Tracheal Instillation Research Articles

Animal Models of Chronic Obstructive Pulmonary Disease Exacerbations: A Review of the Current Status

Chronic obstructive pulmonary disease (COPD), characterized by airflow limitation and manifested as emphysema and chronic airway obstruction, is a major cause of morbidity and mortality worldwide, resulting in an economic and social burden that is both substantial and increasing. The natural history of COPD involves systemic manifestations, such as skeletal muscle wasting and cardiovascular impairment, and frequent exacerbations. The latter are caused by bacterial or viral infections and have major implications for patients and healthcare systems. There are no effective therapies to prevent or reverse these events. Smoking cessation remains the most effective intervention for reducing disease progression. Animal models of COPD and of COPD exacerbations have been developed to advance understanding of the pathogenesis of COPD and the role of infections in its severity. Cigarette smoke exposure, tracheal instillation of elastase, and genetic manipulation are commonly used to reproduce baseline COPD conditions, each with its advantages and disadvantages. Intratracheal instillation of lipopolysaccharide (LPS), bacteria, or viruses are the most common interventions used to exacerbate baseline COPD. This review highlights the three major animal models used for induction of emphysema and its exacerbations. Further exploration of these models should facilitate identification of new therapeutic approaches for COPD.

3,5,4'-Tri-O-acetylresveratrol decreases seawater inhalation-induced acute lung injury by interfering with the NF-κB and i-NOS pathways.

Drowning is a cause of accidental mortality. However, survival may result in acute lung injury. The aim of the present study was to evaluate the effects of 3,5,4'-tri-O-acetylresveratrol (AC-Res) on acute lung injury (ALI) induced by seawater inhalation in rats. ALI models were established by the tracheal instillation of artificial seawater with or without 50 mg/kg AC-Res pretreatment for 7 days. Lung samples from different groups were harvested 4 h after the model was established. Histological changes, blood vessel permeability, inflammatory factor secretion and expression states of the nuclear factor-κB (NF-κB) and inducible NOS (i-NOS) pathway were assessed to evaluate seawater‑induced lung injury and the protective effects of acetylated resveratrol. The results showed that seawater inspiration led to physiological structure changes and an increased permeability of blood vessels. In addition, seawater stimulation enhanced the expression levels of nitric oxide (NO), tumor necrosis factor α (TNF-α) and interleukin-1 β (IL-1β) secretion in vitro and in vivo. Notably, seawater inhalation increased NF-κB and i-NOS expression in lungs and cells. On the other hand, pretreatment of AC-Res inhibited the abnormal expression of the NF-κB and i-NOS pathways, followed by decreased NO, TNF-α and IL-1β secretion, protein and cell content in bronchoalveolar lavage fluid (BALF) and Evans blue, protein and cell infiltration from blood vessels into lung tissues. The results therefore suggest that AC-Res attenuated seawater inhalation induced‑ALI by interfering with the NF-κB and i-NOS pathways.

Decreased lung compliance increases preload dynamic tests in a pediatric acute lung injury model

BackgroundPreload dynamic tests, pulse pressure variation (PPV) and stroke volume variation (SVV) have emerged as powerful tools to predict response to fluid administration. The influence of factors other than preload in dynamic preload test is currently poorly understood in pediatrics. The aim of our study was to assess the effect of tidal volume (VT) on PPV and SVV in the context of normal and reduced lung compliance in a piglet model. Material and methodTwenty large-white piglets (5.2±0.4kg) were anesthetized, paralyzed and monitored with pulse contour analysis. PPV and SVV were recorded during mechanical ventilation with a VT of 6 and 12mL/kg (low and high VT, respectively), both before and after tracheal instillation of polysorbate 20. ResultsBefore acute lung injury (ALI) induction, modifications of VT did not significantly change PPV and SVV readings. After ALI, PPV and SVV were significantly greater during ventilation with a high VT compared to a low VT (PPV increased from 8.9±1.2 to 12.4±1.1%, and SVV from 8.5±1.0 to 12.7±1.2%, both P<0.01). ConclusionsThis study found that a high VT and reduced lung compliance due to ALI increase preload dynamic tests, with a greater influence of the latter. In subjects with ALI, lung compliance should be considered when interpreting the preload dynamic tests.

Instillation and Fixation Methods Useful in Mouse Lung Cancer Research

The ability to instill live agents, cells, or chemicals directly into the lung without injuring or killing the mice is an important tool in lung cancer research. Although there are a number of methods that have been published showing how to intubate mice for pulmonary function measurements, none are without potential problems for rapid tracheal instillation in large cohorts of mice. In the present paper, a simple and quick method is described that enables an investigator to carry out such instillations in an efficient manner. The method does not require any special tools or lighting and can be learned with very little practice. It involves anesthetizing a mouse, making a small incision in the neck to visualize the trachea, and then inserting an intravenous catheter directly. The small incision is quickly closed with tissue adhesive, and the mice are allowed to recover. A skilled student or technician can do instillations at an average rate of 2 min/mouse. Once the cancer is established, there is frequently a need for quantitative histologic analysis of the lungs. Traditionally pathologists usually do not bother to standardize lung inflation during fixation, and analyses are often based on a scoring system that can be quite subjective. While this may sometime be sufficiently adequate for gross estimates of the size of a lung tumor, any proper stereological quantification of lung structure or cells requires a reproducible fixation procedure and subsequent lung volume measurement. Here we describe simple reliable procedures for both fixing the lungs under pressure and then accurately measuring the fixed lung volume. The only requirement is a laboratory balance that is accurate over a range of 1 mg-300 g. The procedures presented here thus could greatly improve the ability to create, treat, and analyze lung cancers in mice.

Angiotensin-Converting Enzyme 2 Improves Arterial Hypoxemia in Meconium-Induced Acute Lung Injury in Piglets

Objective. Meconium aspiration induces acute lung injury (ALI) in neonates born through meconium-stained amniotic fluid. As yet, there is no specific therapy for improving the outcome. Recently, angiotensin-converting enzyme 2 (ACE2), which inactivates angiotensin II (Ang II), has been shown to ameliorate murine ALI. Design. To evaluate the therapeutic potential of this substance, we studied ACE2 in a piglet model of ALI induced by meconium aspiration. Subjects. Twelve anesthetized piglets were subjected in an animal research laboratory. ALI was induced by tracheal meconium instillation. Thereafter, six animals were randomly assigned to the ACE2 group, while another 6 served as control. Measurements. Systemic, pulmonary hemodynamic, and blood gas exchange parameters and Ang II levels were examined before ALI induction and at various time points after administering ACE2 or saline. In addition, ventilation-perfusion distribution of the lung was assessed by the multiple inert gas elimination technique (MIGET). Main Results. Animals treated with ACE2 maintained significantly higher arterial partial pressures of oxygen (Pao2) and lower arterial partial pressures of carbon dioxide (Paco2), respectively. Furthermore, Ang II, which was substantially increased, returned to basal values. Conclusion. In summary, ACE2 improves blood gas exchange in meconium-induced ALI in piglets.

Effect of Liuweibuqi capsule, a Chinese patent medicine, on the JAK1/STAT3 pathway and MMP9/TIMP1 in a chronic obstructive pulmonary disease rat model

ObjectiveTo observe effect of Liuweibuqi Capsule, a Traditional Chinese Medicine (TCM), on the janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway and matrix metalloproteinases (MMPs) in a chronic obstructive pulmonary disease (COPD) rat model with lung deficiency in terms of TCM's pattern differentiation. MethodsRats were randomly divided into a normal group, model group, Liuweibuqi group, Jinshuibao group, and spleen aminopeptidase group (n= 10). Aside from the normal group, all rats were exposed to smoke plus lipopolysaccharide tracheal instillation to establish the COPD model with lung deficiency. Models were established after 28 days and then the normal and model groups were given normal saline (0.09 g/kg), Liuweibuqi group was given Liuweibuqi capsule (0.35 g/kg), Jinshuibao group was given Jinshuibao capsules (0.495 g/kg), and the spleen group was given spleen aminopeptidase (0.33 mg/kg), once a day for 30 days. Changes in symptoms, signs, and lung histology were observed. Lung function was measured with a spirometer. Serum cytokines were detected using enzyme-linked immunosorbent assay, and changes in the JAK/STAT pathway, MMP-9, and MMPs inhibitor 1 (TIMP1) were detected by immunohistochemistry, RT-PCR, and western blotting, respectively. ResultsCompared with the normal group, lung tissue was damaged, and lung function was reduced in the model control group. Additionally, the levels of interleukin (IL)-1β, γ interferon (IFN-γ), and IL-6 were higher, while IL-4 and IL-10 were lower in the model control group than those in the normal group. The expressions of JAK1, STAT3, p-STAT3, and MMP-9 mRNA and protein in lung tissue were higher, and TIMP1 mRNA and protein was lower in the model group compared with the normal group. After treatment, compared with the model group, the expression of inflammatory cytokines was lower in each treatment group, and expressions of JAK/STAT pathway, MMPs were lower. Compared with the positive control groups, the Jinshuibao and spleen aminopeptidase groups, lung function was better, and JAK1, STAT3, and p-STAT3 protein were lower and TIMP1 was higher in the Liuweibuqi group. ConclusionLiuweibuqi capsules can improve the symptoms of COPD possibly by regulating the expression of the JAK1/STAT3 pathway and MMP9/TIMP1.

Upregulation of α1A-Subtype Adrenergic Signaling is Beneficial in Failing Right Ventricle (RV)

Compared to non-failing RV, we reported that in failing RV the inotropic response to stimulation of α1-adrenergic receptors (α1-ARs) was increased. Recently we found that a single α1-AR subtype (α1A) was responsible. However, the significance of increased α1A responses remains unclear.Goal: Determine if increased α1A-mediated inotropy is beneficial in RV failure and if the α1A-subtype is a potential therapeutic target.Methods: We used a RV-specific failure model in mice. Tracheal instillation of the fibrogenic antibiotic bleomycin induced pulmonary fibrosis, pulmonary hypertension, and RV failure within 2-3 wk. We assessed survival and investigated the cellular mechanisms contributing to increased α1A inotropy in failing RV. Furthermore, in the context of bleomycin induced RV failure, we determined if there was a beneficial effect of chronic (2 wk.) treatment of mice with the α1A agonist A61603 using osmotic mini-pump infusion.Results: For non-failing RV, acute α1A stimulation with A61603 induced a negative inotropic effect. In contrast, for failing RV, acute α1A stimulation induced a positive inotropic effect. A subset of animals did not manifest this switch in the inotropic response, and had significantly worse survival, suggesting that upregulation of the α1A inotropic response was beneficial in failing RV. Upregulation of α1A inotropy in failing RV was associated with increased Ca2+ transients, and increased phosphorylation of the myosin regulatory light chain (increased myofilament Ca2+ sensitivity). Moreover, consistent with a beneficial effect of increased α1A activation in failing RV, chronic treatment of mice with A61603 for 2 wk. resulted in less myocardial fibrosis, lower serum troponin level, and better in-vivo function assessed with echocardiography.Conclusion: Increased α1A subtype activation is beneficial in the failing RV. The α1A-subtype is a potential therapeutic target in failing RV.

Receptor for advanced glycation end-products regulates lung fluid balance via protein kinase C-gp91(phox) signaling to epithelial sodium channels.

The receptor for advanced glycation end-products (RAGE), a multiligand member of the Ig family, may play a crucial role in the regulation of lung fluid balance. We quantified soluble RAGE (sRAGE), a decoy isoform, and advanced glycation end-products (AGEs) from the bronchoalveolar lavage fluid of smokers and nonsmokers, and tested the hypothesis that AGEs regulate lung fluid balance through protein kinase C (PKC)-gp91(phox) signaling to the epithelial sodium channel (ENaC). Human bronchoalveolar lavage samples from smokers showed increased AGEs (9.02 ± 3.03 μg versus 2.48 ± 0.53 μg), lower sRAGE (1,205 ± 292 pg/ml versus 1,910 ± 263 pg/ml), and lower volume(s) of epithelial lining fluid (97 ± 14 ml versus 133 ± 17 ml). sRAGE levels did not predict ELF volumes in nonsmokers; however, in smokers, higher volumes of ELF were predicted with higher levels of sRAGE. Single-channel patch clamp analysis of rat alveolar epithelial type 1 cells showed that AGEs increased ENaC activity measured as the product of the number of channels (N) and the open probability (Po) (NPo) from 0.19 ± 0.08 to 0.83 ± 0.22 (P = 0.017) and the subsequent addition of 4-hydroxy-2, 2, 6, 6-tetramethylpiperidine-N-oxyl decreased ENaC NPo to 0.15 ± 0.07 (P = 0.01). In type 2 cells, human AGEs increased ENaC NPo from 0.12 ± 0.05 to 0.53 ± 0.16 (P = 0.025) and the addition of 4-hydroxy-2, 2, 6, 6-tetramethylpiperidine-N-oxyl decreased ENaC NPo to 0.10 ± 0.03 (P = 0.013). Using molecular and biochemical techniques, we observed that inhibition of RAGE and PKC activity attenuated AGE-induced activation of ENaC. AGEs induced phosphorylation of p47(phox) and increased gp91(phox)-dependent reactive oxygen species production, a response that was abrogated with RAGE or PKC inhibition. Finally, tracheal instillation of AGEs promoted clearance of lung fluid, whereas concomitant inhibition of RAGE, PKC, and gp91(phox) abrogated the response.

Role of Foxp3/Treg and RORγt/Th17 cell imbalance in rat model of chronic obstructive pulmonary disease

To observe the changes in forkhead/winged helix transcription factor p3(Foxp3), regulatory T cells (Treg), retinoid-related orphan receptor gamma (RORγt) in rat model of chronic obstructive pulmonary disease (COPD). Twenty Sprague-Dawley (SD) rats were randomly divided into normal control group and COPD model group, with 10 rats in each group. The COPD model was reproduced by smoke inhalation and tracheal instillation of lipopolysaccharide (LPS), and no such treatment was conducted in normal control group. Twenty-eight days after the model reproduction, the pulmonary function was determined, the pathological changes of lung tissue were observed with haematoxylin-eosin (HE) staining, interleukins (IL-6, IL-10) in serum were detected by enzyme-linked immunosorbent assay (ELISA), CD4⁺ CD25⁺ Foxp3⁺ Treg of peripheral blood was determined by flow cytometry, and the expressions of Foxp3, RORγt, IL-17 protein in lung tissue were assayed by Western Blot. Under light microscope, significal interstitial infiltration of inflammatory cells was found in alveoli and interstitial tissue of the lung, and destruction of alveolar tissue, alveolar wall thinning, and even rupture to fuse into bullae, and bleeding into alveoli in different degress could be observed. Compared with the normal control group, forced vital capacity (FVC), forced expiratory volume in 0.3 second (FEV0.3), FEV0.3/FVC, peak expiratory flow (PEF) in model group were significantly decreased [FVC (mL): 8.04 ± 2.03 vs. 9.97 ± 2.14, FEV0.3 (mL): 6.16 ± 2.23 vs. 8.84 ± 2.12, FEV0.3/FVC: 0.70 ± 0.09 vs. 0.85 ± 0.11, PEF (mL/s): 33.56 ± 4.76 vs. 40.14 ± 5.64, P<0.05 or P<0.01]. Serum IL-6 was obviously increased (ng/L: 93.17 ±20.96 vs. 76.28 ± 13.24, P<0.05), IL-10 was significantly decreased (ng/L: 78.62 ± 15.17 vs. 104.34 ± 19.46, P<0.01), and CD4⁺ CD25⁺ FoxP3(+)Treg was significantly diminished [(2.75 ± 0.83)% vs. (4.16 ± 1.14)%, P<0.01] in model group compared with those in the normal control group. The expression of Foxp3 protein in lung tissue in model group was significantly down-regulated compared with that in the normal control group (gray scale: 0.38 ± 0.15 vs. 0.63 ± 0.11, P<0.01), and RORγt and IL-17 protein expressions were significantly up-regulated [RORγt (gray scale): 0.96 ± 0.23 vs. 0.47 ± 0.11, IL-17 (gray scale): 1.02 ± 0.24 vs. 0.34 ± 0.08, both P<0.01]. Correlation analysis showed that FEV0.3 was positively correlated with Foxp3 (r=0.585, P<0.05), and FEV0.3/FVC was negatively correlated with IL-6 and RORγt (r=-0.655, r=-0.607, both P<0.05). PEF was positively correlated with Treg (r=0.573, P<0.05), and negatively correlated with IL-17 (r=-0.198, P<0.05). IL-6 was negatively correlated with Foxp3(r=-0.603, P<0.05),and positively correlated with RORγt (r=0.588, P<0.05). IL-10 was positively correlated with Treg (r=0.573, P<0.05). Treg was positively correlated with Foxp3 (r=0.607, P<0.05), and negatively correlated with IL-17 (r=-0.569, P<0.05). Foxp3 was negatively correlated with RORγt (r=-0.591, P<0.05). RORγt was positively correlated with IL-17 (r=0.578, P<0.05). There is a relationship among decreased pulmonary function, inflammation and imbalance of Foxp3/Treg and RORγt/Th17 in COPD.

High-density lipoproteins potentiate α1-antitrypsin therapy in elastase-induced pulmonary emphysema.

Several studies report that high-density lipoproteins (HDLs) can carry α1-antitrypsin (AAT; an elastase inhibitor). We aimed to determine whether injection of exogenous HDL, enriched or not in AAT, may have protective effects against pulmonary emphysema. After tracheal instillation of saline or elastase, mice were randomly treated intravenously with saline, human plasma HDL (75 mg apolipoprotein A1/kg), HDL-AAT (75 mg apolipoprotein A1-3.75 mg AAT/kg), or AAT alone (3.75 mg/kg) at 2, 24, 48, and 72 hours. We have shown that HDL-AAT reached the lung and prevented the development of pulmonary emphysema by 59.3% at 3 weeks (alveoli mean chord length, 22.9 ± 2.8 μm versus 30.7 ± 4.5 μm; P < 0.001), whereas injection of HDL or AAT alone only showed a moderate, nonsignificant protective effect (28.2 ± 4.2 μm versus 30.7 ± 5 μm [P = 0.23] and 27.3 ± 5.66 μm versus 30.71 ± 4.96 μm [P = 0.18], respectively). Indeed, protection by HDL-AAT was significantly higher than that observed with HDL or AAT (P = 0.006 and P = 0.048, respectively). This protective effect was associated (at 6, 24, and 72 h) with: (1) a reduction in neutrophil and macrophage number in the bronchoalveolar lavage fluid; (2) decreased concentrations of IL-6, monocyte chemoattractant protein-1, and TNF-α in both bronchoalveolar lavage fluid and plasma; (3) a reduction in matrix metalloproteinase-2 and matrix metalloproteinase-9 activities; and (4) a reduction in the degradation of fibronectin, a marker of tissue damage. In addition, HDL-AAT reduced acute cigarette smoke-induced inflammatory response. Intravenous HDL-AAT treatment afforded a better protection against elastase-induced pulmonary emphysema than AAT alone, and may represent a significant development for the management of emphysema associated with AAT deficiency.

Depletion of Natural Killer Cells Increases Mice Susceptibility in a Pseudomonas aeruginosa Pneumonia Model*

Pseudomonas aeruginosa infection is a clinically relevant infection involved in pneumonia in ICUs. Understanding the type of immune response initiated by the host during pneumonia would help defining new strategies to interfere with the bacteria pathogenicity. In this setting, the role of natural killer cells remains controversial. We assessed the role of systemic natural killer cells in a Pseudomonas aeruginosa mouse pneumonia model. Experimental study. Research laboratory from a university hospital. RjOrl:SWISS and BALB/cJ mice (weight, 20-24 g). Lung injuries were assessed by bacterial load, myeloperoxidase activity, endothelial permeability (pulmonary edema), immune cell infiltrate (histological analysis), proinflammatory cytokine release, and Ly6-G immunohistochemistry. Bacterial loads were assessed in the lungs and spleen. Natural killer cell number and status were assessed in spleen (flow cytometry and quantitative polymerase chain reaction). Depletion of natural killer cells was achieved through an IV anti-asialo-GM1 antibody injection. Pseudomonas aeruginosa tracheal instillation led to an acute pneumonia with a rapid decrease of bacterial load in lungs and with an increase of endothelial permeability, proinflammatory cytokines (tumor necrosis factor-α and interleukin-1β), and myeloperoxidase activity followed by Ly6-G positive cell infiltrate in lungs. Pseudomonas aeruginosa was detected in the spleen. Membrane markers of activation and maturation (CD69 and KLRG1 molecules) were increased in splenic natural killer cells during Pseudomonas aeruginosa infection. Splenic natural killer cells activated upon Pseudomonas aeruginosa infection produced interferon-γ but not interleukin-10. Ultimately, mice depleted of natural killer cells displayed an increased neutrophil numbers in the lungs and an increased mortality rate without bacterial load modifications in the lungs, indicating that mice depleted of natural killer cells were much more susceptible to infection compared with control animals. We report for the first time that natural killer cells play a major role in the mice susceptibility toward a Pseudomonas aeruginosa-induced acute pneumonia model.

Effect of Arginase Inhibition on Pulmonary L-Arginine Metabolism in Murine Pseudomonas Pneumonia

RationaleInfection of the lung with Pseudomonas aeruginosa results in upregulation of nitric oxide synthases (NOS) and arginase expression, and both enzymes compete for L-arginine as substrate. Nitric oxide (NO) production may be regulated by arginase as it controls L-arginine availability for NOS. We here studied the effect of systemic arginase inhibition on pulmonary L-arginine metabolism in Pseudomonas pneumonia in the mouse.MethodsMice (C57BL/6, 8–10 weeks old, female) underwent direct tracheal instillation of Pseudomonas (PAO-1)-coated agar beads and were treated by repeated intra-peritoneal injections of the arginase inhibitor 2(S)-amino-6-boronohexanoic acid (ABH) or PBS until lungs were harvested on day 3 of the infection. L-arginine metabolites were quantified using liquid chromatography-tandem mass spectrometry, NO metabolites nitrate and nitrite by Griess reagent and cytokines by ELISA.ResultsNO metabolite concentrations (48.5±2.9 vs. 10.9±2.3 µM, p<0.0001), as well as L-ornithine (29.6±1.7 vs 2.3±0.4 µM, p<0.0001), the product of arginase activity, were increased in Pseudomonas infected lungs compared to naïve controls. Concentrations of the NOS inhibitor asymmetric dimethylarginine (ADMA) were also increased (0.44±0.02 vs. 0.16±0.01 µM, p<0.0001). Arginase inhibition in the infected animals resulted in a significant decrease in L-ornithine (14.6±1.6 µM, p<0.0001) but increase in L-arginine concentration (p<0.001), L-arginine/ADMA ratio (p<0.001), L-arginine availability for NOS (p<0.001), and NO metabolite concentrations (67.3±5.7 µM, p<0.05). Arginase inhibitor treatment also resulted in an increase in NO metabolite levels in animals following intratracheal injection of LPS (p = 0.015). Arginase inhibition was not associated with an increase in inflammatory markers (IFN-γ, IL-1β, IL-6, MIP-2, KC or TNF-α) in lung. Concentrations of the L-ornithine-dependent polyamines putrescine, spermidine and spermine were increased in Pseudomonas infected lungs (p<0.001, respectively) but were unaffected by ABH treatment.ConclusionsSystemic arginase inhibition with ABH during Pseudomonas pneumonia in mice results in an increase in pulmonary NO formation but no pro-inflammatory effect.

Multitracer stable isotope quantification of arginase and nitric oxide synthase activity in a mouse model of pseudomonas lung infection.

Cystic fibrosis airways are deficient for L-arginine, a substrate for nitric oxide synthases (NOSs) and arginases. The rationale for this study was to quantify NOS and arginase activity in the mouse lung. Anesthetized unventilated mice received a primed constant stable isotope intravenous infusion containing labeled L-arginine, ornithine, and citrulline. The isotopic enrichment of each of the infused isotopomers and its product amino acids were measured in plasma and organ homogenates using liquid chromatography-tandem mass spectrometry. The effect of infection was studied three days after direct tracheal instillation of Pseudomonas-coated agar beads. In the infusion model, lung infection resulted in a significant (28-fold) increase in NOS activity in lung but not in trachea, kidney, liver, or plasma. Absolute rates of arginase activity in solid tissues could not be calculated in this model. In an isolated lung perfusion model used for comparison increased NOS activity in infected lungs was confirmed (28.5-fold) and lung arginase activity was increased 9.7-fold. The activity of L-arginine metabolizing enzymes can be measured using stable isotope conversion in the mouse. Accumulation of L-ornithine in the whole mouse model hindered the exact quantification of arginase activity in the lung, a problem that was overcome utilizing an isolated lung perfusion model.

Intubation in the delivery room: Experience with nasal midazolam

BackgroundNeonates are often intubated in the delivery room (DR) without anesthesia because vascular access is impossible. AimsTo assess neonatal comfort and adverse events after use of nasal midazolam (nMDZ) for intubation in the DR. Study designProspective data collection over 6months on the intubation of neonates with respiratory distress requiring tracheal instillation of surfactant. SubjectsTwenty-seven neonates with median (Q25–75) gestational age and birthweight of, respectively, 29 (27–33)weeks and 1270 (817–1942)g received a 0.1mg/kg dose of nMDZ, and intubation was performed at the onset of tonus resolution or apnea. Outcome measuresComfort was assessed with a scale of hetero-pain assessment and electrical skin conductance monitoring. Continuous pulse oximetry was recorded in the first postnatal hour, with oscillometric blood pressure measurement every 10min. ResultsSeventy percent of the patients required a single dose, with intubation performed 4.8 (3–9)min after administration. Combined electro-clinical assessment found adequate comfort during the procedure in 68% of neonates. Mean blood pressure decreased from 39 (34–44)mmHg before to 31 (25–33)mmHg 1h following nMDZ (p=0.011). ConclusionnMDZ provided rapid and effective sedation to intubate neonates in the DR but potentially exposed them to hypotension, thus requiring close hemodynamic monitoring.

18:1/18:1-Dioleoyl-phosphatidylglycerol prevents alveolar epithelial apoptosis and profibrotic stimulus in a neonatal piglet model of acute respiratory distress syndrome

Background18:1/18:1-Dioleoyl-phosphatidylgycerol (DOPG) is a surfactant phospholipid that is nearly non-detectable in neonatal surfactant films. When alveolar macrophages are exposed to DOPG in vitro, secretory phospholipase A2 (sPLA2) production is blocked, resulting in suppressed macrophage activity and improved surfactant function. We investigated whether the addition of DOPG to a commercially available surfactant preparation would improve lung function in a neonatal piglet model of acute respiratory distress syndrome. Materials and methodsRespiratory failure was achieved by triple-hit lung injury (repeated broncho-alveolar lavage, injurious ventilation, tracheal lipopolysaccharide instillation, each intervention 24 h apart) in twenty-four domestic piglets aged 2–6 days and subject to mechanical ventilation. Following each lung injury protocol the piglets were treated with surfactant alone or with surfactant + DOPG. ResultsWithin 72 h of mechanical ventilation, we observed significantly improved gas exchange (oxygenation and ventilation), lung mechanics (compliance and resistance of the respiratory system), and pulmonary oedema (extra-vascular lung water index) in the surfactant + DOPG group. This favourable clinical effect could be attributed to improved surfactant function, reduced sPLA2 secretion, inhibition of macrophage migration, reduced alveolar epithelial apoptosis, and suppression of amphiregulin and TGF-β1 expression in pulmonary tissues as a prerequisite for fibrous lung repair. ConclusionsWe conclude that surfactant fortified by DOPG preserves lung function, and prevents alveolar epithelial injury and fibrous stimulus by reduction of sPLA2 in a neonatal model of acute respiratory distress syndrome without any relevant discernable side effects. Hence, DOPG supplementation in a neonatal lung exerts important function protecting effects and seems to be justified in cases of overwhelming pulmonary inflammation.

Comparison of Nasal Continuous Positive Airway Pressure Therapy with and without Prophylactic Surfactant in Preterm Neonates

Background: Preterm labor is at risk for respiratory distress syndrome (RDS) and sometimes requires to mechanical ventilation (MV) and surfactant therapy. Continuous positive airway pressure (CPAP) and Nasal Continuous positive airway pressure (NCPAP) are the methods of respiratory support especially for using in RDS of neonates. In other method surfactant is administered to babies via tracheal instillation. The aim of study was comparing nasal continuous positive airway pressure (NCPAP) with and without prophylactic surfactant therapy in preterm neonates’ RDS. Patients and methods: This is a randomized clinical trial study that performed on eighty newborns (28-34 weeks) who were born in Ali-Ebne-Abitaleb Hospital of Zahedan University of Medical Sciences from October 2008 to September 2010. Subjects were randomly classified in two groups whom received nasal CPAP alone or with surfactant (40 patients in each group).The Including criteria for entrance to research were approved suggested RDS, gestational age less than 34 weeks and neonates 72 hours after delivery . The data collected through direct observation and questionnaire contained fourteen items. Groups received either surfactant (Curosurf, Parma, Italy) with NCPAP or NCPAP alone. The data analyzed were conducted through SPSS 17.00 version and followed by Crosstab (Pearson Chi-square). Results: Results revealed that six neonates who received only NCPAP and four patients who received NCAPA plus surfactant required to MACHANICAL VENTILATION (MV) therapy. After a week, 33 neonates who received only NCPAP and 36 patients who received CPAP plus prophylactic surfactant remained alive. There was no significant difference between the groups regarding adverse outcomes (P=0.518). Conclusions: According to the results of this study, NCPAP is affordable as a safe protocol for RDS in preterm neonates. Further research especially with control or placebo groups is required to clarify and validate our findings.

Biophysical determinants of alveolar epithelial plasma membrane wounding associated with mechanical ventilation

Mechanical ventilation may cause harm by straining lungs at a time they are particularly prone to injury from deforming stress. The objective of this study was to define the relative contributions of alveolar overdistension and cyclic recruitment and "collapse" of unstable lung units to membrane wounding of alveolar epithelial cells. We measured the interactive effects of tidal volume (VT), transpulmonary pressure (PTP), and of airspace liquid on the number of alveolar epithelial cells with plasma membrane wounds in ex vivo mechanically ventilated rat lungs. Plasma membrane integrity was assessed by propidium iodide (PI) exclusion in confocal images of subpleural alveoli. Cyclic inflations of normal lungs from zero end-expiratory pressure to 40 cmH2O produced VT values of 56.9 ± 3.1 ml/kg and were associated with 0.12 ± 0.12 PI-positive cells/alveolus. A preceding tracheal instillation of normal saline (3 ml) reduced VT to 49.1 ± 6 ml/kg but was associated with a significantly greater number of wounded alveolar epithelial cells (0.52 ± 0.16 cells/alveolus; P < 0.01). Mechanical ventilation of completely saline-filled lungs with saline (VT = 52 ml/kg) to pressures between 10 and 15 cmH2O was associated with the least number of wounded epithelial cells (0.02 ± 0.02 cells/alveolus; P < 0.01). In mechanically ventilated, partially saline-filled lungs, the number of wounded cells increased substantially with VT, but, once VT was accounted for, wounding was independent of maximal PTP. We found that interfacial stress associated with the generation and destruction of liquid bridges in airspaces is the primary biophysical cell injury mechanism in mechanically ventilated lungs.

Acute Effects of Cigarette Smoke Extract on Alveolar Epithelial Sodium Channel Activity and Lung Fluid Clearance

Cigarette smoke contains high levels of reactive species. Moreover, cigarette smoke can induce cellular production of oxidants. The purpose of this study was to determine the effect of cigarette smoke extract (CSE)-derived oxidants on epithelial sodium channel (ENaC) activity in alveolar type 1 (T1) and type 2 (T2) cells and to measure corresponding rates of fluid clearance in mice receiving a tracheal instillation of CSE. Single-channel patch clamp analysis of T1 and T2 cells demonstrate that CSE exposure increases ENaC activity (NPo), measured as the product of the number of channels (N) and a channels open probability (Po), from 0.17 ± 0.07 to 0.34 ± 0.10 (n = 9; P = 0.04) in T1 cells. In T2 cells, CSE increased NPo from 0.08 ± 0.03 to 0.35 ± 0.10 (n = 9; P = 0.02). In both cell types, addition of tetramethylpiperidine and glutathione attenuated CSE-induced increases in ENaC NPo. Biotinylation and cycloheximide chase assays indicate that CSE-derived ROS increases channel activity, in part, by maintaining cell surface expression of the α-ENaC subunit. In vivo studies show that tracheal instillation of CSE promoted alveolar fluid clearance after 105 minutes compared with vehicle control (n = 10/group; P < 0.05).

High tidal volume ventilation does not exacerbate acid-induced lung injury in infant rats

The impact of mechanical ventilation with high VT-low PEEP in infant rats with preinjured lungs is unknown. After tracheal instillation of saline or acid, two week old rats were ventilated with VT 7mL/kg and PEEP 5cm H2O or VT 21mL/kg and PEEP 1cm H2O for 4h. Airway resistance and the coefficient of tissue elastance, measured via low-frequency forced-oscillation technique, and quasi-static pressure-volume curves deteriorated less with high VT-low PEEP when compared with low VT-high PEEP. IL-6 concentration in bronchoalveolar lavage fluid (BALF) did not differ between all ventilated groups. Moreover, differences in BALF protein concentration and histological lung injury scores were independent of applied ventilation strategies. In contrast to experimental studies with adult rats, short-term mechanical ventilation with high VT-low PEEP is not deleterious when compared to low VT-high PEEP in both healthy and pre-injured infant rat lungs. Our results call for caution when extrapolating data from adult studies and highlight the need for age-specific animal models.

Surfactant Administration via Thin Catheter during Spontaneous Breathing: Randomized Controlled Trial in Alzahra hospital

Introduction Respiratory distress syndrome (RDS) is a current cause of morbidity in premature infants resulted from surfactant deficiency. The primary aim of this randomized study was to describe the feasibility of early administration of surfactant via a thin catheter during spontaneous breathing (TEC) and compare its outcomes with the InSurE (Intubate, Surfactant, Extubate) procedure Materials and Methods Premature infants with RDS, who were ≤ 32 weeks old and stabilized with nasal continuous positive airway pressure (nCPAP), were randomized to receive surfactant either by the TEC or InSurE technique. Tracheal instillation of 200 mg/kg Curosorf via 5-F catheter during spontaneous breathing under nCPAP was performed in the intervention group (n=38). In the InSurE group (n=40), infants were intubated, received positive pressure ventilation for 30 seconds after surfactant instillation, and placed on nCPAP immediately Results Necrotizing enterocolitis rate was significantly lower in TEC group, than InSurE group, [p<0.02, RR=0.49, CI=%95(0.39_0.62)]. But rate of other morbidities did not significant difference between two groups. Although Mechanical ventilation duration did not significant different between two groups (p=0.2), but Total CPAP duration was shorter in TEC group, significantly [P<0.01, RR=8.2, CI=%95 (-37.9_-4.8)]. Need for O2 supplement and mean of total hospital duration was also lower in TEC group, but was not significant, statistically Conclusion The TEC technique is feasible for the treatment of respiratory distress syndrome in infants with very low birth weight and decreases total nCPAP duration, significantly. But Judgment of substitution TEC instead of InSurE as a routine technique need to more studies.