A two-hit model of sepsis plus hyperoxia causes lung permeability and inflammation

Mouse models of acute lung injury (ALI) have been instrumental for studies of the biologic underpinnings of lung inflammation and permeability, but murine models of sepsis generate minimal lung injury. Our goal was to create a murine sepsis model of ALI that reflects the inflammation, lung edema, histologic abnormalities and physiologic dysfunction that characterize ALI. Using a cecal slurry (CS) model of polymicrobial abdominal sepsis and exposure to hyperoxia (95%), we systematically varied timing and dose of the CS injection, fluids and antibiotics and dose of hyperoxia. We found that CS alone had a high mortality rate that was improved with the addition of antibiotics and fluids. Despite this, we did not see evidence of ALI as measured by bronchoalveolar lavage (BAL) cell count, total protein, CXCL-1 or by lung wet:dry weight ratio. Addition of hyperoxia (95% FiO2) to CS immediately after CS injection increased BAL cell counts and CXCL-1 and lung wet:dry weight ratio but was associated with 40% mortality. Splitting the hyperoxia treatment into two 12 hour exposures (0-12 hours24-36 hours) after CS injection increased survival to 75% and caused significant lung injury compared to CS alone as measured by increased BAL total cell count (92500 vs 240000, p=0.0004), BAL protein (71 vs 103 ug/ml, p=0.0030, and lung wet:dry weight ratio (4.5 vs 5.5 p=0.0005), and compared to sham as measured by increased BAL CXCL-1 (20 vs 2372 pg/ml, p<0.0001), and histologic lung injury score (1.9 vs 4.2, p=0.0077). Additionally, our final model showed evidence of lung epithelial (increased BAL and plasma RAGE) and endothelial (increased Syndecan-1 and sulfated glycosaminoglycans) injury. In conclusion, we have developed a clinically relevant mouse model of sepsis-induced ALI using IP injection of CS, antibiotics and fluids, and hyperoxia. This clinically relevant model can be used for future studies of sepsis-induced ALI.

Shen-su-yin, a Traditional Herbal Medicine, Attenuates Lipopolysaccharide-induced Acute Lung Injury in Rats Through Its Anti-inflammatory and Anti-oxidative Effects

Most components of Shen-su-yin (SSY), an herbal formula, have anti-inflammatory and antioxidant activities. The present study was designed to investigate potential effects and mechanisms of SSY on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in rats. 48 rats were randomly divided into 4 groups: control (Ctrl) group, LPS-induced ALI group, low- (SSY-LD) and high- (SSY-HD) dose SSY-treated ALI group. SSY was administered to SSY-treated rats immediately after LPS induction. After 24 hours, blood gas analysis and lactate determination were performed; and bronchoalveolar lavage fluid was collected for detecting protein concentration and levels of cytokines. Lung tissues were obtained for Western blot analysis, histopathological analysis, wet-to-dry weight ratio calculation and measurement of oxidative stress levels. SSY improved oxygenation index and mean arterial pressure, decreased levels of lactate and heart rate, alleviated lung histopathology indexes including lung injury score, wet-to-dry weight ratio and exudation of protein as well as inflammatory cells in ALI rats. Furthermore, SSY reduced levels of pro-inflammatory and oxidative mediums, while increasing levels of anti-inflammatory cytokine and anti-oxidative activity in lung tissues. SSY also suppressed NF-κB signalling pathway and further activated Keap1-Nrf2-ARE signalling pathway activated by LPS. Moreover, all the effects caused by SSY in the SSY-HD group were more encouraging than those in the SSY-LD group. The results indicate that the preventive use of SSY can alleviate ALI through the anti-inflammatory and antioxidant effects mediated by inhibition of NF-κB signalling pathway and activation of Keap1-Nrf2-ARE signalling pathway, and the effect of high dose is better.

Pharmacological Effects of Marine-Derived Enterococcus faecium EA9 against Acute Lung Injury and Inflammation in Cecal Ligated and Punctured Septic Rats

Microorganisms obtained from the marine environment may represent a potential therapeutic value for multiple diseases. This study explored the possible protective role of marine-derived potential probiotic Enterococcus faecium EA9 (E. faecium) against pulmonary inflammation and oxidative stress using the cecal ligation and puncture (CLP) model of sepsis in Wistar rats. Animals were pretreated with E. faecium for 10 days before either sham or CLP surgeries. Animals were sacrificed 72 hours following the surgical intervention. The histological architecture of lung tissues was evaluated as indicated by the lung injury score. In addition, the extend of pulmonary edema was determined as wet/dry weight ratio. The inflammatory cytokines were estimated in lung tissues, including tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and interleukin-1 beta (IL-1β) using the enzyme-linked-immunosorbent-assay (ELISA) technique. Moreover, markers for lipid peroxidation such as thiobarbituric acid reaction substances (TBARs), and endogenous antioxidants, including reduced glutathione (GSH) were determined in lung tissues. Finally, the enzymatic activities of antioxidant enzymes such as catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione reductase (GR) were assayed in the lungs. Pretreatment with E. faecium markedly attenuated CLP-induced lung injury and pulmonary edema. Markers for inflammation, including TNF-α, IL-6, and IL-1β were augmented in the lung tissues of CLP animals, while E. faecium ameliorated their augmented levels. E. faecium pretreatment also restored the elevated TBARS levels and the prohibited CAT, SOD, and GPx enzymatic activities in CLP animals. GSH levels were corrected by E. faecium in CLP animals. The inflammatory and lipid peroxidation mediators were positively correlated, while antioxidant enzymatic activities were negatively correlated with CLP-induced lung injury and pulmonary edema. Collectively, marine-derived Enterococcus faecium EA9 might be considered as a prospective therapeutic tool for the management of pulmonary dysfunction associated with sepsis.

Modulation of oxidative and nitrosative stress attenuates microvascular hyperpermeability in ovine model of Pseudomonas aeruginosa sepsis

In sepsis, microvascular hyperpermeability caused by oxidative/nitrosative stress (O&NS) plays an important role in tissue edema leading to multi-organ dysfunctions and increased mortality. We hypothesized that a novel compound R-107, a modulator of O&NS, effectively ameliorates the severity of microvascular hyperpermeability and preserves multi-organ function in ovine sepsis model. Sepsis was induced in twenty-two adult female Merino sheep by intravenous infusion of Pseudomonas aeruginosa (PA) (1 × 1010 CFUs). The animals were allocated into: 1) Control (n = 13): intramuscular injection (IM) of saline; and 2) Treatment (n = 9): IM of 50 mg/kg R-107. The treatment was given after the PA injection, and monitored for 24-h. R-107 treatment significantly reduced fluid requirement (15–24 h, P < 0.05), net fluid balance (9–24 h, P < 0.05), and water content in lung/heart/kidney (P = 0.02/0.04/0.01) compared to control. R-107 treatment significantly decreased lung injury score/modified sheep SOFA score at 24-h (P = 0.01/0.04), significantly lowered arterial lactate (21–24 h, P < 0.05), shed syndecan-1 (3–6 h, P < 0.05), interleukin-6 (6–12 h, P < 0.05) levels in plasma, and significantly attenuated lung tissue 3-nitrotyrosine and vascular endothelial growth factor-A expressions (P = 0.03/0.002) compared to control. There was no adverse effect in R-107 treatment. In conclusion, modulation of O&NS by R-107 reduced hyperpermeability markers and improved multi-organ function.

Modulation of Oxidative and Nitrosative Stress Attenuates Microvascular Hyperpermeability in Ovine Model of Pseudomonas Aeruginosa Sepsis

In sepsis, microvascular hyperpermeability caused by oxidative/nitrosative stress (O&NS) plays an important role in tissue edema leading to multi-organ dysfunctions and increased mortality. We hypothesized that a novel compound R-107, a modulator of O&NS, effectively ameliorates the severity of microvascular hyperpermeability and preserves multi-organ function in ovine sepsis model. Sepsis was induced in twenty-two adult female Merino sheep by intravenous infusion of Pseudomonas aeruginosa (PA) (1x1010 CFUs). The animals were allocated into: 1) Control (n = 13): intramuscular injection (IM) of saline; and 2) Treatment (n = 9): IM of 50 mg/kg R-107. The treatment was given after the PA injection, and monitored for 24-hour. R-107 treatment significantly reduced fluid requirement (15–24 hours, p < 0.05), net fluid balance (9–24 hours, p < 0.05), and water content in lung/heart/kidney (p = 0.02/0.04/0.01) compared to control. R-107 treatment significantly decreased lung injury score/modified sheep SOFA score at 24-hour (p = 0.01/0.04), significantly lowered arterial lactate (21–24 hours, p < 0.05), shed syndecan-1 (3–6 hours, p < 0.05), interleukin-6 (6–12 hours, p < 0.05) levels in plasma, and significantly attenuated lung tissue 3-nitrotyrosine and vascular endothelial growth factor-A expressions (p = 0.03/0.002) compared to control. There was no adverse effect in R-107 treatment. In conclusion, modulation of O&NS by R-107 reduced hyperpermeability markers and improved multi-organ function.

Obesity Attenuates Ventilator-Induced Lung Injury by Modulating the STAT3–SOCS3 Pathway

BackgroundVentilator-induced lung injury (VILI) is characterized by vascular barrier dysfunction and suppression of alveolar fluid clearance (AFC). Obesity itself leads to chronic inflammation, which may initiate an injurious cascade to the lungs and simultaneously induce a protective feedback. In this study, we investigated the protective mechanism of obesity on VILI in a mouse model.MethodsThe VILI model was set up via 6-h mechanical ventilation with a high tidal volume. Parameters including lung injury score, STAT3/NFκB pathway, and AFC were assessed. Mice with diet-induced obesity were obtained by allowing free access to a high-fat diet since the age of 3 weeks. After a 9-week diet intervention, these mice were sacrificed at the age of 12 weeks. The manipulation of SOCS3 protein was achieved by siRNA knockdown and pharmaceutical stimulation using hesperetin. WNK4 knockin and knockout obese mice were used to clarify the pathway of AFC modulation.ResultsObesity itself attenuated VILI. Knockdown of SOCS3 in obese mice offset the protection against VILI afforded by obesity. Hesperetin stimulated SOCS3 upregulation in nonobese mice and provided protection against VILI. In obese mice, the WNK4 axis was upregulated at the baseline, but was significantly attenuated after VILI compared with nonobese mice. At the baseline, the manipulation of SOCS3 by siRNA and hesperetin also led to the corresponding alteration of WNK4, albeit to a lesser extent. After VILI, WNK4 expression correlated with STAT3/NFκB activation, regardless of SOCS3 status. Obese mice carrying WNK4 knockout had VILI with a severity similar to that of wild-type obese mice. The severity of VILI in WNK4-knockin obese mice was counteracted by obesity, similar to that of wild-type nonobese mice only.ConclusionsObesity protects lungs from VILI by upregulating SOCS3, thus suppressing the STAT3/NFκB inflammatory pathway and enhancing WNK4-related AFC. However, WNK4 activation is mainly from direct NFκB downstreaming, and less from SOCS3 upregulation. Moreover, JAK2–STAT3/NFκB signaling predominates the pathogenesis of VILI. Nevertheless, the interaction between SOCS3 and WNK4 in modulating VILI in obesity warrants further investigation.

Early high-dose vitamin C in post-cardiac arrest syndrome (VITaCCA): study protocol for a randomized, double-blind, multi-center, placebo-controlled trial

Background High-dose intravenous vitamin C directly scavenges and decreases the production of harmful reactive oxygen species (ROS) generated during ischemia/reperfusion after a cardiac arrest. The aim of this study is to investigate whether short-term treatment with a supplementary or very high-dose intravenous vitamin C reduces organ failure in post-cardiac arrest patients. Methods This is a double-blind, multi-center, randomized placebo-controlled trial conducted in 7 intensive care units (ICUs) in The Netherlands. A total of 270 patients with cardiac arrest and return of spontaneous circulation will be randomly assigned to three groups of 90 patients (1:1:1 ratio, stratified by site and age). Patients will intravenously receive a placebo, a supplementation dose of 3 g of vitamin C or a pharmacological dose of 10 g of vitamin C per day for 96 h. The primary endpoint is organ failure at 96 h as measured by the Resuscitation-Sequential Organ Failure Assessment (R-SOFA) score at 96 h minus the baseline score (delta R-SOFA). Secondary endpoints are a neurological outcome, mortality, length of ICU and hospital stay, myocardial injury, vasopressor support, lung injury score, ventilator-free days, renal function, ICU-acquired weakness, delirium, oxidative stress parameters, and plasma vitamin C concentrations. Discussion Vitamin C supplementation is safe and preclinical studies have shown beneficial effects of high-dose IV vitamin C in cardiac arrest models. This is the first RCT to assess the clinical effect of intravenous vitamin C on organ dysfunction in critically ill patients after cardiac arrest. Trial registration ClinicalTrials.gov NCT03509662. Registered on April 26, 2018. https://clinicaltrials.gov/ct2/show/NCT03509662European Clinical Trials Database (EudraCT): 2017-004318-25. Registered on June 8, 2018. https://www.clinicaltrialsregister.eu/ctr-search/trial/2017-004318-25/NL

Survival of COVID-19 Patients With Respiratory Failure is Related to Temporal Changes in Gas Exchange and Mechanical Ventilation

Background: Respiratory failure due to coronavirus disease of 2019 (COVID-19) often presents with worsening gas exchange over a period of days. Once patients require mechanical ventilation (MV), the temporal change in gas exchange and its relation to clinical outcome is poorly described. We investigated whether gas exchange over the first 5 days of MV is associated with mortality and ventilator-free days at 28 days in COVID-19. Methods: In a cohort of 294 COVID-19 patients, we used data during the first 5 days of MV to calculate 4 daily respiratory scores: PaO2/FiO2 (P/F), oxygenation index (OI), ventilatory ratio (VR), and Murray lung injury score. The association between these scores at early (days 1-3) and late (days 4-5) time points with mortality was evaluated using logistic regression, adjusted for demographics. Correlation with ventilator-free days was assessed (Spearman rank-order coefficients). Results: Overall mortality was 47.6%. Nonsurvivors were older ( P < .0001), more male ( P = .029), with more preexisting cardiopulmonary disease compared to survivors. Mean PaO2 and PaCO2 were similar during this timeframe. However, by days 4 to 5 values for all airway pressures and FiO2 had diverged, trending lower in survivors and higher in nonsurvivors. The most substantial between-group difference was the temporal change in OI, improving 15% in survivors and worsening 11% in nonsurvivors ( P < .05). The adjusted mortality OR was significant for age (1.819, P = .001), OI at days 4 to 5 (2.26, P = .002), and OI percent change (1.90, P = .02). The number of ventilator-free days correlated significantly with late VR (−0.166, P < .05), early and late OI (−0.216, P < .01; −0.278, P < .01, respectively) and early and late P/F (0.158, P < .05; 0.283, P < .01, respectively). Conclusion: Nonsurvivors of COVID-19 needed increasing intensity of MV to sustain gas exchange over the first 5 days, unlike survivors. Temporal change OI, reflecting both PaO2 and the intensity of MV, is a potential marker of outcome in respiratory failure due to COVID-19.

Inhibition of microsomal prostaglandin E synthase-1 ameliorates acute lung injury in mice

Background To examine the effects of BI 1029539 (GS-248), a novel selective human microsomal prostaglandin E synthase-1 (mPGES-1) inhibitor, in experimental models of acute lung injury (ALI) and sepsis in transgenic mice constitutively expressing the mPGES1 (Ptges) humanized allele. Methods Series 1: Lipopolysaccharide (LPS)-induced ALI. Mice were randomized to receive vehicle, BI 1029539, or celecoxib. Series 2: Cecal ligation and puncture-induced sepsis. Mice were randomized to receive vehicle or BI 1029539. Results Series 1: BI 1029539 or celecoxib reduced LPS-induced lung injury, with reduction in neutrophil influx, protein content, TNF-ɑ, IL-1β and PGE2 levels in bronchoalveolar lavage (BAL), myeloperoxidase activity, expression of mPGES-1, cyclooxygenase (COX)-2 and intracellular adhesion molecule in lung tissue compared with vehicle-treated mice. Notably, prostacyclin (PGI2) BAL concentration was only lowered in celecoxib-treated mice. Series 2: BI 1029539 significantly reduced sepsis-induced BAL inflammatory cell recruitment, lung injury score and lung expression of mPGES-1 and inducible nitric oxide synthase. Treatment with BI 1029539 also significantly prolonged survival of mice with severe sepsis. Anti-inflammatory and anti-migratory effect of BI 1029539 was confirmed in peripheral blood leukocytes from healthy volunteers. Conclusions BI 1029539 ameliorates leukocyte infiltration and lung injury resulting from both endotoxin-induced and sepsis-induced lung injury.

Rapid postmortem ventilation improves donor lung viability by extending the tolerable warm ischemic time after cardiac death in mice

Uncontrolled donation after cardiac death (uDCD) contributes little to ameliorating donor lung shortage due to rapidly progressive warm ischemia after circulatory arrest. Here, we demonstrated non-hypoxia improves donor lung viability in a novel uDCD lung transplant model undergoing rapid ventilation after cardiac death and compared the evolution of ischemia-reperfusion injury in mice that underwent pulmonary artery ligation (PAL). The tolerable warm ischemia time at 37ºC was initially determined in mice using a modified PAL model. The donor lung following PAL was also transplanted into syngeneic mice and compared to those that underwent rapid ventilation or no ventilation at 37ºC prior to transplantation. Twenty-four hours following reperfusion, lung histology, PaO2/FIO2 ratio, and inflammatory mediators were measured. Four hours of PAL had little impact on PaO2/FIO2 ratio and acute lung injury score in contrast to significant injury induced by 5 hours of PAL. Four-hour PAL lungs showed an early myeloid-dominant inflammatory signature when compared to naïve lungs and substantially injured five-hour PAL lungs. In the context of transplantation, unventilated donor lungs showed severe injury after reperfusion, whereas ventilated donor lungs showed minimal changes in PaO2/FIO2 ratio, histologic score, and expression of inflammatory markers. Taken together, the tolerable warm ischemia time of murine lungs at 37oC can be extended by maintaining alveolar ventilation for up to 4 hours. Non-hypoxic lung warm ischemia-reperfusion injury shows an early transcriptional signature of myeloid cell recruitment and extracellular matrix proteolysis prior to blood-gas barrier dysfunction and significant tissue damage.