Limiting Overdistention or Collapse when Mechanically Ventilating Injured Lungs: A Randomized Study in a Porcine Model.

RATIONALE: It is unknown whether preventing overdistention or collapse is more important when titrating positive end-expiratory pressure (PEEP) in acute respiratory distress syndrome (ARDS). OBJECTIVE: To compare PEEP targeting either minimal overdistention, minimal collapse or using a compromise between collapse and overdistention in a randomized trial, and assess the impact on respiratory mechanics, gas exchange, inflammation, and hemodynamics. METHODS: In a porcine model of ARDS, lung collapse and overdistention were estimated by electrical impedance tomography during a decremental PEEP titration. Pigs were randomized to three groups and ventilated for 12 hours: PEEP set at ≤3% of overdistention (low overdistention); ≤3% of collapse (low collapse); and crossing point of collapse and overdistention (crossing-point). MEASUREMENTS AND MAIN RESULTS: Thirty-six pigs (12/group) were included. Median PEEP were 7(IQR:6-8)cmH2O, 11(10-11)cmH2O, and 15(12-16)cmH2O in the three groups, p<0.001. With low overdistension, 6(50%) pigs died whereas survival was 100% in both other groups. Cause of death was hemodynamic in nature, with high transpulmonary gradient and high epinephrine requirements. Compared to the other groups, pigs surviving with low overdistension had worse respiratory mechanics and gas exchange during the entire protocol. Minimal differences existed between crossing-point and low collapse animals in physiological parameters but postmortem alveolar density was more homogeneous in crossing-point. Inflammatory markers were not significantly different. CONCLUSIONS: PEEP to minimize overdistention resulted in high mortality in an animal model of ARDS. Minimizing collapse or choosing a compromise between collapse and overdistention may result in less lung injury, with potential benefits of the compromise approach.

Therapeutic stem cell-derived alveolar-like macrophages display bactericidal effects and resolve Pseudomonas aeruginosa-induced lung injury.

Bacterial lung infections lead to greater than 4 million deaths per year with antibiotic treatments driving an increase in antibiotic resistance and a need to establish new therapeutic approaches. Recently, we have generated mouse and rat stem cell-derived alveolar-like macrophages (ALMs), which like primary alveolar macrophages (1'AMs), phagocytose bacteria and promote airway repair. Our aim was to further characterize ALMs and determine their bactericidal capabilities. The characterization of ALMs showed that they share known 1'AM cell surface markers, but unlike 1'AMs are highly proliferative in vitro. ALMs effectively phagocytose and kill laboratory strains of P. aeruginosa (P.A.), E. coli (E.C.) and S. aureus, and clinical strains of P.A. In vivo, ALMs remain viable, adapt additional features of native 1'AMs, but proliferation is reduced. Mouse ALMs phagocytose P.A. and E.C. and rat ALMs phagocytose and kill P.A. within the lung 24 h post-instillation. In a pre-clinical model of P.A.-induced lung injury, rat ALM administration mitigated weight loss and resolved lung injury observed seven days post-instillation. Collectively, ALMs attenuate pulmonary bacterial infections and promote airway repair. ALMs could be utilized as an alternative or adjuvant therapy where current treatments are ineffective against antibiotic-resistant bacteria or to enhance routine antibiotic delivery.

Repeated endo-tracheal tube disconnection generates pulmonary edema in a model of volume overload: an experimental study.

BACKGROUND: An abrupt lung deflation in rodents results in lung injury through vascular mechanisms. Ventilator disconnections during endo-tracheal suctioning in humans often cause cardio-respiratory instability. Whether repeated disconnections or lung deflations cause lung injury or oedema is not known and was tested here in a porcine large animal model. METHODS: Yorkshire pigs (~ 12 weeks) were studied in three series. First, we compared PEEP abruptly deflated from 26 cmH2O or from PEEP 5 cmH2O to zero. Second, pigs were randomly crossed over to receive rapid versus gradual PEEP removal from 20 cmH2O. Third, pigs with relative volume overload, were ventilated with PEEP 15 cmH2O and randomized to repeated ETT disconnections (15 s every 15 min) or no disconnection for 3 h. Hemodynamics, pulmonary variables were monitored, and lung histology and bronchoalveolar lavage studied. RESULTS: As compared to PEEP 5 cmH2O, abrupt deflation from PEEP 26 cmH2O increased PVR, lowered oxygenation, and increased lung wet-to-dry ratio. From PEEP 20 cmH2O, gradual versus abrupt deflation mitigated the changes in oxygenation and vascular resistance. From PEEP 15, repeated disconnections in presence of fluid loading led to reduced compliance, lower oxygenation, higher pulmonary artery pressure, higher lung wet-to-dry ratio, higher lung injury score and increased oedema on morphometry, compared to no disconnects. CONCLUSION: Single abrupt deflation from high PEEP, and repeated short deflations from moderate PEEP cause pulmonary oedema, impaired oxygenation, and increased PVR, in this large animal model, thus replicating our previous finding from rodents. Rapid deflation may thus be a clinically relevant cause of impaired lung function, which may be attenuated by gradual pressure release.

Alveolar-like Macrophages Attenuate Respiratory Syncytial Virus Infection.

Respiratory Syncytial Virus (RSV) is the leading cause of acute lower respiratory infections in young children and infection has been linked to the development of persistent lung disease in the form of wheezing and asthma. Despite substantial research efforts, there are no RSV vaccines currently available and an effective monoclonal antibody targeting the RSV fusion protein (palivizumab) is of limited general use given the associated expense. Therefore, the development of novel approaches to prevent RSV infection is highly desirable to improve pediatric health globally. We have developed a method to generate alveolar-like macrophages (ALMs) from pluripotent stem cells. These ALMs have shown potential to promote airway innate immunity and tissue repair and so we hypothesized that ALMs could be used as a strategy to prevent RSV infection. Here, we demonstrate that ALMs are not productively infected by RSV and prevent the infection of epithelial cells. Prevention of epithelial infection was mediated by two different mechanisms: phagocytosis of RSV particles and release of an antiviral soluble factor different from type I interferon. Furthermore, intratracheal administration of ALMs protected mice from subsequent virus-induced weight loss and decreased lung viral titres and inflammation, indicating that ALMs can impair the pathogenesis of RSV infection. Our results support a prophylactic role for ALMs in the setting of RSV infection and warrant further studies on stem cell-derived ALMs as a novel cell-based therapy for pulmonary viral infections.

Does lack of glutathione peroxidase 1 gene expression exacerbate lung injury induced by neonatal hyperoxia in mice?

Supplemental oxygen (O2) increases the risk of lung injury in preterm infants, owing to an immature antioxidant system. Our objective was to determine whether impairing antioxidant defense by decreasing glutathione peroxidase 1 (GPx1) gene expression increases the injurious effects of hyperoxia (Hyp). GPx1+/+ and GPx1-/- C57Bl/6J mice were exposed to 21% O2 (Air) or 40% O2 (Hyp) from birth to postnatal day 7 (P7d); they were euthanized on P7d or maintained in air until adulthood [postnatal day 56 (P56d)] to assess short-term and long-term effects, respectively. We assessed lung architecture, three markers of pulmonary oxidative stress (P7d, P56d), macrophages in lung tissue (P7d), immune cells in bronchoalveolar lavage fluid (BALF; P56d), and GPx1-4 and catalase gene expression in lung tissue (P7d, P56d). On P7d, macrophages were decreased by lack of GPx1 expression and further decreased by hyperoxia. GPx1 expression was increased in GPx1+/+Hyp mice and decreased in both GPx1-/- groups. On P56d, heme oxygenase-1 was increased by hyperoxia when GPx1 was absent. There were significantly more immune cells from Hyp groups than from the GPx1+/+Air group and a greater proportion of lymphocytes in GPx1-/-Hyp mice. GPx1 expression was significantly decreased in GPx1-/- mice; GPx2-4 and catalase expression was increased in GPx1-/-Hyp mice compared with other groups. Tissue fraction was decreased in GPx1-/-Air mice; bronchiolar smooth muscle was decreased in GPx1-/- mice. GPx1 does not clearly exacerbate hyperoxia-induced increases in oxidative stress or lung injury but may alter pulmonary immune function. Increased expression of GPx2-4 and catalase in GPx1-/-Hyp mice suggests gene redundancy within the model.

Early Postnatal Hyperoxia in Mice Leads to Severe Persistent Vitreoretinopathy.

Purpose: To describe a mouse model of hyperoxia-induced vitreoretinopathy that replicated some of the clinical and pathologic features encountered in infants with severe retinopathy of prematurity and congenital ocular conditions such as persistent hyperplastic primary vitreous. Methods: Experimental mice (C57BL/6J) were exposed to 65% oxygen between postnatal days (P)0 to P7 and studied at P10, P14, and 3, 5, 8, 20, and 40 weeks. Controls were exposed to normoxic conditions. Fundus imaging and fluorescein angiography were performed at all time points, and spectral-domain optical coherence tomography (SD-OCT) and electroretinography were performed at 8- and 20-week time points. Eyes were processed for resin histology, frozen sections, and retinal whole mounts. Immunostaining was performed to visualize vasculature isolectin B4 (Ib4), collagen type IV, glial fibrillary acidic protein, and α-smooth muscle actin. Results: Early exposure to hyperoxia resulted in bilateral vitreous hemorrhages at 3 weeks. From 5 weeks onward there were extensive zones of retinal degeneration, scarring or gliosis, retinal folding, and detachments caused by traction of α-smooth muscle actin-positive vitreous membranes. Tortuous retinal vessels, together with hyperplastic and persistence of hyaloid vessels are evident into adulthood. In the early stages (P10-3 weeks), branches from the tunica vasculosa lentis (TVL) supplied the marginal retina until retinal vessels were established. The peripheral retina remained poorly vascularized into adulthood. Electroretinography revealed 50% to 60% diminution in retinal function in adult mice that strongly correlated with vitreal changes identified using SD-OCT. Conclusions: This animal model displays a mixture of vitreoretinal pathologic changes that persist into adulthood. The model may prove valuable in experimental investigations of therapeutic approaches to blinding conditions caused by vitreous and retinal abnormalities.

Impact of Dietary Tomato Juice on Changes in Pulmonary Oxidative Stress, Inflammation and Structure Induced by Neonatal Hyperoxia in Mice (Mus musculus).

Many preterm infants require hyperoxic gas for survival, although it can contribute to lung injury. Experimentally, neonatal hyperoxia leads to persistent alterations in lung structure and increases leukocytes in bronchoalveolar lavage fluid (BALF). These effects of hyperoxia on the lungs are considered to be caused, at least in part, by increased oxidative stress. Our objective was to determine if dietary supplementation with a known source of antioxidants (tomato juice, TJ) could protect the developing lung from injury caused by breathing hyperoxic gas. Neonatal mice (C57BL6/J) breathed either 65% O2 (hyperoxia) or room air from birth until postnatal day 7 (P7d); some underwent necropsy at P7d and others were raised in room air until adulthood (P56d). In subsets of both groups, drinking water was replaced with TJ (diluted 50:50 in water) from late gestation to necropsy. At P7d and P56d, we analyzed total antioxidant capacity (TAC), markers of oxidative stress (nitrotyrosine and heme oxygenase-1 expression), inflammation (interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α) expression), collagen (COL) and smooth muscle in the lungs; we also assessed lung structure. We quantified macrophages in lung tissue (at P7d) and leukocytes in BALF (at P56d). At P7d, TJ increased pulmonary TAC and COL1α1 expression and attenuated the hyperoxia-induced increase in nitrotyrosine and macrophage influx; however, changes in lung structure were not affected. At P56d, TJ increased TAC, decreased oxidative stress and reversed the hyperoxia-induced increase in bronchiolar smooth muscle. Additionally, TJ alone decreased IL-1ß expression, but following hyperoxia TJ increased TNF-α expression and did not alter the hyperoxia-induced increase in leukocyte number. We conclude that TJ supplementation during and after neonatal exposure to hyperoxia protects the lung from some but not all aspects of hyperoxia-induced injury, but may also have adverse side-effects. The effects of TJ are likely due to elevation of circulating antioxidant concentrations.

Neonatal exposure to mild hyperoxia causes persistent increases in oxidative stress and immune cells in the lungs of mice without altering lung structure.

Preterm infants often require supplemental oxygen due to lung immaturity, but hyperoxia can contribute to an increased risk of respiratory illness later in life. Our aim was to compare the effects of mild and moderate levels of neonatal hyperoxia on markers of pulmonary oxidative stress and inflammation and on lung architecture; both immediate and persistent effects were assessed. Neonatal mice (C57BL6/J) were raised in either room air (21% O2), mild (40% O2), or moderate (65% O2) hyperoxia from birth until postnatal day 7 (P7d). The mice were killed at either P7d (immediate effects) or lived in air until adulthood (P56d, persistent effects). We enumerated macrophages in lung tissue at P7d and immune cells in bronchoalveolar lavage fluid (BALF) at P56d. At P7d and P56d, we assessed pulmonary oxidative stress [heme oxygenase-1 (HO-1) and nitrotyrosine staining] and lung architecture. The data were interrogated for sex differences. At P7d, HO-1 gene expression was greater in the 65% O2 group than in the 21% O2 group. At P56d, the area of nitrotyrosine staining and number of immune cells were greater in the 40% O2 and 65% O2 groups relative to the 21% O2 group. Exposure to 65% O2, but not 40% O2, led to larger alveoli and lower tissue fraction in the short term and to persistently fewer bronchiolar-alveolar attachments. Exposure to 40% O2 or 65% O2 causes persistent increases in pulmonary oxidative stress and immune cells, suggesting chronic inflammation within the adult lung. Unlike 65% O2, 40% O2 does not affect lung architecture.