Neutrophils In Bronchoalveolar Lavage Fluid Research Articles

Lippia javanica (Zumbani) herbal tea infusion attenuates allergic airway inflammation via inhibition of Th2 cell activation and suppression of oxidative stress

BackgroundLippia javanica (lemon bush) is commonly used in the treatment of respiratory ailments, including asthma in southern African countries but there is no scientific evidence to support this claim. This study investigated the anti-inflammatory, antioxidant and anti-asthmatic effects of L. javanica using a rat model of asthma.MethodsA 5% w/v L. javanica tea infusion was prepared and characterised by liquid chromatography–mass spectrometer (LC-MS). Animals were intraperitoneally sensitized with ovalbumin (OVA) and subsequently challenged intranasal with OVA on day 15 except the control group. Animals were grouped (n = 5/group) for treatment: unsensitised control, sensitised control, sensitised + prednisolone and sensitised + L. javanica at 50 mg/kg/day and 100 mg/kg/day – equivalent to 1 and 2 cups of tea per day, respectively. After 2 weeks of treatment, bronchoalveolar lavage fluid (BALF) was collected for total and differential white blood cell (WBC) count. Nitric oxide (NO), lipid peroxidation and antioxidants were also assessed in BALF. Ovalbumin specific IgE antibody and inflammatory cytokines: IL-4, IL-5, IL-13 and TNF-alpha were measured in serum. Lung and muscle tissues were histological examined.ResultsL. javanica was rich in phenolic compounds. OVA sensitisation resulted in development of allergic asthma in rats. L. javanica treatment resulted in a reduction in total WBC count as well as eosinophils, lymphocytes and neutrophils in BALF. L. javanica inhibited Th2-mediated immune response, which was evident by a decrease in serum IgE and inflammatory cytokines: IL-4, IL-5, IL-13 and TNF-α. L. javanica treatment also reduced malondialdehyde (MDA) and NO, and increased superoxide dismutase, glutathione and total antioxidant capacity. Histology showed significant attenuation of lung infiltration of inflammatory cells, alveolar thickening, and bronchiole smooth muscle thickening.ConclusionL. javanica suppressed allergic airway inflammation by reducing Th2-mediated immune response and oxidative stress in OVA-sensitized rats which may be attributed to the presence of phenolic compound in the plant. This finding validates the traditional use of L. javanica in the treatment of respiratory disorders.

Progranulin attenuated asthma inflammation by inhibiting the expression of IL-6

Objective: To investigate the role and mechanism of progranulin (PGRN) in asthma. Methods: Control group and model group were set up in wild and IL-6 knockout (IL-6 ko) mice, respectively. For asthma model, mice were intraperitoneally sensitized with 100 μg OVA on days 0 and 7, followed by aerosol challenges with 5% OVA for 30 min per day from day 14 to 21, and mice were sacrificed 24 h after the last challenge. The mice in control group were treated in the same way with PBS. Bronchoalveolar lavage fluid (BALF) was collected for leukocytes count and differential count. The pathological changes of lung tissues were observed by H&E staining. The cytokines in lung homogenate, serum and BALF were detected by Q-PCR and ELISA. The in vitro model of asthma was induced by stimulating A549 or BEAS-2B cells with IL-13. Each group was replicated in three wells and four groups were designed: PBS group, IL-13 treatment group, IL-13 + rhPGRN treatment group, inhibitors of p38 phosphorylation (SB203508) treatment group. The cells or supernatant were collected after 0~48 h. PGRN and IL-6 levels were determined by Q-PCR and ELISA, the level of p38 phosphorylation was tested by Western blot (WB). Results: Compared with control group, PGRN levels were decreased in lung homogenate and BALF (P＜0.05), and PGRN presented a downtrend in serum, however, the level of IL-6 in BALF was increased in asthma mice (P＜0.01). In IL-6 ko asthma mice, compared with the wild asthma mice, leukocytes, especially neutrophils in BALF were decreased (P＜0.05), but PGRN was increased (P＜0.05), lung pathological damage was significantly alleviated. In vitro experiments, compared with PBS group, PGRN level was decreased (P＜0.05), IL-6 level was increased (P＜0.01), phosphorylation of p38 was activated in IL-13 treatment group. Compared with IL-13 treatment group, in IL-13 + PGRN treatment group, IL-6 level was decreased (P＜0.05); phosphorylation of p38 was inhibited (P＜0.05); and the production of IL-6 (P＜0.05) was decreased after treatment with inhibitor of p38 phosphorylation. Conclusion: PGRN inhibited the production of IL-6 by suppressing the p38 phosphorylation to alleviate asthmatic airway inflammation.

Bronchoalveolar lavage fluid cell subsets associate with the disease course in Löfgren's and non-Löfgren's sarcoidosis patients

BackgroundSarcoidosis is a multisystem granulomatous inflammatory disorder, that predominantly involves the lungs. Patients with Löfgren's syndrome (LS) are characterized by acute onset and usually have the HLA-DRB1*03 (DR3positive) allele and a good prognosis. Non-LS patients are usually DR3negative and are more likely to develop chronic disease. The study aimed to identify bronchoalveolar lavage fluid (BALF) cells that could associate with disease severity (reduced pulmonary function tests (PFTs), advanced chest radiographs, need for treatment) and/or chronicity (duration >2 years) in newly diagnosed LS and non-LS patients, respectively. MethodsWe retrospectively included data from 955 non-LS patients, 477 LS patients, and 295 healthy controls (HC) in this study. Intra-group comparison of patients with resolving versus chronic disease was performed in LS and non-LS, respectively. Non-LS patients were divided into two subgroups according to the binary BALF cell concentrations for intra-group comparison (i.e. higher or lower than the 95th percentile of the BALF cells references in healthy individuals). ResultsLS patients with a non-resolving disease course had higher BALF lymphocytes, neutrophils, and eosinophils than LS with a favourable outcome. In non-LS subjects increased BALF of the same cells and in addition also of basophils and mast cells were more likely associated with more severe disease course. ConclusionIncreased BALF cells display prognostic significance in sarcoidosis. Certain BALF profiles should promote the clinician to monitor these patients more closely as they may associate non-resolving disease, in turn, resulting in future irreversible functional impairment.

Loss of IL-33 enhances elastase-induced and cigarette smoke extract-induced emphysema in mice

BackgroundIL-33, which is known to induce type 2 immune responses via group 2 innate lymphoid cells, has been reported to contribute to neutrophilic airway inflammation in chronic obstructive pulmonary disease. However, its role in the pathogenesis of emphysema remains unclear.MethodsWe determined the role of interleukin (IL)-33 in the development of emphysema using porcine pancreas elastase (PPE) and cigarette smoke extract (CSE) in mice. First, IL-33−/− mice and wild-type (WT) mice were given PPE intratracheally. The numbers of inflammatory cells, and the levels of cytokines and chemokines in the bronchoalveolar lavage (BAL) fluid and lung homogenates, were analyzed; quantitative morphometry of lung sections was also performed. Second, mice received CSE by intratracheal instillation. Quantitative morphometry of lung sections was then performed again.ResultsIntratracheal instillation of PPE induced emphysematous changes and increased IL-33 levels in the lungs. Compared to WT mice, IL-33−/− mice showed significantly greater PPE-induced emphysematous changes. No differences were observed between IL-33−/− and WT mice in the numbers of macrophages or neutrophils in BAL fluid. The levels of hepatocyte growth factor were lower in the BAL fluid of PPE-treated IL-33−/− mice than WT mice. IL-33−/− mice also showed significantly greater emphysematous changes in the lungs, compared to WT mice, following intratracheal instillation of CSE.ConclusionThese observations suggest that loss of IL-33 promotes the development of emphysema and may be potentially harmful to patients with COPD.

The effect of lower airway inflammation on inflammatory cytokine gene expression in bronchoalveolar lavage fluid and whole blood in racing Thoroughbreds

BackgroundImmunological mechanisms involved in the pathogenesis of mild to moderate equine asthma (MEA) are not completely understood. There are limited data on bronchoalveolar lavage fluid (BALF) and blood inflammatory cytokine profiles in racehorses with MEA, and the effect of racing on inflammatory cytokines is unknown. Hypothesis/objectivesWe hypothesized that inflammatory cytokine gene expression in BALF and resting blood would be higher in racehorses with lower airway inflammation compared to healthy controls, and that gene expression in blood collected immediately post-race would be increased compared to resting blood in racehorses with lower airway inflammation. Animals38 racing Thoroughbreds (samples: 30 resting blood, 22 post-race BALF, 41 post-race blood). MethodsProspective observational study. Inflammatory cytokine gene expression was determined in resting blood, post-race BALF and post-race blood from racehorses with lower airway inflammation and controls. ResultsLower airway inflammation was diagnosed in 79 % of racehorses (23 % neutrophilic, 67 % mastocytic, and 10 % mixed). There was no difference in gene expression in BALF or resting blood between racehorses with lower airway inflammation and controls. IL-8 gene expression was higher in post-race blood compared to resting peripheral blood, regardless of disease (p = 0052). BALF neutrophil proportions increased with increasing IL-1β gene expression in all sample types (p = 0.0025). BALF mast cell proportions increased with increasing TNF-α gene expression in post-race blood (p = 0.015). Conclusions and clinical importanceLower airway inflammation was common in a population of racehorses without respiratory signs or exercise intolerance. Exercise alone increased peripheral blood IL-8 gene expression. Inflammatory cytokine gene expression was not increased in BALF or resting blood in horses with subclinical lower airway inflammation, precluding its diagnostic utility in clinical practice.

SUL-151 Decreases Airway Neutrophilia as a Prophylactic and Therapeutic Treatment in Mice after Cigarette Smoke Exposure.

Chronic obstructive pulmonary disease (COPD) caused by cigarette smoke (CS) is featured by oxidative stress and chronic inflammation. Due to the poor efficacy of standard glucocorticoid therapy, new treatments are required. Here, we investigated whether the novel compound SUL-151 with mitoprotective properties can be used as a prophylactic and therapeutic treatment in a murine CS-induced inflammation model. SUL-151 (4 mg/kg), budesonide (500 μg/kg), or vehicle were administered via oropharyngeal instillation in this prophylactic and therapeutic treatment setting. The number of immune cells was determined in the bronchoalveolar lavage fluid (BALF). Oxidative stress response, mitochondrial adenosine triphosphate (ATP) production, and mitophagy-related proteins were measured in lung homogenates. SUL-151 significantly decreased more than 70% and 50% of CS-induced neutrophils in BALF after prophylactic and therapeutic administration, while budesonide showed no significant reduction in neutrophils. Moreover, SUL-151 prevented the CS-induced decrease in ATP and mitochondrial mtDNA and an increase in putative protein kinase 1 expression in the lung homogenates. The concentration of SUL-151 was significantly correlated with malondialdehyde level and radical scavenging activity in the lungs. SUL-151 inhibited the increased pulmonary inflammation and mitochondrial dysfunction in this CS-induced inflammation model, which implied that SUL-151 might be a promising candidate for COPD treatment.

Nandina domestica Leaf Extracts Ameliorate Lung Injury by Decreasing Inflammatory Cytokine Levels and Neutrophil Accumulation in Mice Injected with Fine Dust

Exposure to dust particles can lead to respiratory problems, diseases, and even death. In this study, we examined the effects of Nandina domestica (ND) on bronchoalveolar lavage fluid (BALF) and lung of intra-nazal-trachea (INT) injected mice with fine dust mixture of coal, flay ash and diesel exhausted particle (CFD). ND significantly reduced the counts of neutrophils in BALF and histological changes in lung tissues including infiltration of inflammatory cells and goblet cell hyperplasia with lowering lung injury score and periodic acid Schiff (PAS) score. In addition, the elevated inflammatory cytokines such as interleukin (IL)-17 and tumor necrosis factor-α (TNF-α) in BALF, macrophage inflammatory protein-2 (MIP-2) and C-X-C motif chemokine 1 (CXCL-1) significantly decreased to a greater extent by ND treatment. Furthermore, ND significantly decreased levels of asymmetric dimethyl-arginine (ADMA) and symmetric dimethyl-arginine (SDMA) elevated by CFD in BALF. Therefore, these data suggest that ND ameliorated lung injury by suppressing inflammatory cytokines as well as by reducing the accumulation of neutrophils.

Nandina domestica Leaf Extracts Ameliorate Lung Injury by Decreasing Inflammatory Cytokine Levels and Neutrophil Accumulation in Mice Injected with Fine Dust

Exposure to dust particles can lead to respiratory problems, diseases, and even death. In this study, we examined the effects of Nandina domestica (ND) on bronchoalveolar lavage fluid (BALF) and lung of intra-nazal-trachea (INT) injected mice with fine dust mixture of coal, flay ash and diesel exhausted particle (CFD). ND significantly reduced the counts of neutrophils in BALF and histological changes in lung tissues including infiltration of inflammatory cells and goblet cell hyperplasia with lowering lung injury score and periodic acid Schiff (PAS) score. In addition, the elevated inflammatory cytokines such as interleukin (IL)-17 and tumor necrosis factor-α (TNF-α) in BALF, macrophage inflammatory protein-2 (MIP-2) and C-X-C motif chemokine 1 (CXCL-1) significantly decreased to a greater extent by ND treatment. Furthermore, ND significantly decreased levels of asymmetric dimethyl-arginine (ADMA) and symmetric dimethyl-arginine (SDMA) elevated by CFD in BALF. Therefore, these data suggest that ND ameliorated lung injury by suppressing inflammatory cytokines as well as by reducing the accumulation of neutrophils.

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.

Houttuynia Cordata Essential Oil Mitigates Airway Remodeling in Asthma by Rectifying IP3R‐mediated Ca 2+ Disruption

Asthma is a common chronic airway inflammatory disease that leads to airway narrowing during the attacks. Beta-adrenergic agonists provide temporary relaxation of airway during asthma attacks. While long-term usage of corticosteroids could improve asthma but it may bring adverse effects to the patients. Thus, an alternative or supplementary therapeutic approach is urgently needed. One potential therapeutic approach is targeting intracellular calcium (Ca2+) signaling in airway smooth muscle cells. It is reported that these intracellular calcium signals played an essential role in contractility regulations and airway remodeling. Meanwhile, Houttuynia Cordata (HC) is a frequently used traditional Chinese medicinal herb in asthma treatment due to its anti-allergic and anti-inflammatory properties. Thus, we hypothesized that HC may modulate intracellular Ca2+ signals and could be a promising candidate to relieve asthmatic airway remodeling. In this study, we investigated the role of intracellular Ca2+ channels in airway remodeling and elucidated the therapeutic mechanism of HC, particularly its effect on intracellular Ca2+ channels. We used interleukin 4 and 13 treated human bronchial smooth muscle cells (hBSMC), and house-dust mite sensitized BALB/C mice as an acute asthmatic cell and animal models. We found that the mRNA and protein levels of IP3R1, TPC2, and SERCA2A were elevated in asthmatic hBSMC, which are reconcilable to the observation in the asthmatic mouse model. Apart from enhanced Ca2+ channels expression, we found that the IP3-mediated calcium release in asthmatic hBSMC was also increased, suggesting IP3R plays an indispensable pathogenic role in asthma development. Intriguingly, pre-treatment of HC essential oil in cells candose-dependently reduced ATP-triggered Ca2+ release. To evaluate the in vivo effect of HC essential oil, we nebulized the HC essential oil to pre-treat the house-dust mite sensitized asthmatic mouse. Our result shows that HC essential oil pre-treatment significantly reduced neutrophil and eosinophil counts in bronchoalveolar lavage fluid. In addition, asthmatic airway remodeling indicators such as increased airway thickness, airway smooth muscle cells hyperplasia, hyperresponsiveness, and airway ring contractions were also mitigated by the HC pre-treatment. These results suggest HC essential oil can prevent airway remodeling and asthma development in asthmatic mouse model. Taken together, our current data suggested that HC essential oil mitigates asthma development by rectifying the disrupted Ca2+ signaling and prevents airway narrowing. HC may serve as a potential alternative treatment choice to prevent or treat asthma.

Deficiency of voltage-gated proton channel Hv1 aggravates ovalbumin-induced allergic lung asthma in mice

Asthma is a chronic airway inflammation that caused by many factors. The voltage-gated proton channel Hv1 has been proposed to extrude excessive protons produced by NADPH oxidase (NOX) from cytosol to maintain its activity during respiratory bursts. Here, we showed that loss of Hv1 aggravates ovalbumin (OVA)-induced allergic lung asthma in mice. The numbers of total cells, eosinophils and neutrophils in bronchoalveolar lavage fluid (BALF) of Hv1-deficiency (KO) mice are obviously increased after OVA challenge compared with that of wild-type (WT) mice. Histopathological staining reveals that Hv1-deficiency aggravates OVA-induced inflammatory cell infiltration and goblet cell hyperplasia in lung tissues. The expression of IL-4, IL-5 and IL-13 are markedly increased in lung tissues of OVA-challenged KO mice compared with that of WT mice. Furthermore, the expression levels of NOX2, NOX4 and DUOX1 are dramatically increased, while the expression levels of SOD2 and catalase are significantly reduced in lung tissues of OVA-challenged KO mice compared with that of WT mice. The production of ROS in lung tissues of KO mice is significantly higher than that of WT mice after OVA challenge. Our data suggest that Hv1-deficiency might aggravate the development of allergic asthma through increasing ROS production.

NaHS Alleviated Cell Apoptosis and Mitochondrial Dysfunction in Remote Lung Tissue after Renal Ischemia and Reperfusion via Nrf2 Activation-Mediated NLRP3 Pathway Inhibition.

Objective Acute kidney injury (AKI) is a common and severe complication in critically ill patients, often caused by renal ischemia-reperfusion (RIR). Previous studies have confirmed that lung injury, rather than renal injury, is one of the leading causes of AKI-induced death. The pathophysiological mechanisms of acute lung injury (ALI) resulting from AKI are very complex and remain unclear. In the present study, we aimed to explore the protective effects and potential mechanism of sodium hydrosulfide (NaHS) on lung injury in RIR mice. Methods The RIR model was established in wild-type and Nrf2−/− mice. Different groups of mice were treated with NaHS and MCC950. Lung tissues were harvested to detect lung injury, mitochondrial function, cell apoptosis, the NLRP3 inflammasome, and Nrf2 pathway-related molecules. Results RIR led to a deterioration in lung histology, the wet/dry weight ratio, PaO2/FiO2, and mitochondrial function, in addition to stimulating the activation of the NLRP3 and Nrf2 pathways. MCC950 alleviated mitochondrial dysfunction, lung apoptosis, and histology injury in the lungs after RIR. NaHS treatment markedly improved the lung histological scores, the wet/dry weight ratio, bronchoalveolar lavage fluid (BALF) cell counts, BALF neutrophil counts, BALF neutrophil elastase activity, BALF protein concentration, PaO2/FiO2, mitochondrial morphology, the red/green fluorescence intensity that indicates changes in mitochondrial membrane potential, respiratory control rate (RCR), ATP, reactive oxygen species (ROS) release, and cell apoptosis via Nrf2-mediated NLRP3 pathway inhibition. Conclusion NaHS protected against RIR-induced lung injury, mitochondrial dysfunction, and inflammation, which is associated with Nrf2 activation-mediated NLRP3 pathway inhibition.

Equine respiratory viruses, airway inflammation and performance in thoroughbred racehorses

Equine asthma is a common cause of poor performance in racehorses but it is unclear if respiratory viruses contribute to its etiology. The objective of the study was to determine if respiratory viruses were associated with clinical signs and bronchoalveolar lavage fluid (BALF) cytology in Thoroughbred racehorses. Equine herpesviruses (EHV-1, 2, 4, 5) and equine rhinitis A and B viruses (ERBV, ERAV) genomes were quantified by qPCR in nasopharyngeal, tracheal, and BALF samples collected after racing. The relationships between virus detection and load and clinical signs, performance, BALF cytology, and environmental exposures were examined with generalized linear mixed models. Ninety-two samples were collected from 31 horses. EHV-1 and ERAV were not found; EHV-4 was detected in only one sample. EHV-2, EHV-5 and ERBV were more likely to be detected in upper airway samples than in BALF (P < 0.0001). Neither respiratory virus detection nor load was associated with clinical signs or performance. Nasopharyngeal detection and load of ERBV and tracheal detection and load of EHV-5 were associated with increased proportions of neutrophils in BALF (P < 0.003). However, nasopharyngeal detection and load of EHV-5 was not (P = 0.11). Nasopharyngeal detection and load of EHV-2 were associated with decreased BALF mast cell proportions. Respirable dust exposures were significantly higher in horses with detection of ERBV when compared to horses with no detectable ERBV (P < 0.001). Our results suggest that ERBV, EHV-2 and EHV-5 are commonly present in upper airways of healthy racehorses; however, the role they play in the etiology of equine asthma remains unclear.

Effects of Bedding Material on Equine Lower Airway Inflammation: A Crossover Study Comparing Peat and Wood Shavings.

Bedding materials affect stable air hygiene, and thus the development and exacerbation of equine asthma. There is limited knowledge concerning the effects of different types of bedding material on equine lower airway inflammation. The objective of our study was to investigate the effects of bedding materials on respiratory signs, tracheal mucus score, and lower airway cytology in healthy adult horses. The study design was a prospective controlled study, and the subjects were healthy adult riding school horses (n = 32) from a single stable. Wood shavings were compared to peat, which was used as a reference bedding material. Lower airway endoscopy and sampling (tracheal wash and bronchoalveolar lavage fluid) for cytological examination were performed after each 35-day bedding period. No difference between bedding periods was observed in the respiratory rate or tracheal mucus score. Tracheal wash neutrophil percentage with the wood shavings was higher compared to the previous (P = 0.040) or following (P = 0.0045) peat period. Bronchoalveolar lavage fluid neutrophil percentage with the wood shavings was higher compared to the following peat period (P < 0.001). We conclude that, between the two bedding materials used in this study, peat caused less neutrophilic lower airway inflammation in horses. The information gained from this study may assist veterinarians and horse owners in selecting bedding materials, especially for horses suffering from equine asthma.

Astragalin Inhibits Cigarette Smoke-Induced Pulmonary Thrombosis and Alveolar Inflammation and Disrupts PAR Activation and Oxidative Stress-Responsive MAPK-Signaling.

Epidemiological evidence shows that smoking causes a thrombophilic milieu that may play a role in the pathophysiology of chronic obstructive pulmonary disease (COPD) as well as pulmonary thromboembolism. The increased nicotine level induces a prothrombotic status and abnormal blood coagulation in smokers. Since several anticoagulants increase bleeding risk, alternative therapies need to be identified to protect against thrombosis without affecting hemostasis. Astragalin is a flavonoid present in persimmon leaves and green tea seeds and exhibits diverse activities of antioxidant and anti-inflammation. The current study investigated that astragalin attenuated smoking-induced pulmonary thrombosis and alveolar inflammation. In addition, it was explored that molecular links between thrombosis and inflammation entailed protease-activated receptor (PAR) activation and oxidative stress-responsive mitogen-activated protein kinase (MAPK)-signaling. BALB/c mice were orally administrated with 10–20 mg/kg astragalin and exposed to cigarette smoke for 8 weeks. For the in vitro study, 10 U/mL thrombin was added to alveolar epithelial A549 cells in the presence of 1–20 µM astragalin. The cigarette smoking-induced the expression of PAR-1 and PAR-2 in lung tissues, which was attenuated by the administration of ≥10 mg/kg astragalin. The oral supplementation of ≥10 mg/kg astragalin to cigarette smoke-challenged mice attenuated the protein induction of urokinase plasminogen activator, plasminogen activator inhibitor-1and tissue factor, and instead enhanced the induction of tissue plasminogen activator in lung tissues. The astragalin treatment alleviated cigarette smoke-induced lung emphysema and pulmonary thrombosis. Astragalin caused lymphocytosis and neutrophilia in bronchoalveolar lavage fluid due to cigarette smoke but curtailed infiltration of neutrophils and macrophages in airways. Furthermore, this compound retarded thrombin-induced activation of PAR proteins and expression of inflammatory mediators in alveolar cells. Treating astragalin interrupted PAR proteins-activated reactive oxygen species production and MAPK signaling leading to alveolar inflammation. Accordingly, astragalin may interrupt the smoking-induced oxidative stress–MAPK signaling–inflammation axis via disconnection between alveolar PAR activation and pulmonary thromboembolism.

Flow Cytometric Analysis of Systemic and Airway Neutrophil Maturation and Activation in Lung Transplant Patients

Purpose The neutrophil is the most abundant blood leukocyte in humans and the first cell involved in an immune response like infection, rejection or injury of the lung. Neutrophils were long considered as simple short-lived cells. However, several distinct neutrophil subsets, maturation and activation stages with functional heterogeneity are identified. This spiked our interest to perform an in-depth neutrophil characterization, focusing on their maturation and activation characteristics, in broncho-alveolar lavage fluid (BALF) and peripheral blood of lung transplantation (LTx) patients who received standard immunosuppression. Methods BALF and blood neutrophils were microscopically evaluated and identified by multicolor flow cytometry (CD16+/CD66b+) to examine expression of maturation/activation markers and adhesion molecules (CD10, CD62L, CD11b, CD11c, CD15, HLA-DR) and chemoattractant receptors (CXCR1-4, CCR1-3, FPR1-2, C5aR and BLTR1) within healthy control subjects (n = 11), stable LTx patients (n = 8-14), and LTx patients with infection (n = 9) or chronic lung allograft dysfunction (n = 2-6). Results LTx patients had a significantly higher (p Conclusion We demonstrated a compromised maturation of neutrophils in peripheral blood and BALF of LTx patients versus healthy controls. In addition, we observed a more activated status of BALF neutrophils with a surprisingly increased expression of CXCR3 on their membrane in stable LTx patients.

Baicalin regulates the development of pediatric asthma via upregulating microRNA-103 and mediating the TLR4/NF-κB pathway

Pediatric asthma seriously endangers the well-being and health of children worldwide. Baicalin (BA) protects against diverse disorders, including asthma. Therefore, this study explored the mechanism of BA in pediatric asthma. The ovalbumin (OVA)-induced asthmatic mouse model was established to evaluate BA efficacy from aspects of oxidative stress, inflammation, blood cells in bronchoalveolar lavage fluid (BALF) and collagen deposition. Differentially expressed microRNAs (miRs) in BA-treated mice were analyzed. Effects of BA on PDGF-BB-induced smooth muscle cells (SMCs) were assessed. miR downstream mRNA and the related pathway were predicted and verified, and their effects on asthmatic mice were evaluated. BA effectively reversed OVA-induced oxidative stress and inflammation, as well as decreased the number of total cells, eosinophils and neutrophils in BALF, and collagen deposition. miR-103 was significantly upregulated after BA treatment. BA inhibited the abnormal proliferation of PDGF-BB-induced SMCs, which was prevented by miR-103 knockdown. miR-103 targeted TLR4 and regulated the extent of NF-κB phosphorylation. In vivo, miR-103 inhibition weakened the alleviating effects of BA on asthma, which was then reversed after silencing of TLR4. We highlighted that BA has the potency to halt the pediatric asthma progression via miR-103 upregulation and the TLR4/NF-κB axis inhibition.

Dust exposure and pulmonary inflammation in Standardbred racehorses fed dry hay or haylage: A pilot study

Respirable dust exposure is linked to airway inflammation in racehorses. Feeding haylage may reduce dust exposure by 60–70%. The objective of this study was to compare dust exposure, airway cytology, and inflammatory cytokine concentrations between horses fed haylage or hay over 6 weeks while in training. Seven healthy Standardbred horses were randomly assigned to be fed alfalfa hay (n = 3) or grass-alfalfa mix haylage (n = 4) for six weeks while training on a treadmill. Dust exposure was measured gravimetrically at the breathing zone. Endotoxin and β-glucan concentrations in respirable dust were measured. Bronchoalveolar lavage fluid (BALF) cytology was determined at baseline and after 2, 4, and 6 weeks. Cytokine concentrations (interferon-γ, tumor necrosis factor-α, and interleukin-4) were measured in BALF at baseline and week 6. The effect of forage on exposure, airway cytology and cytokines were evaluated using generalized linear mixed models.Respirable dust and β-glucan exposures were lower in horses fed haylage than hay (0.02 ± 0.001 mg/m3 vs. 0.06 ± 0.01 mg/m3; P = 0.03, and 69 ± 18 pg/m3 vs. 160 ± 21 pg/m3; P = 0.02, respectively). In horses eating haylage, BALF neutrophil proportion decreased between baseline (2.2 ± 0.5%), week 2 (0.8 ± 0.3%; P = 0.01) and week 6 (0.7 ± 0.2%; P = 0.03). By week 6, horses fed haylage had lower BALF neutrophilia than horses fed hay (4.0 ± 0.7 %; P = 0.0004). Interleukin-4 concentration in BALF was higher at week 6 (14.4 ± 4.6 pg/mL) in horses fed hay compared to baseline (2.9 ± 4.6 pg/mL; P = 0.007). In conclusion, feeding haylage instead of hay to horses in training can reduce exposure to respirable irritants and mitigate airway neutrophilia.

MNK as a potential pharmacological target for suppressing LPS-induced acute lung injury in mice

Acute lung injury (ALI) or its more severe form, known as acute respiratory distress syndrome (ARDS), is characterized by an initial exudative phase, expression of proinflammatory mediators, activation of inflammatory leukocytes, and impairment of the lung endothelium and epithelium. Despite numerous, novel therapeutic strategies have been developed regarding the pathophysiology of ALI, current treatment is mainly supportive, as specific therapies have not been established in the past few decades. The MAP kinase-interacting kinases (MNK1 and MNK2) are serine threonine kinases which are activated by mitogen-activated protein kinases (MAPKs), regulate protein synthesis by phosphroylating eukaryotic translation initiation factor 4E (eIF4E). Although studies have shown that MAPKs pathway is involved in anti-inflammatory and preventing tissue injury processes, the role of MNKs in ALI has, until now, remained relatively unexplored. Here, we investigated whether partial inhibition of MAPKs pathway by targeting MNKs was effective in the prevention and treatment of ALI. C57BL6 mice were pretreated with MNK1 and MNK2 inhibitor (CGP57380, 30 mg/kg) for 30 min and then challenged with 5 mg/kg LPS for 6 h. The results showed that pretreatment with CGP57380 not only significantly attenuated LPS-induced lung wet/dry ratio, as well as protein content, total cells and neutrophils in bronchoalveolar lavage fluid (BALF), but also decreased the production of pro-inflammatory mediators such as interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-α) and keratinocyte-derived chemoattractant (KC). In addition, CGP57380 was observed to significantly suppress LPS-stimulated phosphorylation of eIF4E and MAPKs in the mouse bone marrow-derived macrophages (BMDMs). The involvement of MNK2 in lung injury was further evident by MNK2 knockout mice. MNK2 deficiency resulted in the attenuated lung histopathological changes, as also reflected by reductions in neutrophil counts, and the less LPS-induced the production of IL-6, TNF-α and KC in mouse BALF. Taken together, these findings demonstrated for the first time that MNK inhibition could effectively reduce the LPS-induced ALI in mice, suggesting a novel and potential application for MNK-based therapy to treat this serious disease.

Inhibiting the ROCK Pathway Ameliorates Acute Lung Injury in Mice following Myocardial Ischemia/reperfusion

ABSTRACT To clarify the role of Y-27632, a selective inhibitor of Rho-associated coiled-coil forming protein kinase (ROCK), in acute lung injury (ALI) induced by myocardial ischemia/reperfusion (I/R). Mice were randomized into Sham, I/R, and Y-27632 (10, 20 or 30 mg/kg) + I/R groups, and hemodynamics, infarcted area, the protein concentration, neutrophils in bronchoalveolar lavage fluid (BALF), malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) levels were assessed. Pathological changes were evaluated by hematoxylin-eosin (HE) staining; protein and gene expression were measured by Western blotting, enzyme-linked immunosorbent assay (ELISA), immunohistochemistry and quantitative real-time reverse-transcription polymerase chain reaction (qRT-PCR); and apoptosis was assessed by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) staining. ROCK1 and ROCK2 expression was up-regulated in lung tissues of I/R mice compared to sham mice. Y-27632 decreased the protein concentration and the neutrophils in BALF in I/R mice, improved hemodynamics and reduced infarct size (IS)/area at risk (AAR) ratio. In addition, pathological changes in lung tissues of Y-27632-treated mice were mitigated, and these alterations were accompanied by decreases in MDA levels in lung tissues and increases in SOD and GSH-Px levels. Moreover, in I/R group, the number of apoptotic cells in lung tissue was higher than that in sham group, and p53, Caspase-3 and Bax expression was up-regulated; however, following treatment with Y-27632 (10, 20 and 30 mg/kg), these changes were reversed. Inhibition of ROCK pathway by Y-27632 ameliorated ALI in myocardial I/R mice by mitigating oxidative stress, inflammation and cell apoptosis.