L-ergothioneine reduces nitration of lactoferrin and loss of antibacterial activity associated with nitrosative stress.

Lactoferrin (LF) is a multifunctional antimicrobial, anti-inflammatory, and antioxidant protein that occurs naturally in mammals, most notably in exocrine gland tissues and fluids, such as in the eye. Nitrosative stress can promote changes to tyrosine and other amino acid residues of the protein, which also reduces the activity of LF. l-ergothioneine (ET) is a potent anti-inflammatory antioxidant present in the eye and other tissues through nutrition or supplementation and that may play a role in the prevention or treatment of a variety of diseases. Here we investigated the ability of ET to reduce 3-nitrotyrosine (NTyr) formation using two separate substrates, with the goal of determining whether ET can protect the antibacterial function of LF and other proteins when exposed separately to peroxynitrite and tetranitromethane as nitrating reagents. Native human LF was used as a simple protein substrate, and lamb corneal lysate was chosen as one example of mammalian tissue with a more complex mixture of proteins and other biomolecules. Nitration was monitored by absorbance and fluorescence spectroscopy as well as sandwich (nitrated LF) and direct NTyr (corneal lysate) enzyme-linked immunosorbent assays (ELISAs). We found that pretreatment with ET reduced chemical modification of both native LF and corneal lysate samples and loss of antibacterial LF function due to exposure to the nitrating reagents. These initial results suggest that ET, raised to sufficiently elevated levels, could be tailored as a therapeutic agent to reduce effects of nitrosative stress on LF and in turn sustain the protein activity.

Nasopharyngeal Type-I Interferon for Immediately Available Prophylaxis Against Emerging Respiratory Viral Infections.

In addition to SARS-CoV-2 and its variants, emerging viruses that cause respiratory viral infections will continue to arise. Increasing evidence suggests a delayed, possibly suppressed, type 1 interferon (IFN-I) response occurs early during COVID-19 and other viral respiratory infections such as SARS and MERS. These observations prompt considering IFN-ß as a prophylactic or early intervention for respiratory viral infections. A rationale for developing and testing intranasal interferon beta (IFN-ß) as an immediately available intervention for new respiratory viral infections that will arise unexpectedly in the future is presented and supported by basic and clinical trial observations. IFN-ß prophylaxis could limit the spread and consequences of an emerging respiratory viral infection in at-risk individuals while specific vaccines are being developed.

Depressed prostaglandins and leukotrienes in veterans with Gulf War illness.

Background: There is need to understand biological markers and mechanisms in Gulf War illness (GWI). Goal: To examine whether and how eicosanoids - prostaglandins and leukotrienes - are altered in veterans with GWI. Methods: Seventy participants including 37 GWI and 33 healthy controls, shared exposure information, and had plasma eicosanoids assessed - prostaglandin F2 alpha (pgf2α), prostaglandin D2 (pgd2), leukotriene B4 (lb4) among others. Values were compared for GWI versus controls. Eicosanoid intercorrelations were compared in cases vs. controls. For the most significantly altered eicosanoid in GWI, exposure and symptom relations were assessed. Results: Prostaglandins and leukotrienes were depressed in GWI, strongest for pgf2α, then lb4. Eicosanoid intercorrelations differed in GWI vs. controls. Fuel-solvent, pesticide, radioactive chemicals and metal exposures related negatively to pgf2α; as, in GWI, did chemical attack and vaccines. Multivariate predictors included fuels-solvents and radioactive chemicals (negative); tetanus vaccine and herbicides (positive). Fuels-solvents and radioactive chemicals predicted lower pgf2α in cases, controls, and all participants controlled for case status. Lower pgf2α related to GWI "Kansas criteria" domains of pain, respiratory, and (borderline significantly) skin symptoms. Conclusion: Multiple eicosanoids are depressed in GWI, particularly pgf2α and lb4. Prior fuel-solvent exposures, radioactive chemicals, and (in GWI cases) vaccines were linked to lower pgf2α.

Injury Characteristics and von Willebrand Factor for the Prediction of Acute Respiratory Distress Syndrome in Patients With Burn Injury: Development and Internal Validation.

OBJECTIVE: To derive and validate a prediction model for the development of ARDS in burn-injured patients. SUMMARY BACKGROUND DATA: Burn injury carries the highest incidence of acute respiratory distress syndrome (ARDS) among all predisposing conditions, but few studies exist on risk factors in these patients. Studies employing biomarkers and clinical risk factors for predicting ARDS mortality have recently been examined but none exist for onset of ARDS nor in patients with burn injury. METHODS: This was a prospective multicenter study of 113 patients with isolated burn injury or inhalation injury. Clinical variables and plasma biomarkers representative of endothelial injury, epithelial injury, or inflammation were collected within 24 hours of admission. The most parsimonious model was chosen by considering discrimination, calibration, and model fit. RESULTS: Among the biomarkers measured in patients with burn injuries, a one-standard deviation increase in log-transformed levels of the A2 domain of von Willebrand factor in the first 24 hours was most strongly associated with the development of ARDS (OR 7.72; 95% CI: 1.64-36.28, P = 0.03). Of candidate models, a 3-variable model with %TBSA, inhalation injury, and von Willebrand factor-A2 had comparable discrimination to more complex models (area under the curve: 0.90; 95% CI 0.85-0.96). The 3-variable model had good model fit by Hosmer-Lemeshow test (P = 0.74) and maintained similar discrimination after accounting for performance optimism (Bootstrapped area under the curve: 0.90; 95% CI: 0.84-0.95). CONCLUSIONS: The 3-variable model with %TBSA, inhalation injury, and von Willebrand factor could be used to better identify at-risk patients for both the study and prevention of ARDS in patients with burn injury.

Development of a sandwich ELISA with potential for selective quantification of human lactoferrin protein nitrated through disease or environmental exposure.

Lactoferrin (LF) is an important multifunctional protein that comprises a large fraction of the protein mass in certain human fluids and tissues, and its concentration is often used to assess health and disease. LF can be nitrated by multiple routes, leading to changes in protein structure, and nitrated proteins can negatively impact physiological health via nitrosative stress. Despite an awareness of the detrimental effects of nitrated proteins and the importance of LF within the body, cost-effective methods for detecting and quantifying nitrated lactoferrin (NLF) are lacking. We developed a procedure to selectively quantify NLF using sandwich enzyme-linked immunosorbent assay (ELISA), utilizing a polyclonal anti-LF capture antibody paired with a monoclonal anti-nitrotyrosine detector antibody. The assay was applied to quantify NLF in samples of pure LF nitrated via two separate reactions at molar ratios of excess nitrating agent to the total number of tyrosine residues between 10/1 and 100/1. Tetranitromethane (TNM) was used as a laboratory surrogate for an environmental pathway selective for production of 3-nitrotyrosine, and sodium peroxynitrite (ONOO-) was used as a surrogate for an endogenous nitration pathway. UV-vis spectroscopy (increased absorbance at 350 nm) and fluorescence spectroscopy (emission decreased by > 96%) for each reaction indicate the production of NLF. A lower limit of NLF detection using the ELISA method introduced here was calculated to be 0.065 µg mL-1, which will enable the detection of human-physiologically relevant concentrations of NLF. Our approach provides a relatively inexpensive and practical way to assess NLF in a variety of systems. Graphical abstract We developed a procedure to selectively quantify nitrated lactoferrin (NLF) protein using a sandwich enzyme-linked immunosorbent assay (ELISA) and verified results against several spectroscopic techniques. Our approach provides an inexpensive and practical way to assess NLF in a variety of systems.

Inhalation of two putative Gulf War toxins by mice.

We employed our inhalation methodology to examine whether biomarkers of inflammation and oxidative stress would be produced in mice following inhalation of aerosols containing carbonaceous particles or the vapor of pesticides prevalent during the first Gulf War. Exposure to two putative Gulf War Illness toxins, fine airborne particles and the pesticide malathion, increased biomarkers of inflammation and oxidative stress in Friend virus B (FVB) female mice. Mice inhaling particles 24 h before had increased lung lavage and plasma Leukotriene B4 (LTB4) (a biomarker of inflammation) and PGF2α (a biomarker of oxidative stress) levels, lung lavage protein and lung lavage lactic dehydrogenase (LDH) levels. These changes were a function of particle density and exposure time. Compared to particle inhalation, mice inhaling malathion 24 h before had small increase in plasma LTB4 and PGF2α levels but no increase in lung lavage LTB4, lung lavage protein, lung lavage LDH, and lung lavage alveolar macrophage (AM) levels compared to unexposed control mice. AM from particle-exposed mice contained phagocytosed particles, while AM from malathion-exposed mice showed no abnormalities. Our results indicate that inhaling particles or malathion can alter inflammatory and oxidative biomarkers in mice and raise the possibility that these toxins may have altered inflammation and oxidative stress biomarkers in Gulf War-exposed individuals.

Brief Glutamine Pretreatment Increases Alveolar Macrophage CD163/Heme Oxygenase-1/p38-MAPK Dephosphorylation Pathway and Decreases Capillary Damage but Not Neutrophil Recruitment in IL-1/LPS-Insufflated Rats.

BACKGROUND: Glutamine (GLN) attenuates acute lung injury (ALI) but its effect on alveolar macrophages is unknown. We hypothesized that GLN pretreatment would induce the anti-inflammatory CD163/heme oxygenase (HO)-1/p38-MAPK dephosphorylation pathway in alveolar macrophages and reduce ALI in rats insufflated with interleukin-1 (IL-1) and lipopolysaccharide (LPS). METHODS: Male Sprague-Dawley rats were randomized to the following groups: GLN-IL-1/LPS-, GLN+IL-1/LPS-, GLN-IL-1/LPS+, and GLN+IL-1/LPS+. GLN pretreatment was given via gavage (1 g/kg L-alanyl-L-glutamine) daily for 2 days. ALI was subsequently induced by insufflating 50 ng IL-1 followed by 5mg/kg E.coli LPS. After 24h, bronchoalveolar lavage (BAL) protein, lactate dehydrogenase (LDH) and neutrophil concentrations were analyzed. BAL alveolar macrophage CD163+ expression, HO-1 and p38-MAPK concentrations were measured, as well as alveolar macrophage tumor necrosis factor (TNF)-α and interleukin (IL)-10 concentrations. Histology and immunofluorescence studies were also performed. RESULTS: Following IL-1/LPS insufflation, GLN pretreated rats had significantly decreased BAL protein and LDH concentrations, but not BAL neutrophil counts, compared to non-GLN pretreated rats. The number of alveolar macrophages and the number of CD163+ macrophages were significantly increased in GLN pretreated IL-1/LPS-insufflated rats compared to non-GLN pretreated, IL-1/LPS-insufflated rats. GLN pretreatment before IL-1/LPS also significantly increased HO-1 concentrations and dephosphorylated p38-MAPK levels but not cytokine levels in alveolar macrophages. Immunofluorescence localized CD163 and HO-1 in alveolar macrophages. CONCLUSION: Short-term GLN pretreatment activates the anti-inflammatory CD163/HO-1/p38-MAPK dephosphorylation pathway of alveolar macrophages and decreases capillary damage but not neutrophil recruitment in IL-1/LPS-insufflated rats.

Perioperative lung protective ventilation in obese patients.

The perioperative use and relevance of protective ventilation in surgical patients is being increasingly recognized. Obesity poses particular challenges to adequate mechanical ventilation in addition to surgical constraints, primarily by restricted lung mechanics due to excessive adiposity, frequent respiratory comorbidities (i.e. sleep apnea, asthma), and concerns of postoperative respiratory depression and other pulmonary complications. The number of surgical patients with obesity is increasing, and facing these challenges is common in the operating rooms and critical care units worldwide. In this review we summarize the existing literature which supports the following recommendations for the perioperative ventilation in obese patients: (1) the use of protective ventilation with low tidal volumes (approximately 8 mL/kg, calculated based on predicted -not actual- body weight) to avoid volutrauma; (2) a focus on lung recruitment by utilizing PEEP (8-15 cmH2O) in addition to recruitment maneuvers during the intraoperative period, as well as incentivized deep breathing and noninvasive ventilation early in the postoperative period, to avoid atelectasis, hypoxemia and atelectrauma; and (3) a judicious oxygen use (ideally less than 0.8) to avoid hypoxemia but also possible reabsorption atelectasis. Obesity poses an additional challenge for achieving adequate protective ventilation during one-lung ventilation, but different lung isolation techniques have been adequately performed in obese patients by experienced providers. Postoperative efforts should be directed to avoid hypoventilation, atelectasis and hypoxemia. Further studies are needed to better define optimum protective ventilation strategies and analyze their impact on the perioperative outcomes of surgical patients with obesity.

Early effect of tidal volume on lung injury biomarkers in surgical patients with healthy lungs.

BACKGROUND: The early biological impact of short-term mechanical ventilation on healthy lungs is unknown. The authors aimed to characterize the immediate tidal volume (VT)-related changes on lung injury biomarkers in patients with healthy lungs and low risk of pulmonary complications. METHODS: Twenty-eight healthy patients for knee replacement surgery were prospectively randomized to volume-controlled ventilation with VT 6 (VT6) or 10 (VT10) ml/kg predicted body weight. General anesthesia and other ventilatory parameters (positive end-expiratory pressure, 5 cm H2O, FIO2, 0.5, respiratory rate titrated for normocapnia) were managed similarly in the two groups. Exhaled breath condensate and blood samples were collected for nitrite, nitrate, tumor necrosis factor-α, interleukins-1ß, -6, -8, -10, -11, neutrophil elastase, and Clara Cell protein 16 measurements, at the onset of ventilation and 60 min later. RESULTS: No significant differences in biomarkers were detected between the VT groups at any time. The coefficient of variation of exhaled breath condensate nitrite and nitrate decreased in the VT6 but increased in the VT10 group after 60-min ventilation. Sixty-minute ventilation significantly increased plasma neutrophil elastase levels in the VT6 (35.2 ± 30.4 vs. 56.4 ± 51.7 ng/ml, P = 0.008) and Clara Cell protein 16 levels in the VT10 group (16.4 ± 8.8 vs. 18.7 ± 9.5 ng/ml, P = 0.015). Exhaled breath condensate nitrite correlated with plateau pressure (r = 0.27, P = 0.042) and plasma neutrophil elastase (r = 0.44, P = 0.001). Plasma Clara Cell protein 16 correlated with compliance (r = 0.34, P = 0.014). CONCLUSIONS: No tidal volume-related changes were observed in the selected lung injury biomarkers of patients with healthy lungs after 60-min ventilation. Plasma neutrophil elastase and plasma Clara Cell protein 16 might indicate atelectrauma and lung distention, respectively.

Chronic inflammatory diseases and the Acute Respiratory Distress Syndrome (ARDS).

The Acute Respiratory Distress Syndrome (ARDS) is a highly fatal pro-inflammatory oxidative respiratory disease. Relatively recently, the modulating effects of chronic inflammatory processes on ARDS susceptibility have been recognized in a number of clinical studies. Herein, we briefly review some of the chronic conditions that have been reported to increase (cigarette smoking and alcohol abuse) or decrease (diabetes and obesity) susceptibility to ARDS. We also propose some potential pathways that may hold clues regarding the pathogenesis and/or therapy for ARDS.

Early increase in alveolar macrophage prostaglandin 15d-PGJ2 precedes neutrophil recruitment into lungs of cytokine-insufflated rats.

Early detection and prevention is an important goal in acute respiratory distress syndrome research. We determined the concentration of the anti-inflammatory 15-deoxy-Δ(12,14)-prostaglandin-J2 (15d-PGJ2) and other components of the cyclopentenone prostaglandin cascade in relation to lung inflammation in cytokine (IL-1/LPS)-insufflated rats. We found that 15d-PGJ2 levels increase in the bronchoalveolar lavage (BAL) fluid of rats insufflated with cytokines 2 h before. BAL 15d-PGJ2 increases preceded neutrophil recruitment, lung injury, and oxidative stress in the lungs of cytokine-insufflated rats. 15d-PGJ2 was localized in alveolar macrophages that decreased following cytokine insufflation. 15d-PGJ2 may constitute an early biomarker of lung inflammation and may reflect an endogenous attempt to regulate ongoing inflammation in macrophages and elsewhere after cytokine insufflation.

Functional benefits of ergothioneine and fruit- and vegetable-derived nutraceuticals: overview of the supplemental issue contents.

For good reason, there is increasing interest in assessing the clinical efficacy of dietary supplements, naturally occurring compounds, and nutraceuticals intended for improving health and reducing disease. This is also a pressing interest in mitigating the effects of age-dependent chronic diseases. This opportunity argues for the need to develop a clear understanding of the basic molecular mechanisms responsible for the actions of dietary biofactors that can contribute to the slowing or preventing of diseases and the possibility of enhancing these improvements by coupling them with healthy lifestyle changes.

Effect of ergothioneine on acute lung injury and inflammation in cytokine insufflated rats.

OBJECTIVE: The Acute Respiratory Distress Syndrome (ARDS), the most severe form of Acute Lung Injury (ALI), is a highly-fatal, diffuse non-cardiogenic edematous lung disorder. The pathogenesis of ARDS is unknown but lung inflammation and lung oxidative stress are likely contributing factors. Since no specific pharmacologic intervention exists for ARDS, our objective was to determine the effect of treatment with ergothioneine-a safe agent with multiple anti-inflammatory and antioxidant properties on the development of lung injury and inflammation in rats insufflated with cytokines found in lung lavages of ARDS patients. METHOD: Sprague-Dawley rats (3-10/group) were given 15 mg/kg or 150 mg/kg l-ergothioneine intravenously 1h before or 18 h after cytokine (IL-1 and IFNγ) insufflation. Lung injury (lavage LDH levels) and lung inflammation (lavage neutrophil numbers) were measured 24h after cytokine insufflation. RESULTS: Ergothioneine pre- and post-treatment generally decreased lung injury and lung inflammation in cytokine insufflated rats. CONCLUSION: Ergothioneine should be considered for additional testing as a potential therapy for treating and preventing ARDS.

Preventing the acute respiratory distress syndrome.

Laudable supportive advances have been made to improve the care of patients with the Acute Respiratory Distress Syndrome (ARDS) but no pharmacologic interventions are known to reduce the high mortality of this disorder once it is established. This commentary discusses some of the challenges that arise in preventing ARDS in at-risk individuals and the likely dependence of this approach on biomarker panels that can be done in real time.

Hyperpolarized 83Kr MRI of lungs.

Hyperpolarized (hp) (83)Kr (spin I=9/2) is a promising gas-phase contrast agent that displays sensitivity to the surface chemistry, surface-to-volume ratio, and surface temperature of the surrounding environment. This proof-of-principle study demonstrates the feasibility of ex vivo hp (83)Kr magnetic resonance imaging (MRI) of lungs using natural abundance krypton gas (11.5% (83)Kr) and excised, but otherwise intact, rat lungs located within a custom designed ventilation chamber. Experiments comparing the (83)Kr MR signal intensity from lungs to that arising from a balloon with no internal structure inflated to the same volume with krypton gas mixture suggest that most of the observed signal originated from the alveoli and not merely the conducting airways. The (83)Kr longitudinal relaxation times in the rat lungs ranged from 0.7 to 3.7s but were reproducible for a given lung. Although the source of these variations was not explored in this work, hp (83)Kr T(1) differences may ultimately lead to a novel form of MRI contrast in lungs. The currently obtained 1200-fold signal enhancement for hp (83)Kr at 9.4T field strength is found to be 180 times below the theoretical upper limit.

Effects of fine carbonaceous particles containing high and low unpaired electron spin densities on lungs of female mice.

The negative impacts on human health that accompany inhalation of atmospheric particles are documented in numerous epidemiologic studies, but the effect of specific chemical properties of the particles is generally unknown. We developed and employed technology for generating inhalable aerosols of carbonaceous air pollution particles that have specific physical and chemical properties. We find that inhaling particles with greater unpaired electron spin (free radical) densities stimulates greater lung inflammatory and oxidative stress responses. Cultured alveolar macrophages take up more particles of greater free radical content, develop mitochondrial abnormalities, and release more leukotriene B(4) (LTB(4)) than alveolar macrophages exposed to lesser free-radical-containing particles in vitro. Mice exposed to high free radical particles in vivo also develop mitochondrial abnormalities in alveolar macrophages and increased oxidative stress, which is reflected by increases in lung nitrotyrosine staining and lung lavage nitrogen oxide levels compared with those of lesser free radical density. These results provide insight for the unexplained geographic differences and have implications for fossil fuel combustion conditions and the impact of fine particles on health and disease.

Detection of tobacco smoke deposition by hyperpolarized krypton-83 MRI.

Despite the importance of the tobacco smoke particulate matter in the lungs to the etiology of pulmonary disease in cigarette smokers, little is currently known about the spatial distribution of particle deposition or the persistence of the resulting deposits in humans, and no satisfactory technique currently exists to directly observe tobacco smoke condensate in airways. In this proof-of-principle work, hyperpolarized (hp) 83Kr MRI and NMR spectroscopy are introduced as probes for tobacco smoke deposition in porous media. A reduction in the hp-83Kr longitudinal (T1) relaxation of up to 95% under near-ambient humidity, pressure and temperature conditions was observed when the krypton gas was brought into contact with surfaces that had been exposed to cigarette smoke. This smoke-induced acceleration of the 83Kr self-relaxation was observed for model glass surfaces that, in some experiments, were coated with bovine lung surfactant extract. However, a similar effect was not observed with hp-(129)Xe indicating that the 83Kr sensitivity to smoke deposition was not caused by paramagnetic species but rather by quadrupolar relaxation due to high adsorption affinity for the smoke deposits. The 83Kr T1 differences between smoke-treated and untreated surfaces were sufficient to produce a strong contrast in variable flip angle FLASH hp-83Kr MRI, suggesting that hp-83Kr may be a promising contrast agent for in vivo pulmonary MRI.

Hyperpolarized 83Kr and 129Xe NMR relaxation measurements of hydrated surfaces: implications for materials science and pulmonary diagnostics.

In this proof of principle work, a technique is introduced to study hydrated surfaces using hyperpolarized (hp) 83Kr NMR spectroscopy. The longitudinal (T1) relaxation of hp-83Kr is shown to be extremely sensitive to the presence of adsorbed water on hydrophilic borosilicate and hydrophobic siliconized glass surfaces. The krypton surface relaxation is found to be largely independent of the total gas pressure applied to the studied materials, and the presented technique is therefore fairly robust. However, the relaxational properties of hp-83Kr can be "tuned" by adjusting the composition of the optical pumping gas mixture. This effect may be important for practical applications such as hp-83Kr MR imaging and can be achieved without sacrificing signal intensity. Complementary information to that of hp-83Kr surface relaxation data can be obtained from hp-129Xe relaxation measurements that are sensitive to the presence of paramagnetic surface sites. In contrast to the signal decay of hp-129Xe, the longitudinal relaxation of 83Kr is largely unaffected by paramagnetic impurities, and in some materials, 83Kr and 129Xe show comparable T1 times that are caused by two completely different relaxation mechanisms. Finally, the relaxation times of 83Kr in contact with bovine lung surfactant coated glass pores that are similar in size to mammalian alveoli are presented. The results suggest that in vivo MR studies may be feasible and could provide valuable information about changes in pulmonary surface chemistry.