Protein Concentration In Lavage Fluid Research Articles

Metformin relieves acute respiratory distress syndrome by reducing miR-138 expression.

To investigate whether metformin can relieve acute respiratory distress syndrome (ARDS). Its potential mechanism was also explored. The ARDS model was established by injecting LPS into mice that received metformin in advance and the mice in the control group. Pulmonary edema was detected by W/D ratios (wet-to-dry weight ratios), and the vascular exudation was reflected by the protein content and cell number of alveolar lavage fluid. Meanwhile, MPO (myeloperoxidase) activity assay was performed to analyze the neutrophil aggregation. The expression of inflammatory cytokines, including TNF-α, IL-1β, IL-6, and IL-17, were detected by enzyme-linked immunosorbent assay (ELISA). This series of experiments reflected the alleviation effect of metformin on ARDS. To further study the mechanism, we cultured alveolar macrophages (NR8383) in vitro and treated them with LPS and metformin. Western blot was used to detect the phosphorylation levels of p38, ERK, NF-kB, and SIRT1 expression level. Bioinformatics method was then used to predict the binding of miR-138 to SIRT1. The mRNA and protein expression of SIRT1 was detected in NR8383 cells transfected with miR-138 inhibitor. The dual luciferase gene reporter assay was used to detect the relative luciferase activities of miR-138 and SIRT1. Pulmonary edema, vascular exudation, and neutrophil accumulation were observed in the ARDS model mice, and the levels of inflammatory cytokines including TNF-α, IL-1b, IL-6, and IL-17 were significantly increased. After metformin treatment, these pulmonic damage indicators were found to be partially reversed. At the same time, metformin could significantly reduce LPS-induced death. After NR8383 was treated with metformin and LPS, the expression of SIRT1 was higher than that of LPS treatment alone, but the expression of p-p38, p-ERK, and p-NF-κB was significantly decreased. After the addition of metformin in NR8383 after LPS treatment, the expression level of miR-138-5p was significantly decreased, and miR-138-5p was confirmed to target SIRT1 and regulate its expression. Metformin could reduce LPS-induced pulmonic injury and increase expression of inflammatory factors. A possible mechanism might be that metformin-induced low expression of mir-138-5p could target SIRT1 to increase its expression and suppress the MAPK pathway, thus alleviating ARDS.

Dexmedetomidine Protects Against Traumatic Brain Injury-Induced Acute Lung Injury in Mice.

BackgroundTraumatic brain injury (TBI) leads to acute lung injury (ALI), in which the inflammatory response plays an important role in its pathophysiology. Recent studies suggest that dexmedetomidine (Dex) plays a protective role in acute inflammatory diseases. However, whether Dex has a protective effect on TBI-induced ALI is not clear. The aim of this study was to investigate the effect of Dex on TBI-induced ALI in mice.Material/MethodsMice were randomly divided into 5 groups: 1) sham group; 2) TBI group; 3) TBI+Dex group; 4) TBI+atipamezole (Atip) group; and 5) TBI+Dex+Atip group. Dex (50 μg/kg) was intraperitoneal injected immediately after TBI. The α2 adrenergic antagonist Atip (250 μg/kg) was intraperitoneal injected 15 minutes prior to Dex treatment. Then 24 hours later, the protein concentration in the bronchoalveolar lavage fluid (BALF), lung wet to dry weight ratio, hematoxylin and eosin (H&E) staining of lungs, the level of high-mobility group box protein 1(HMGB1) in serum, and the receptor for advanced glycation end products (RAGE) expression in lung were detected.ResultsDex ameliorated the score of lung histological examination, as well as the severity of pulmonary edema and permeability. Moreover, Dex was observed to significantly suppress the expression of HMGBI and RAGE. However, the protective effects of Dex were partially reversed by the administration of Atip.ConclusionsDex may protect against TBI-induced ALI via the HMGB1-RAGE signal pathway, and this protective effect is partly dependent on its α2 adrenoceptor agonist action.

Tumor necrosis factor-alpha protein concentrations in bronchoalveolar lavage fluid from healthy horses and horses with severe equine asthma

The objective of this study was to determine if TNF-α protein concentration differs in bronchoalveolar lavage fluid (BALF) obtained from healthy horses, horses with naturally occurring exacerbations of severe equine asthma and horses in remission from severe equine asthma. Tumor necrosis factor-alpha (TNF- α) protein concentrations were determined in BALF by commercial equine ELISA. Horses with naturally occurring exacerbation of severe equine asthma were found to have significantly lower BALF TNF-α protein concentrations than healthy horses (p = 0.0026). There was no significant difference in BALF TNF-α concentration between horses in exacerbation and remission from disease, but there was a decrease in median TNF-α concentration between healthy horses and horses with clinical exacerbation of severe equine asthma. These findings suggest, that similar to human asthma, the role of TNF-α in chronic lower airway inflammation of horses may differ between disease phenotypes and disease state. Furthermore, the method with which TNF-α is measured (mRNA expression vs. protein concentration) may affect results when studying the role of TNF-α in horses with severe equine asthma.

Angiotensin II type 2 receptor agonist Compound 21 attenuates pulmonary inflammation in a model of acute lung injury.

PurposeAlthough the role of the angiotensin II type 2 (AT2) receptor in acute lung injury is not yet completely understood, a protective role of this receptor subtype has been suggested. We hypothesized that, in a rodent model of acute lung injury, stimulation of the AT2 receptor with the direct agonist Compound 21 (C21) might have a beneficial effect on pulmonary inflammation and might improve pulmonary gas exchange.Materials and methodsMale adult rats were divided into a treatment group that received pulmonary lavage followed by mechanical ventilation (LAV, n=9), a group receiving pulmonary lavage, mechanical ventilation, and direct stimulation of the AT2 receptor with C21 (LAV+C21, n=9), and a control group that received mechanical ventilation only (control, n=9). Arterial blood gas analysis was performed every 30 min throughout the 240-min observation period. Lung tissue and plasma samples were obtained at 240 min after the start of mechanical ventilation. Protein content and surface activity of bronchoalveolar lavage fluid were assessed and the wet/dry-weight ratio of lungs was determined. Transcriptional and translational regulation of pro- and antiinflammatory cytokines IL-1β, tumor necrosis factor-alpha, IL-6, IL-10, and IL-4 was determined in lungs and in plasma.ResultsPulmonary lavage led to a significant impairment of gas exchange, the formation of lung edema, and the induction of pulmonary inflammation. Protein content of lavage fluid was increased and contained washed-out surfactant. Direct AT2 receptor stimulation with C21 led to a significant inhibition of tumor necrosis factor-alpha and IL-6 expressions in the lungs, whereas the expressions of IL-1, IL-10, and IL-4 remained unchanged. During the 240-min observation period, AT2 receptor stimulation did not improve pulmonary gas exchange or lung edema.ConclusionIn this rodent model of acute lung injury after repeated pulmonary lavage, AT2 receptor stimulation attenuates pulmonary inflammation but does not improve gas exchange.

Luteolin attenuates acute lung injury in experimental mouse model of sepsis

The present investigation was undertaken to assess the result of pretreatment of luteolin in sepsis-induced acute lung injury in mice and its mechanism of action. Luteolin was administered intraperitoneally one hour before caecal ligation and puncture (CLP) surgery in mice. Acute lung injury was assessed by estimation of different parameters like lung edema, protein content, cytokines level, oxidative stress, inducible nitric oxide synthase (iNOS), intercellular adhesion molecule (ICAM)-1 expression and histopathology. Pretreatment of mice with luteolin showed decrease lung edema and protein content in tissue and bronchoalveolar lavage fluid (BALF). However, mice pretreated with luteolin showed reduction (p = 0.92) in blood and lung tissue bacterial counts however it was non significant. Further, luteolin showed significant reduction in interleukin (IL)-6 and IL-1β in lung tissue which are the proinflammatory cytokines. However, plasma IL-1β and tissue tumor necrosis factor (TNF)-α level decrease (p = 0.24; p = 0.19) with this pretreatment. Further, ICAM-1 mRNA expression and nuclear factor (NF)-kappa B protein expression were significantly (p < 0.01) decreased in luteolin pretreated septic mice. The lung iNOS level, iNOS mRNA and protein expressions were markedly (p = 0.25; p = 0.50; p = 0.06) altered with luteolin pretreatment, respectively. Also, significant reduction in lipid peroxidation and increase in the activity of antioxidant enzymes like superoxide dismutase (SOD) and catalase was noted with luteolin pretreatment. However, luteolin did not alter (p = 0.36) the non enzymatic antioxidant GSH activity in septic mice. Histopathology of lung tissue showed reduction in lung injury with the luteolin pretreatment in septic mice. The study suggests that luteolin showed attenuation in sepsis-induced acute lung injury in mice through suppression in ICAM-1, NF-kappa B, oxidative stress and partially iNOS pathways.

Preservation with α1-antitrypsin improves primary graft function of murine lung transplants.

BackgroundVascular damage and primary graft dysfunction increase with prolonged preservation times of transplanted donor lungs. Hence, storage and conservation of donated lungs in protein-free, dextran-containing electrolyte solutions, like Perfadex, is limited to about 6 hours. We hypothesized that transplanted lungs are protected against neutrophil-mediated proteolytic damage by adding α1-anti-trypsin (AAT), a highly abundant human plasma proteinase inhibitor, to Perfadex.MethodsA realistic clinically oriented murine model of lung transplantation was used to simulate the ischemia–reperfusion process. Lung grafts were stored at 4°C in Perfadex solution supplemented with AAT or an AAT mutant devoid of elastase-inhibiting activity for 18 hours. We examined wild-type and proteinase 3/neutrophil elastase (PR3/NE) double-deficient mice as graft recipients. Gas exchange function and infiltrating neutrophils of the transplanted lung, as well as protein content and neutrophil numbers in the bronchoalveolar lavage fluid, were determined.ResultsAAT as a supplement to Perfadex reduced the extent of primary graft dysfunction and early neutrophil responses after extended storage for 18 hours at 4°C and 4-hour reperfusion in the recipients. Double-knockout recipients that lack elastase-like activities in neutrophils were also protected from early reperfusion injury, but not lung grafts that were perfused with a reactive center mutant of AAT devoid of elastase-inhibiting activity.ConclusionsPR3 and NE, the principal targets of AAT, are major triggers of post-ischemic reperfusion damage. Their effective inhibition in the graft and recipient is a promising strategy for organ usage after storage for >6 hours.

Vaspin protects against lipopolysaccharide-induced acute respiratory distress syndrome in mice by inhibiting inflammation and protecting vascular endothelium via PI3K/Akt signal pathway

To investigate the effects of Vaspin on lipopolysaccharide (LPS)-induced acute respiratory distress syndrome (ARDS) in mice and explore the possible mechanism. Forty male C57B/L6 mice were randomized equally into control group, LPS group, Vaspin group and wortmannin group with corresponding treatments. The pathological changes of the lung tissues were evaluated by HE staining, and the severity of pulmonary edema was measured according to the wet/dry ratio (W/D) of the lung tissue. The lung permeability was evaluated by detecting total protein concentrations in the bronchoalveolar lavage fluid (BALF) using bicinchoninic acid (BCA) assay. Myeloperoxidase (MPO) activity in the lung tissue was detected using a MPO assay kit, and the levels of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in the lungs were measured using ELISA. Immunohistochemical staining was performed to detect the expression of vascular cell adhesion molecule-1 (VCAM-1) and Western blotting was used to detect the protein expressions of cleaved caspase-3 and p-Akt in the lung tissues. Compared with the control group, the mice in LPS group displayed typical ARDS pathological changes in the lungs with significantly increased W/D, total protein concentrations in BALF, lung MPO activity, levels of IL-1β and TNF-α, and pulmonary expressions of VCAM-1 and cleaved caspase-3 (P<0.05) but decreased expression of p-Akt (P<0.05). These changes induced by LPS were significantly alleviated by the administration of Vaspin (P<0.05). The protective effects of Vaspin against ARDS were obviously attenuated by the PI3K inhibitor wortmannin (P<0.05). Vaspin protects against LPS-induced ARDS in mice possibly by inhibiting inflammation and protecting vascular endothelium through upregulation of the PI3K/Akt signal pathway.

Protective effects of tiotropium alone or combined with budesonide against cadmium inhalation induced acute neutrophilic pulmonary inflammation in rats.

As a potent bronchodilator, the anti-inflammatory effects of tiotropium and its interaction with budesonide against cadmium-induced acute pulmonary inflammation were investigated. Compared to values obtained in rats exposed to cadmium, cytological analysis indicated a significant decrease of total cell and neutrophil counts and protein concentration in bronchoalveolar lavage fluid (BALF) in rats pretreated with tiotropium (70μg/15ml or 350μg/15ml). Zymographic tests showed a decrease of MMP-2 activity in BALF in rats pretreated only with high concentration of tiotropium. Histological examination revealed a significant decrease of the severity and extent of inflammatory lung injuries in rats pretreated with both tested concentrations of tiotropium. Though tiotropium (70μg/15ml) or budesonide (250μg/15ml) could not reduce cadmium-induced bronchial hyper-responsiveness, their combination significantly decreased bronchial contractile response to methacholine. These two drugs separately decreased the neutrophil number and protein concentration in BALF but no significant interaction was observed when both drugs were combined. Although no inhibitory effects on MMP-2 and MMP-9 was observed in rats pretreated with budesonide alone, the combination with the ineffective dose of tiotropium induced a significant reduction on these parameters. The inhibitory effect of tiotropium on lung injuries was not influenced by budesonide which alone induced a limited action on the severity and extent of inflammatory sites. Our findings show that tiotropium exerts anti-inflammatory effects on cadmium-induced acute neutrophilic pulmonary inflammation. The combination of tiotropium with budesonide inhibits cadmium-induced inflammatory injuries with a synergistic interaction on MMP-2 and MMP-9 activity and airway hyper-responsiveness.

Potentiation of Sodium Metabisulfite Toxicity by Propylene Glycol in Both in Vitro and in Vivo Systems.

Many consumer products used in our daily lives result in inhalation exposure to a variety of chemicals, although the toxicities of the active ingredients are not well known; furthermore, simultaneous exposure to chemical mixtures occurs. Sodium metabisulfite (SM) and propylene glycol (PG) are used in a variety of products. Both the cytotoxicity and the sub-acute inhalation toxicity of each chemical and their mixtures were evaluated. Assays for cell viability, membrane damage, and lysosome damage demonstrated that SM over 100 μg/ml induced significant cytotoxicity; moreover, when PG, which was not cytotoxic, was mixed with SM, the cytotoxicity of the mixture was enhanced. Solutions of 1, 5, and 20% SM, each with 1% PG solution, were prepared, and the whole body of rats was exposed to aerosols of the mixture for 6 h/day, 5 days/week for 2 weeks. The rats were sacrificed 1 (exposure group) or 7 days (recovery group) after termination of the exposure. The actual concentration of SM in the low-, medium-, and high-exposure groups was 3.91 ± 1.26, 35.73 ± 6.01, and 80.98 ± 5.47 mg/m3, respectively, and the actual concentration of PG in each group was 6.47 ± 1.25, 8.68 ± 0.6, and 8.84 ± 1.77 mg/m3. The repeated exposure to SM and PG caused specific clinical signs including nasal sound, sneeze, and eye irritation which were not found in SM single exposure. In addition, the body weight of treatment group rats decreased compared to that of the control group rats in a time-dependent manner. The total protein concentration and lactate dehydrogenase activity in the bronchoalveolar lavage fluid (BALF) increased. Histopathological analysis of the lungs, liver, and nasal cavity was performed. Adverse effects were observed in the nasal cavity, with squamous cell metaplasia identified in the front of the nasal cavity in all high-exposure groups, which completely recovered 7 days after exposure was terminated. Whereas inhalation of SM for 2 weeks only reduced body weight in the high-dose group, inhalation of SM and PG mixtures for 2 weeks significantly decreased body weight and induced metaplasia of the respiratory epithelium into squamous cells in the medium- and high-dose groups. In conclusion, PG potentiated the toxicity of SM in human lung epithelial cells and the inhalation toxicity in rats.

Glucose-6-phosphate dehydrogenase inhibition attenuates acute lung injury through reduction in NADPH oxidase-derived reactive oxygen species.

Acute lung injury (ALI) is a heterogeneous disease with the hallmarks of alveolar capillary membrane injury, increased pulmonary oedema and pulmonary inflammation. The most common direct aetiological factor for ALI is usually parenchymal lung infection or haemorrhage. Reactive oxygen species (ROS) generated by nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX2) are thought to play an important role in the pathophysiology of ALI. Glucose-6-phosphate dehydrogenase (G6PD) plays an important role both in production of ROS as well as their removal through the supply of NADPH. However, how G6PD modulation affects NOX2-mediated ROS in the airway epithelial cells (AECs) during acute lung injury has not been explored previously. Therefore, we investigated the effect of G6PD inhibitor, 6-aminonicotinamide on G6PD activity, NOX2 expression, ROS production and enzymatic anti-oxidants in AECs in a mouse model of ALI induced by lipopolysaccharide (LPS). ALI led to increased G6PD activity in the AECs with concomitant elevation of NOX2, ROS, SOD1 and nitrotyrosine. G6PD inhibitor led to reduction of LPS-induced airway inflammation, bronchoalveolar lavage fluid protein concentration as well as NOX2-derived ROS and subsequent oxidative stress. Conversely, ALI led to decreased glutathione reductase activity in AECs, which was normalized by G6PD inhibitor. These data show that activation of G6PD is associated with enhancement of oxidative inflammation in during ALI. Therefore, inhibition of G6PD might be a beneficial strategy during ALI to limit oxidative damage and ameliorate airway inflammation.

The comparison of heparan sulfate and its fragments on the protection against extracellular histones during the pathogenesis of acute respiratory distress syndrome

Objective: In order to explore the role of heparan sulfate (HS) during the pathogenesis of acute respiratory distress syndrome (ARDS) , the protective effect of HS and its fragments against extracellular histones was compared. Methods: Calf thymus histones (CTH) were injected via femoral vein to induce ARDS in rats. HS, HS fragments or saline was intraperitoneally injected (10mg/kg, Q6h, 24h) to test the protective effect against CTH. The ratio of wet/dry lung weight, protein content in bronchoalveolar lavage fluid (BALF) , total leukocyte and neutrophil count in BALF were measured. Results: After CTH injection, the ratio of wet/dry lung weight (5.7±0.95) was much higher than the saline control group (3.1±0.15). The protein content (0.47±0.086mg/ml) , total leukocyte[ (97.4±15.6l) ×10(4)/ml] and neutrophil (18±3.4/LPF) in BALF were obviously increased compared with the saline control group. The intervention of HS evidently decreased ratio of wet/dry lung weight (4.2±0.41) , protein content[ (0.26±0.019) mg/ml], leukocyte[ (61.3±5.74) ×10(4)/ml] and neutrophil (12±1.8/LPF) in BALF. HS fragments also decreased ratio of wet/dry lung weight, protein content, leukocyte and neutrophil count in BALF though the strength was much less than HS. Conclusion: HS and its fragments could provide protection against extracellular histones during the pathogenesis of ARDS. For the protective effect full length HS was much better than HS fragments.

Protective effects of esmolol on lung function in septic rat

Objective To explore the protective effects of short-acting β1 receptor blocker esmolol on lung injury in septic rats and explore its mechanism. Methods Fifty-four male Sprague-Dawley rats were randomly(random number) divided into three groups (n=18 in each), namely sham operation group(Sham), sepsis group(Spesis),esmolol group(ES).Each group were further divided into three subgroups of 6 h,12 h and 24 h(n=6 in each subgroup).The sepsis models were established with cecal ligation and puncture (CLP) in rats of Sepsis group and ES group, while rats of SH group were treated with laparotomy without CLP. Dwelled cannulation in the left internal jugular vein was performed after the model establishment. Saline in the rate of 1 mL/h was continuously pumped intravenously into the rats of SH group and Sepsis group by micro pump for 6 h, and esmolol solution in the rate of 1 mL/h [15 mg/(kg·h)] was continuously pumped into the rats of ES group for 6h.Rats in each subgroup were sacrificed at 6h,12h and 24 h after modeling, separately. The levels of catecholamine (CA) in plasma and levels of NF-κB in lung tissue were measured by ELISA. The protein levels of TLR4 and NF-κB in lung tissue were detected by Western Blot. The protein content of alveolar lavage fluid, wet/dry weight ratio(W/D) of lung tissue, lung coefficient and lung water content were measured. The pathological changes of lung tissues were observed under an optical microscope. Results ①The levels of CA in plasma in Sepsis and ES groups at 6 h after modeling was significantly increased compared with Sham group (P<0.05), and the level of CA in plasma increased significantly in ES group compared with Sham and Sepsis groups (P<0.05). At 12 h after modeling, level of CA in plasma was still significantly increased in ES group compared with SH group (P<0.05); At 24 h after modeling, level of CA in plasma was increased significantly in ES group compared with Sepsis group (P<0.05).②At 6 h,12 h and 24 h after modeling, levels of NF-κB in lung tissue were significantly increased in Sepsis group and ES group compared with SH group (P<0.05),and levels of NF-κB in lung tissue in ES group were also significantly lower than those in Sepsis group at given intervals (P<0.05).③ At 6 h,12 h and 24 h after modeling, protein content of alveolar lavage fluid, wet/dry weight ratio (W/D) in lung tissue, lung coefficient and lung water content in Sepsis group and ES group were significantly increased compared with SH group (P<0.05), and protein content of alveolar lavage fluid, wet/dry weight ratio (W/D) in lung tissue, lung coefficient and lung water content in ES group also significantly lower than those in Sepsis group at given intervals (P<0.05).④Western Blot revealed at 6 h,12 h and 24 h after modeling, the protein levels of TLR4 and NF-κB in lung tissue in Sepsis group and ES group were significantly increased compared with SH group (P<0.05),and the protein levels of TLR4 and NF-κB in lung tissue of ES group were also significantly lower than those in Sepsis group at all given intervals (P<0.05).⑤The lightest degree of pathological change was observed in SH group, the most serious degree of pathological change was found in Sepsis group, and the pathological change was significantly alleviated in ES group compared with Sepsis group whereas the pathological change was significantly increased compared with SH group at all given intervals. Conclusions Esmolol can promote the secretion of catecholamine,inhibit the release of inflammatory cytokines,reduce lung permeability,reduce pulmonary edema,and thus play an important role in protecting the lungs.The mechanism may be that esmolol improves the responsiveness of β-adrenergic receptor to catecholamine,and regulates the level of inflammatory cytokines by inhibiting TLR4—NF-κB—TNF-α signaling pathways,thus exerting protective effect on the lungs. Key words: Sepsis; Esmolol; Catecholamine; TNF-α; NF-κB; Toll-like receptors 4

Curcumin ameliorated ventilator-induced lung injury in rats

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

Microparticles in red cell concentrates prime polymorphonuclear neutrophils and cause acute lung injury in a two-event mouse model

Red cell-derived microparticles (RMPs) are potential mediators of transfusion-related acute lung injury (TRALI). The aim of this study was to investigate the effects of microparticles present in red cell concentrates (RCC) on polymorphonuclear neutrophil (PMN) respiratory burst and acute lung injury (ALI) in mice. Microparticles (MPs) in RCC supernatant were quantified using flow cytometry. The priming activity of either isolated MPs or RCC supernatant toward human PMN was measured in vitro. Mice were injected with lipopolysaccharide (LPS), followed by an infusion of either isolated MPs or heat-treated RCC supernatant. The lungs were harvested to assess myeloperoxidase (MPO) activity, histology and pulmonary edema. Protein content in bronchoalveolar lavage fluid (BALF) was measured. The number of RMPs increased significantly during storage. Both isolated MPs and the supernatants from RCCs that had been stored for 28 and 35days effectively primed the PMN respiratory burst. The infusion of isolated MPs or supernatants that had been stored for >28days into LPS-treated mice caused ALI. The filtered supernatant resulted in significantly ameliorated mouse ALI. MPs that accumulate during RCC storage prime the PMN respiratory burst and cause ALI in a two-event mouse model.

Tanshinone II is a potent candidate for treatment of lipopolysaccharide-induced acute lung injury in rat model.

The present study aimed to investigate the effect of tanshinone II, isolated from Salvia miltiorrhiza Bunge, on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in rats. Male Sprague-Dawley rats were divided into three groups: Control, LPS and tanshinone II. Animals in the tanshinone II and LPS groups were administered 10 mg/kg LPS, whereas those in the control group received an equal volume of normal saline. Tanshinone II treatment group were injected with 30 nm/kg tanshinone II at 1 h after LPS administration. The results revealed that LPS administration increased the bronchoalveolar lavage fluid protein concentration significantly compared with the control group. However, tanshinone II treatment significantly inhibited the LPS-induced increase in protein level. Treatment of the LPS-administered rats with tanshinone II prevented the formation of pulmonary edema, which was evidenced by low values for wet to dry lung weight ratio. The activity of myeloperoxidase and expression of malondialdehyde were significantly lower in lung homogenates from the tanshinone II group compared with the LPS group. Furthermore, tanshinone II treatment inhibited the expression of tumor necrosis factor-α and interleukin-6 in the blood plasma. Tissue sections of the tanshinone II group exhibited normal morphology and absence of neutrophil accumulation. However, in the LPS group, neutrophils accumulated and penetrated into the pulmonary tissues. These results suggested that tanshinone II protects the rats from LPS-induced ALI. Therefore tanshinone II may have clinical applications in the treatment of ALI.

Preventive and Therapeutic Effects of Thymol in a Lipopolysaccharide-Induced Acute Lung Injury Mice Model.

Acute lung injury (ALI) is a life-threatening syndrome which causes a high mortality rate worldwide. In traditional medicine, lots of aromatic plants-such as some Thymus species-are used for treatment of various lung diseases including pertussis, bronchitis, and asthma. Thymol, one of the primary active constituent derived from Thymus vulgaris (thyme), has been reported to exhibit potent anti-microbial, anti-oxidant, and anti-inflammatory activities in vivo and in vitro. The present study aims to investigate the protective effects of thymol in lipopolysaccharide (LPS)-induced lung injury mice model. In LPS-challenged mice, treatment with thymol (100mg/kg) before or after LPS challenge significantly improved pathological changes in lung tissues. Thymol also inhibited the LPS-induced inflammatory cells influx, TNF-α and IL-6 releases, and protein concentration in bronchoalveolar lavage fluid (BALF). Additionally, thymol markedly inhibited LPS-induced elevation of MDA and MPO levels, as well as reduction of SOD activity. Further study demonstrated that thymol effectively inhibited the NF-κB activation in the lung. Taken together, these results suggested that thymol might be useful in the therapy of acute lung injury.

Pharmacological modulation of C-X-C motif chemokine receptor 4 influences development of acute respiratory distress syndrome after lung ischaemia-reperfusion injury.

Activation of C-X-C motif chemokine receptor 4 (CXCR4) has been reported to result in lung protective effects in various experimental models. The effects of pharmacological CXCR4 modulation on the development of acute respiratory distress syndrome (ARDS) after lung injury, however, are unknown. Thus, we studied whether blockade and activation of CXCR4 influences development of ARDS in a unilateral lung ischaemia-reperfusion injury rat model. Anaesthetized, mechanically ventilated animals underwent right lung ischaemia (series 1, 30minutes; series 2, 60minutes) followed by reperfusion for 300minutes. In series 1, animals were treated with vehicle or 0.7μmol/kg of AMD3100 (CXCR4 antagonist) and in series 2 with vehicle, 0.7 or 3.5μmol/kg ubiquitin (non-cognate CXCR4 agonist) within 5minutes of reperfusion. AMD3100 significantly reduced PaO2 /FiO2 ratios, converted mild ARDS with vehicle treatment into moderate ARDS (PaO2 /FiO2 ratio<200) and increased histological lung injury. Ubiquitin dose-dependently increased PaO2 /FiO2 ratios, converted moderate-to-severe into mild-to-moderate ARDS and reduced protein content of bronchoalveolar lavage fluid (BALF). Measurements of cytokine levels (TNFα, IL-6, IL-10) in lung homogenates and BALF showed that AMD3100 reduced IL-10 levels in homogenates from post-ischaemic lungs, whereas ubiquitin dose-dependently increased IL-10 levels in BALF from post-ischaemic lungs. Our findings establish a cause-effect relationship for the effects of pharmacological CXCR4 modulation on the development of ARDS after lung ischaemia-reperfusion injury. These data further suggest CXCR4 as a new drug target to reduce the incidence and attenuate the severity of ARDS after lung injury.

Radiation Mitigating Properties of Intranasally Administered KL4Surfactant in a Murine Model of Radiation-Induced Lung Damage

The threat of exposure to ionizing radiation from a nuclear reactor accident or deliberate terrorist actions is a significant public health concern. The lung is particularly susceptible to radiation-induced injury from external sources or inhalation of radioactive particles from radioactive fallout. Radiation-induced lung disease can manifest with an acute radiation pneumonitis and/or delayed effects leading to pulmonary fibrosis. As prior warning of radiation exposure is unlikely, medical countermeasures (MCMs) to mitigate radiation-induced lung disease that can be given in mass-casualty situations many hours or days postirradiation are needed to prevent both early and late lung damage. In this study, KL4 surfactant (lucinactant) was evaluated as a radiation mitigator in a well-characterized mouse model of targeted thoracic radiation exposure, for its effect on both early (several weeks) and late (18 weeks) lung damage. Here, 120 mg/kg total phospholipid of KL4 surfactant was administered twice daily intranasally, (enabling intrapulmonary inhalation of drug) to C57BL/6 mice 24 h after a single 13.5 Gy dose of thoracic irradiation (LD50 dose). Both early and chronic phase (2 and 4 weeks and 18 weeks postirradiation, respectively) assessments were performed. Mice were evaluated for evidence of reduced arterial blood oxygenation and early and chronic lung and systemic inflammation, lung fibrosis and oxidative stress. Analysis was done by performing lung function/respiration dynamics and measuring cellular protein content of bronchoalveolar lavage fluid (BALF), and levels of cytokines, 8-iso-prostaglandin F2α, hydroxyproline in lung and plasma, along with evaluating lung histology. The results of this study showed that intranasal delivery of KL4 surfactant was able to preserve lung function as evidenced by adequate arterial oxygen saturation and reduced lung inflammation and oxidative stress; total white count and absolute neutrophil count was decreased in BALF, as were plasma pro-inflammatory cytokine levels and biomarker of oxidative stress. KL4 surfactant is a promising MCM for mitigation of lung tissue damage after targeted, thoracic irradiation and has the potential to be developed as a broad-spectrum, multi-use MCM against chemical, biological, radiological or nuclear threat agents with potential to cause lung injury.

NPS 2143, a selective calcium-sensing receptor antagonist inhibits lipopolysaccharide-induced pulmonary inflammation

NPS 2143, a novel and selective antagonist of calcium-sensing receptor (CaSR) has been reported to possess anti-inflammatory activity. In the present study, we examined the protective effect of NPS 2143 on lipopolysaccharide (LPS)-induced acute lung injury (ALI). NPS 2143 pretreatment significantly inhibited the influx of inflammatory cells and the expression of monocyte chemoattractant protein-1 (MCP-1) in the lung of mice with LPS-induced ALI. NPS 2143 decreased the levels of neutrophil elastase (NE) and protein concentration in the bronchoalveolar lavage fluid (BALF). NPS 2143 also reduced the production of inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in the BALF and serum. In addition, NPS 2143 attenuated the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), and increased the activation of AMP-activated protein kinase (AMPK) in the lung. NPS 2143 also downregulated the activation of nuclear factor-kappa B (NF-κB) in the lung. In LPS-stimulated H292 airway epithelial cells, NPS 2143 attenuated the releases of IL-6 and MCP-1. Furthermore, NPS 2143 upregulated the activation of AMPK and downregulated the activation of NF-κB. These results suggest that NPS 2143 could be potential agent for the treatment of inflammatory diseases including ALI.

All-trans retinoic acid attenuates bleomycin-induced pulmonary fibrosis via downregulating EphA2-EphrinA1 signaling

The role of all-trans retinoic acid (ATRA) in pulmonary fibrosis is relatively unknown, although this metabolite modulates cell differentiation, proliferation, and development. We aimed to evaluate the role of ATRA in bleomycin-induced pulmonary fibrosis, and whether the mechanism involves EphA2-EphrinA1 and PI3K-Akt signaling. We evaluated three groups of mice: a control group (intraperitoneal DMSO injection 3 times weekly after PBS instillation), bleomycin group (intraperitoneal DMSO injection 3 times weekly after bleomycin instillation), and bleomycin + ATRA group (intraperitoneal ATRA injection 3 times weekly after bleomycin instillation). The cell counts and protein concentration in the bronchoalveolar lavage fluid (BALF), changes in histopathology, Ashcroft score, hydroxyproline assay, expression of several signal pathway proteins including EphA2-EphrinA1, and PI3K-Akt, and cytokine levels were compared among the groups. We found that bleomycin significantly increased the protein concentration in the BALF, Ashcroft score in lung tissue, and hydroxyproline contents in lung lysates. Furthermore, bleomycin upregulated EphA2, EphrinA1, PI3K 110γ, Akt, IL-6 and TNF-α. However, administration of ATRA attenuated the upregulation of EphA2-EphrinA1 and PI3K-Akt after bleomycin instillation, and decreased pulmonary fibrosis. In addition, ATRA suppressed IL-6 and TNF-α production induced by bleomycin-induced injury. Collectively, these data suggest that ATRA attenuates bleomycin-induced pulmonary fibrosis by regulating EphA2-EphrinA1 and PI3K-Akt signaling.