly, SNG pretreatment significantly reduced the number of total cells, total protein, neutrophils, and lympholytes (all P<0.001) in BALF, and which also decreased the levels of TNF-α, IL-1β, TLR4 mRNA, and NF-κB p65 mRNA (all P<0.001) in the lungs of CLP-induced rats. Moreover, pretreatment of SNG significantly increased the levels of anti-oxidant enzymes GSH, SOD, GSH-Px, and catalase (all P<0.001) in the lung tissue of septic rats. Conclusion:SNG pretreatment has a protective effect on ALI in septic rats, and the specific mechanism may be related to anti-endotoxic, anti-inflammatory, and anti-oxidative properties.

Glucagon-like peptide-1 receptor mediates the beneficial effect of liraglutide in an acute lung injury mouse model involving the thioredoxin-interacting protein.

Repurposing clinically used drugs is among the important strategies in drug discovery. Glucagon-like peptide-1 (GLP-1) and its diabetes-based drugs, such as liraglutide, possess a spectrum of extra-pancreatic functions, while GLP-1 receptor (GLP-1R) is most abundantly expressed in the lung. Recent studies have suggested that GLP-1-based drugs exert beneficial effects in chronic, as well as acute, lung injury rodent models. Here, we show that liraglutide pretreatment reduced LPS induced acute lung injury in mice. It significantly reduced lung injury score, wet/dry lung weight ratio, bronchoalveolar lavage fluid immune cell count and protein concentration, and cell apoptosis in the lung, and it was associated with reduced lung inflammatory cytokine and chemokine gene expression. Importantly, these effects were virtually absent in GLP-1R−/− mice. A well-known function of GLP-1 and GLP-based drugs in pancreatic β-cells is the attenuation of high-glucose stimulated expression of thioredoxin-interacting protein (TxNIP), a key component of inflammasome. LPS-challenged lungs showed elevated TxNIP mRNA and protein expression, which was attenuated by liraglutide treatment in a GLP-1R-dependent manner. Hence, our observations suggest that GLP-1R is essential in mediating beneficial effects of liraglutide in acute lung injury, with the inflammasome component TxNIP as a potential target.

Circulating and tissue biomarkers as predictors of bromine gas inhalation.

The threat from deliberate or accidental exposure to halogen gases is increasing, as is their industrial applications and use as chemical warfare agents. Biomarkers that can identify halogen exposure, diagnose victims of exposure or predict injury severity, and enable appropriate treatment are lacking. We conducted these studies to determine and validate biomarkers of bromine (Br2 ) toxicity and correlate the symptoms and the extent of cardiopulmonary injuries. Unanesthetized rats were exposed to Br2 and monitored noninvasively for clinical scores and pulse oximetry. Animals were euthanized and grouped at various time intervals to assess brominated fatty acid (BFA) content in the plasma, lung, and heart using mass spectrometry. Bronchoalveolar lavage fluid (BALF) protein content was used to assess pulmonary injury. Cardiac troponin I (cTnI) was assessed in the plasma to evaluate cardiac injury. The blood, lung, and cardiac tissue BFA content significantly correlated with the clinical scores, tissue oxygenation, heart rate, and cardiopulmonary injury parameters. Total (free + esterified) bromostearic acid levels correlated with lung injury, as indicated by BALF protein content, and free bromostearic acid levels correlated with plasma cTnI levels. Thus, BFAs and cardiac injury biomarkers can identify Br2 exposure and predict the severity of organ damage.

P53 deficiency potentiates LPS-Induced acute lung injury in vivo.

Acute Lung Injury (ALI) and Acute Respiratory Distress Syndrome (ARDS) represent a significant cause of morbidity and mortality in critically ill hospitalized patients. Emerging evidence suggest that the expression levels of P53 in the lungs are associated with the supportive effects of heat shock protein 90 inhibitors and growth hormone releasing hormone antagonists in the endothelium. In the current study, we employed an in vivo model of intratracheal administration of lipopolysaccharides (LPS)-induced ALI to investigate the role of P53 in counteracting LPS-induced lung inflammatory responses. In wild type mice, LPS induced the expression of IL-1α, IL-1β, and TNFα in the lungs, increased bronchoalveolar lavage fluid protein concentration, and activated cofilin. Remarkably; those responses were more potent in P53 knockout mice, suggesting the crucial role of P53 in orchestrating rigorous endothelial defenses against inflammatory stimuli. The present study supports previous endeavors on the protective role of P53 against lung inflammatory disease, and enrich our knowledge on the development of medical countermeasures against ARDS.

Associations of soluble metals and lung and liver toxicity in mice induced by fine particulate matter originating from a petrochemical complex.

Adverse health effects have been observed in nearby residents due to exposure to petrochemical-derived chemicals. The objective of this study was to examine associations of soluble metals with lung and liver toxicity in fine particulate matter (PM2.5) in the vicinity of a petrochemical complex. PM2.5 was collected in the vicinity of a petrochemical complex of Mailiao Township (Yunlin County, Taiwan) to investigate lung and liver toxicity in BALB/c mice. The PM2.5 concentration was 30.2 ± 11.2μg/m3, and the PM2.5 was clustered in major local emissions (19.1μg/m3) and minor local emissions (14.1μg/m3) using a k-means clustering model. The PM2.5 (50 and 150μg/kg) and PM2.5-equivalent soluble nickel (Ni), vanadium (V), and lead (Pb) concentrations were intratracheally instilled into BALB/c mice. PM2.5 and V significantly decreased the tidal volume after exposure (p< 0.05). The peak expiratory flow (PEF) and peak inspiratory flow (PIF)/PEF ratio were significantly altered by 150μg/kgV (p< 0.05). V and Pb significantly increased total protein and lactate dehydrogenase (LDH) levels in bronchoalveolar lavage fluid (BALF) (p< 0.05). Interleukin (IL)-6 in BALF significantly increased after exposure to Pb (p< 0.05) accompanied by lung inflammatory infiltration. PM2.5 and Pb significantly increased levels of 8-isoprostane (p< 0.05). The level of caspase-3 activity significantly increased after exposure to Pb (p< 0.05). LDH in the liver was significantly increased by PM2.5 (p< 0.05). 8-Isoprostane in the liver was significantly increased by PM2.5 and Pb (p< 0.05). IL-6 in the liver was significantly increased by PM2.5, Ni, V, and Pb after exposure (p< 0.05), accompanied by liver inflammatory infiltration. Our results demonstrated that V in PM2.5 was associated with an increase in 8-isoprostane for all emissions and major local petrochemical emissions. In conclusion, V contributes to in vivo liver toxicity induced by PM2.5 in the vicinity of a petrochemical complex.