Nikolaos A. Maniatis

Endothelial pathomechanisms in acute lung injury

Abstract Acute lung injury (ALI) and its most severe extreme the acute respiratory distress syndrome (ARDS) refer to increased-permeability pulmonary edema caused by a variety of pulmonary or systemic insults. ALI and in particular ARDS, are usually accompanied by refractory hypoxemia and the need for mechanical ventilation. In most cases, an exaggerated inflammatory and pro-thrombotic reaction to an initial stimulus, such as systemic infection, elicits disruption of the alveolo-capillary membrane and vascular fluid leak. The pulmonary endothelium is a major metabolic organ promoting adequate pulmonary and systemic vascular homeostasis, and a main target of circulating cells and humoral mediators under injury; pulmonary endothelium is therefore critically involved in the pathogenesis of ALI. In this review we will discuss mechanisms of pulmonary endothelial dysfunction and edema generation in the lung with special emphasis on the interplay between the endothelium, the immune and hemostatic systems, and highlight how these principles apply in the context of defined disorders and specific insults implicated in ALI pathogenesis.

The endothelium in acute lung injury/acute respiratory distress syndrome

Purpose of reviewSince pulmonary edema from increased endothelial permeability is the hallmark of acute lung injury, a frequently encountered entity in critical care medicine, the study of endothelial responses in this setting is crucial to the development of effective endothelial-targeted treatments. Recent findingsFrom the enormous amount of research in the field of endothelial pathophysiology, we have focused on work delineating endothelial alterations elicited by noxious stimuli implicated in acute lung injury. The bulk of the material covered deals with molecular and cellular aspects of the pathogenesis, reflecting current trends in the published literature. We initially discuss pathways of endothelial dysfunction in acute lung injury and then cover the mechanisms of endothelial protection. Several experimental treatments in animal models are presented, which aid in the understanding of the disease pathogenesis and provide evidence for potentially useful therapies. SummaryMechanistic studies have delivered several interventions, which are effective in preventing and treating experimental acute lung injury and have thus provided objectives for translational studies. Some of these modalities may evolve into clinically useful tools in the treatment of this devastating illness.

Static and dynamic mechanics of the murine lung after intratracheal bleomycin

BackgroundDespite its widespread use in pulmonary fibrosis research, the bleomycin mouse model has not been thoroughly validated from a pulmonary functional standpoint using new technologies. Purpose of this study was to systematically assess the functional alterations induced in murine lungs by fibrogenic agent bleomycin and to compare the forced oscillation technique with quasi-static pressure-volume curves in mice following bleomycin exposure.MethodsSingle intratracheal injections of saline (50 μL) or bleomycin (2 mg/Kg in 50 μL saline) were administered to C57BL/6 (n = 40) and Balb/c (n = 32) mice. Injury/fibrosis score, tissue volume density (TVD), collagen content, airway resistance (RN ), tissue damping (G) and elastance coefficient (H), hysteresivity (η), and area of pressure-volume curve (PV-A) were determined after 7 and 21 days (inflammation and fibrosis stage, respectively). Statistical hypothesis testing was performed using one-way ANOVA with LSD post hoc tests.ResultsBoth C57BL/6 and Balb/c mice developed weight loss and lung inflammation after bleomycin. However, only C57BL/6 mice displayed cachexia and fibrosis, evidenced by increased fibrosis score, TVD, and collagen. At day 7, PV-A increased significantly and G and H non-significantly in bleomycin-exposed C57BL/6 mice compared to saline controls and further increase in all parameters was documented at day 21. G and H, but not PV-A, correlated well with the presence of fibrosis based on histology, TVD and collagen. In Balb/c mice, no change in collagen content, histology score, TVD, H and G was noted following bleomycin exposure, yet PV-A increased significantly compared to saline controls.ConclusionsLung dysfunction in the bleomycin model is more pronounced during the fibrosis stage rather than the inflammation stage. Forced oscillation mechanics are accurate indicators of experimental bleomycin-induced lung fibrosis. Quasi-static PV-curves may be more sensitive than forced oscillations at detecting inflammation and fibrosis.

Pretreatment with atorvastatin attenuates lung injury caused by high-stretch mechanical ventilation in an isolated rabbit lung model.

Objective:We hypothesized that pretreatment with atorvastatin improves alveolar capillary permeability and hemodynamics and, thus, confers protection against lung injury caused by high-stretch mechanical ventilation. Methods:Twenty-four isolated sets of normal rabbit lungs were utilized. Treated animals received atorvastatin (20 mg/kg body weight/day by mouth) for 3 days before surgery. Lungs were perfused constantly (300 mL/min) and ventilated for 1 hr with pressure-control ventilation at either 23 (high pressure; resulting in tidal volume approximately 22 mL/kg) or 11 (low pressure; tidal volume approximately 10 mL/kg) cm H2O peak inspiratory pressure and positive end-expiratory pressure of 3 cm H2O. Four groups were examined: high pressure–no statin, high pressure–statin pretreatment, low pressure–no statin, and low pressure–statin pretreatment. Results:The high-pressure–no statin group sustained more damage than the low-pressure groups. In high-pressure groups, lungs of statin-pretreated vs. no statin-pretreated animals sustained a significantly lower increase in ultrafiltration coefficient (an accurate marker of alveolar capillary permeability; high-pressure–statin pretreatment vs. high-pressure–no statin, −0.013 ± 0.017 g/min/mm Hg/100g vs. 1.723 ± 0.495 g/min/mm Hg/100g; p < .001), lower weight gain (i.e., less edema formation; 4.62 ± 1.50 grams vs. 17.75 ± 4.71 grams; p = .005), improved hemodynamics (i.e., lower increase in mean pulmonary artery pressure; 0.56 ± 0.51 mm Hg vs. 5.62 ± 1.52 mm Hg; p = .04), lower protein concentration in bronchoalveolar lavage fluid (p < .001), and fewer histologic lesions (p = .013). Apoptosis of lung parenchyma cells was not different (p = .97). There was no difference between low-pressure–statin pretreatment and low-pressure–no statin groups regarding these outcomes. Conclusion:In this model, atorvastatin improves alveolar capillary permeability and hemodynamics and, thus, attenuates lung injury caused by high-stretch mechanical ventilation.

Caveolins and Lung Function

The primary function of the mammalian lung is to facilitate diffusion of oxygen to venous blood and to ventilate carbon dioxide produced by catabolic reactions within cells. However, it is also responsible for a variety of other important functions, including host defense and production of vasoactive agents to regulate not only systemic blood pressure, but also water, electrolyte and acid-base balance. Caveolin-1 is highly expressed in the majority of cell types in the lung, including epithelial, endothelial, smooth muscle, connective tissue cells, and alveolar macrophages. Deletion of caveolin-1 in these cells results in major functional aberrations, suggesting that caveolin-1 may be crucial to lung homeostasis and development. Furthermore, generation of mutant mice that under-express caveolin-1 results in severe functional distortion with phenotypes covering practically the entire spectrum of known lung diseases, including pulmonary hypertension, fibrosis, increased endothelial permeability, and immune defects. In this Chapter, we outline the current state of knowledge regarding caveolin-1-dependent regulation of pulmonary cell functions and discuss recent research findings on the role of caveolin-1 in various pulmonary disease states, including obstructive and fibrotic pulmonary vascular and inflammatory diseases.

Inhaled activated protein C protects mice from ventilator-induced lung injury

IntroductionActivated Protein C (APC), an endogenous anticoagulant, improves tissue microperfusion and endothelial cell survival in systemic inflammatory states such as sepsis, but intravenous administration may cause severe bleeding. We have thus addressed the role of APC delivered locally by inhalation in preventing acute lung injury from alveolar overdistention and the subsequent ventilator-induced lung injury (VILI). We also assessed the effects of APC on the activation status of Extracellular- Regulated Kinase 1/2 (ERK) pathway, which has been shown to be involved in regulating pulmonary responses to mechanical stretch.MethodsInhaled APC (12.5 μg drotrecogin-α × 4 doses) or saline was given to tracheotomized C57/Bl6 mice starting 20 min prior to initiation of injurious mechanical ventilation with tidal volume 25 mL/Kg for 4 hours and then hourly thereafter; control groups receiving inhaled saline were ventilated with 8 mL/Kg for 30 min or 4 hr. We measured lung function (respiratory system elastance H), arterial blood gases, surrogates of vascular leak (broncho-alveolar lavage (BAL) total protein and angiotensin-converting enzyme (ACE)-activity), and parameters of inflammation (BAL neutrophils and lung tissue myeloperoxidase (MPO) activity). Morphological alterations induced by mechanical ventilation were examined in hematoxylin-eosin lung tissue sections. The activation status of ERK was probed in lung tissue homogenates by immunoblotting and in paraffin sections by immunohistochemistry. The effect of APC on ERK signaling downstream of the thrombin receptor was tested on A549 human lung epithelial cells by immunoblotting. Statistical analyses were performed using ANOVA with appropriate post-hoc testing.ResultsIn mice subjected to VILI without APC, we observed hypoxemia, increased respiratory system elastance and inflammation, assessed by BAL neutrophil counts and tissue MPO activity. BAL total protein levels and ACE activity were also elevated by VILI, indicating compromise of the alveolo-capillary barrier. In addition to preserving lung function, inhaled APC prevented endothelial barrier disruption and attenuated hypoxemia and the inflammatory response. Mechanistically, we found a strong activation of ERK in lung tissues by VILI, which was prevented by APC, suggestive of pathogenetic involvement of the Mitogen-Activated Kinase pathway. In cultured human lung epithelial cells challenged by thrombin, APC abrogated the activation of ERK and its downstream effector, cytosolic Phospholipase A2.ConclusionsTopical application of APC by inhalation may effectively reduce lung injury induced by mechanical ventilation in mice.

A critical role for gelsolin in ventilator-induced lung injury.

Mechanical ventilation, an essential life-support modality of patients with acute lung injury (ALI) or the acute respiratory distress syndrome (ARDS), exerts its detrimental effects through largely unknown mechanisms. Gelsolin (GSN), an actin-binding protein and a substrate of caspase-3, was recently shown to play a major role in bleomycin- or lipopolysaccharide-induced lung injury. To dissect a possible role of GSN in the pathogenesis of ventilator-induced lung injury (VILI), genetically modified mice lacking GSN expression and wild-type controls underwent mechanical ventilation with high tidal volumes. GSN was found up-regulated in the airways upon VILI, and its genetic ablation led to almost complete disease protection as manifested by reduced edema formation, reduced lung injury, attenuated epithelial apoptosis, diminished cytokine expression, and impaired neutrophil infiltration. GSN fragmentation was shown to be an effector mechanism in VILI-induced apoptosis, while GSN expression was shown to be necessary for efficient neutrophil infiltration, which was found to be a prerequisite for VILI induction in this model. Therefore, intracellular GSN and GSN-mediated responses were shown to be an important player in the pathogenesis of VILI.

Effect of clarithromycin in patients with suspected Gram-negative sepsis: results of a randomized controlled trial

BACKGROUND A previous randomized study showed that clarithromycin decreases the risk of death due to ventilator-associated pneumonia and shortens the time until infection resolution. The efficacy of clarithromycin was tested in a larger population with sepsis. METHODS Six hundred patients with systemic inflammatory response syndrome due to acute pyelonephritis, acute intra-abdominal infections or primary Gram-negative bacteraemia were enrolled in a double-blind, randomized, multicentre trial. Clarithromycin (1 g) was administered intravenously once daily for 4 days consecutively in 302 patients; another 298 patients were treated with placebo. Mortality was the primary outcome; resolution of infection and hospitalization costs were the secondary outcomes. RESULTS The groups were well matched for demographics, disease severity, microbiology and appropriateness of the administered antimicrobials. Overall 28 day mortality was 17.1% (51 deaths) in the placebo arm and 18.5% (56 deaths) in the clarithromycin arm (P = 0.671). Nineteen out of 26 placebo-treated patients with septic shock and multiple organ dysfunctions died (73.1%) compared with 15 out of 28 clarithromycin-treated patients (53.6%, P = 0.020). The median time until resolution of infection was 5 days in both arms. In the subgroup with severe sepsis/shock, this was 10 days in the placebo arm and 6 days in the clarithromycin arm (P = 0.037). The cost of hospitalization was lower after treatment with clarithromycin (P = 0.044). Serious adverse events were observed in 1.3% and 0.7% of placebo- and clarithromycin-treated patients, respectively (P = 0.502). CONCLUSIONS Intravenous clarithromycin did not affect overall mortality; however, administration shortened the time to resolution of infection and decreased the hospitalization costs.

Secreted phosphoprotein-1 directly provokes vascular leakage to foster malignant pleural effusion

Secreted phosphoprotein-1 (SPP1) promotes cancer cell survival and regulates tumor-associated angiogenesis and inflammation, both central to the pathogenesis of malignant pleural effusion (MPE). Here, we examined the impact of tumor- and host-derived SPP1 in MPE formation and explored the mechanisms by which the cytokine exerts its effects. We used a syngeneic murine model of lung adenocarcinoma-induced MPE. To dissect the effects of tumor- versus host-derived SPP1, we intrapleurally injected wild-type and SPP1-knockout C57/BL/6 mice with either wild-type or SPP1-deficient syngeneic lung cancer cells. We demonstrated that both tumor- and host-derived SPP1 promoted pleural fluid accumulation and tumor dissemination in a synergistic manner (P<0.001). SPP1 of host origin elicited macrophage recruitment into the cancer-affected pleural cavity and boosted tumor angiogenesis, whereas tumor-derived SPP1 curtailed cancer cell apoptosis in vivo. Moreover, the cytokine directly promoted vascular hyper-permeability independently of vascular endothelial growth factor. In addition, SPP1 of tumor and host origin differentially affected the expression of proinflammatory and angiogenic mediators in the tumor microenvironment. These results suggest that SPP1 of tumor and host origin impact distinct aspects of MPE pathobiology to synergistically promote pleural fluid formation and pleural tumor progression. SPP1 may present an attractive target of therapeutic interventions for patients with MPE.

Interstitial cortisol obtained by microdialysis in mechanically ventilated septic patients: Correlations with total and free serum cortisol

PURPOSE The aim of this study was to measure subcutaneous tissue cortisol obtained by microdialysis (MD) in 35 mechanically ventilated septic patients. MATERIALS AND METHODS Upon intensive care unit admission, an MD catheter was inserted into the subcutaneous tissue of the thigh. Cortisol (CORT) was determined in a 5:00 to 9:00 am microdialysate sample collected within 72 hours. Concurrently, serum total (T-CORT) and free CORT (F-CORT) were measured. The Acute Physiology and Chronic Health Evaluation (APACHE II) and Sequential Organ Failure Assessment scores were calculated. Both T-CORT less than 10 μg/dL and F-CORT less than 0.8 μg/dL were considered as indicating critical illness-related corticosteroid insufficiency. Adrenal adequacy was defined as T-CORT greater than 20 μg/dL or F-CORT greater than 2.0 μg/dL. RESULTS Total CORT correlated significantly with F-CORT (rs = +0.8, P < .0001). Microdialysis CORT had a lower correlation with T-CORT (rs = +0.6, P < .0001) and F-CORT (rs = +0.7, P < .0001) and a weak correlation with APACHE II score (rs = +0.4, P < .01). On the basis of MD-CORT, the patients were divided in quartiles. Although the median F-CORT and T-CORT levels were significantly different (P < .001) among the MD-CORT quartiles, there was a considerable overlap between the subgroups. All patients with T-CORT less than 10 μg/dL and all but 3 patients with F-CORT less than 0.8 μg/dL had tissue CORT in the lower quartile. However, only 50% and 58% of patients with adequate T-CORT and F-CORT levels, respectively, had concordant MD-CORT in the highest quartile. CONCLUSIONS Microdialysis CORT levels correlate moderately with circulating T-CORT and F-CORT. Of note, several patients presented with discrepant measurements between interstitial and circulating CORT concentrations. Thus, interstitial CORT measurements represent an additional tool to investigate the tissue CORT availability in critically ill patients.