Magnetic Resonance Imaging Quantification of Pulmonary Perfusion using Calibrated Arterial Spin Labeling

This demonstrates a MR imaging method to measure the spatial distribution of pulmonary blood flow in healthy subjects during normoxia (inspired O₂, fraction (F_IO₂) = 0.21) hypoxia (F_IO₂ = 0.125), and hyperoxia (F_IO₂ = 1.00). In addition, the physiological responses of the subject are monitored in the MR scan environment. MR images were obtained on a 1.5 T GE MRI scanner during a breath hold from a sagittal slice in the right lung at functional residual capacity. An arterial spin labeling sequence (ASL-FAIRER) was used to measure the spatial distribution of pulmonary blood flow ^1,2 and a multi-echo fast gradient echo (mGRE) sequence ³ was used to quantify the regional proton (i.e. H₂O) density, allowing the quantification of density-normalized perfusion for each voxel (milliliters blood per minute per gram lung tissue). With a pneumatic switching valve and facemask equipped with a 2-way non-rebreathing valve, different oxygen concentrations were introduced to the subject in the MR scanner through the inspired gas tubing. A metabolic cart collected expiratory gas via expiratory tubing. Mixed expiratory O₂ and CO₂ concentrations, oxygen consumption, carbon dioxide production, respiratory exchange ratio, respiratory frequency and tidal volume were measured. Heart rate and oxygen saturation were monitored using pulse-oximetry. Data obtained from a normal subject showed that, as expected, heart rate was higher in hypoxia (60 bpm) than during normoxia (51) or hyperoxia (50) and the arterial oxygen saturation (SpO₂) was reduced during hypoxia to 86%. Mean ventilation was 8.31 L/min BTPS during hypoxia, 7.04 L/min during normoxia, and 6.64 L/min during hyperoxia. Tidal volume was 0.76 L during hypoxia, 0.69 L during normoxia, and 0.67 L during hyperoxia. Representative quantified ASL data showed that the mean density normalized perfusion was 8.86 ml/min/g during hypoxia, 8.26 ml/min/g during normoxia and 8.46 ml/min/g during hyperoxia, respectively. In this subject, the relative dispersion⁴, an index of global heterogeneity, was increased in hypoxia (1.07 during hypoxia, 0.85 during normoxia, and 0.87 during hyperoxia) while the fractal dimension (Ds), another index of heterogeneity reflecting vascular branching structure, was unchanged (1.24 during hypoxia, 1.26 during normoxia, and 1.26 during hyperoxia). Overview. This protocol will demonstrate the acquisition of data to measure the distribution of pulmonary perfusion noninvasively under conditions of normoxia, hypoxia, and hyperoxia using a magnetic resonance imaging technique known as arterial spin labeling (ASL). Rationale: Measurement of pulmonary blood flow and lung proton density using MR technique offers high spatial resolution images which can be quantified and the ability to perform repeated measurements under several different physiological conditions. In human studies, PET, SPECT, and CT are commonly used as the alternative techniques. However, these techniques involve exposure to ionizing radiation, and thus are not suitable for repeated measurements in human subjects.Download video file.^{(74M, mov)}     

Aerosolized bovine lactoferrin reduces lung injury and fibrosis in mice exposed to hyperoxia

This study investigated the ability of aerosolized bovine lactoferrin (bLF) to protect the lungs from injury induced by chronic hyperoxia. Female CD-1 mice were exposed to hyperoxia (FiO₂ = 80 %) for 7 days to induce lung injury and fibrosis. The therapeutic effects of bLF, administered via an aerosol delivery system, on the chronic lung injury induced by this period of hyperoxia were measured by bronchoalveolar lavage, lung histology, cell apoptosis, and inflammatory cytokines in the lung tissues. After exposure to hyperoxia for 7 days, the survival of the mice was significantly decreased to 20 %. The protective effects of bLF against hyperoxia were further confirmed by significant reductions in lung edema, total cell numbers in bronchoalveolar lavage fluid, inflammatory cytokines (IL-1β and IL-6), pulmonary fibrosis, and apoptotic DNA fragmentation. The aerosolized bLF protected the mice from oxygen toxicity and increased the survival fraction to 66.7 % in the hyperoxic model. The results support the use of an aerosol therapy with bLF in intensive care units to reduce oxidative injury in patients with severe hypoxemic respiratory failure or chronic obstructive pulmonary disease.     

Disruption of cytochrome P4501A2 in mice leads to increased susceptibility to hyperoxic lung injury

Hyperoxia contributes to acute lung injury in diseases such as acute respiratory distress syndrome. Cytochrome P450 (CYP) 1A enzymes have been implicated in hyperoxic lung injury, but the mechanistic role of CYP1A2 in pulmonary injury is not known. We hypothesized that mice lacking the gene Cyp1a2 (which is predominantly expressed in the liver) will be more sensitive to lung injury and inflammation mediated by hyperoxia and that CYP1A2 will play a protective role by attenuating lipid peroxidation and oxidative stress in the lung. Eight- to ten-week-old WT (C57BL/6) or Cyp1a2^−/− mice were exposed to hyperoxia (>95% O₂) or maintained in room air for 24–72 h. Lung injury was assessed by determining the ratio of lung weight/body weight (LW/BW) and by histology. Extent of inflammation was determined by measuring the number of neutrophils in the lung as well as cytokine expression. The Cyp1a2^−/− mice under hyperoxic conditions showed increased LW/BW ratios, lung injury, neutrophil infiltration, and IL-6 and TNF-α levels and augmented lipid peroxidation, as evidenced by increased formation of malondialdehyde– and 4-hydroxynonenal–protein adducts and pulmonary isofurans compared to WT mice. In vitro experiments showed that the F₂-isoprostane PGF₂-α is metabolized by CYP1A2 to a dinor metabolite, providing evidence for a catalytic role for CYP1A2 in the metabolism of F₂-isoprostanes. In summary, our results support the hypothesis that hepatic CYP1A2 plays a critical role in the attenuation of hyperoxic lung injury by decreasing lipid peroxidation and oxidative stress in vivo.     

Involvements of p38 MAPK and oxidative stress in the ozone-induced enhancement of AHR and pulmonary inflammation in an allergic asthma model

Background

Exposure to ambient ozone (O₃) increases the susceptivity to allergens and triggers exacerbations in patients with asthma. However, the detailed mechanisms of action for O₃ to trigger asthma exacerbations are still unclear.

Methods

An ovalbumin (OVA)-established asthmatic mouse model was selected to expose to filtered air (OVA-model) or 1.0 ppm O₃ (OVA-O₃ model) during the process of OVA challenge. Next, the possible involvements of p38 MAPK and oxidative stress in the ozone actions on the asthma exacerbations were investigated on the mice of OVA-O₃ model by treating them with SB239063 (a p38 MAPK inhibitor), and/or the α-tocopherol (antioxidant). Biological measurements were conducted including airway hyperresponsiveness (AHR), airway resistance (Raw), lung compliance (CL), inflammation in the airway lumen and lung parenchyma, the phosphorylation of p38 MAPK and heat shock protein (HSP) 27 in the tracheal tissues, and the malondialdehyde (MDA) content and the glutathione peroxidase (GSH-Px) activity in lung tissues.

Results

In OVA-allergic mice, O3 exposure deteriorated airway hyperresponsiveness (AHR), airway resistance (Raw), lung compliance (CL) and pulmonary inflammation, accompanied by the increased oxidative stress in lung tissues and promoted p38 MAPK and HSP27 phosphorylation in tracheal tissues. Administration of SB239063 (a p38 MAPK inhibitor) on OVA-O3 model exclusively mitigated the Raw, the CL, and the BAL IL-13 content, while α-tocopherol (antioxidant) differentially reduced the BAL number of eosinophils and macrophages, the content of BAL hyaluronan, the peribronchial inflammation, as well as the mRNA expression of TNF-α and IL-5 in the lung tissues of OVA-O3 model. Administration of these two chemical inhibitors similarly inhibited the AHR, the BAL IFN-γ and IL-6 production, the perivascular lung inflammation and the lung IL-17 mRNA expression of OVA-O3 model. Interestingly, the combined treatment of both compounds together synergistically inhibited neutrophil counts in the BALF and CXCL-1 gene expression in the lung.

Conclusions

O₃ exposure during the OVA challenge process promoted exacerbation in asthma. Both p38 MAPK and oxidative stress were found to play a critical role in this process and simultaneous inhibition of these two pathways significantly reduced the O₃-elicited detrimental effects on the asthma exacerbation.

     

Cytokines increase rat lung antioxidant enzymes during exposure to hyperoxia

Pretreatment with the combination of tumor necrosis factor/cachectin (TNF/C) and interleukin 1 (IL-1) increased glucose-6-phosphate dehydrogenase (G6PDH), glutathione reductase (GR), glutathione peroxidase (GPX), catalase (CAT), and superoxide dismutase (SOD) activities in lungs of rats continuously exposed to hyperoxia for 72 h, a time when all untreated rats had already died. Pretreatment with TNF/C and IL-1 also increased, albeit slightly, lung G6PDH and GR activities of rats exposed to hyperoxia for 4 or 16 h. By comparison, no differences occurred in lung antioxidant enzyme activities of TNF/C and IL-1- or saline-pretreated rats exposed to hyperoxia for 36 or 52 h; the latter is a time just before untreated rats began to succumb during exposure to hyperoxia. The results raise the possibility that TNF/C and IL-1 treatment can increase lung antioxidant enzyme activities and that increased lung antioxidant enzymes may contribute to the increased survival of TNF/C and IL-1-pretreated rats in hyperoxia for greater than 72 h.     

Modification of alveolar hyperoxia induced pulmonary vasodilatation by indomethacin

Decrease in pulmonary vascular resistance was observed in neonatal minature pigs breathing 100% O₂ or 95% O₂:5% CO₂. The pulmonary vasodilator response to hyperoxia ventilation was reduced by indomethacin in the intact animal and in the isolated perfused lung preparation. In the isolated perfused lung preparation, it was also shown that lung alveolar pO₂ rather than pulmonary arterial pO₂ was responsible for the pulmonary vasodilation. The study suggests that alveolar hyperoxia induced decrease in pulmonary vascular resistance may be mediated in part by release of prostaglandins. The relevance of this study with oxygen therapy in newborn infants is also discussed.     

Antioxidant and Pulmonary Protective Potential of Fraxinus xanthoxyloides Bark Extract against CCl4-Induced Toxicity in Rats

Fraxinus xanthoxyloides is a perennial shrub belonging to family Oleaceae, traditionally used for malaria, jaundice, pneumonia, inflammation, and rheumatism. Our study is aimed to assess the total phenolics (TPC), flavonoids (TFC), terpenoids contents (TTC) and antioxidant profiling of F. xanthoxyloides methanol bark extract (FXBM) and its fractions, hexane, chloroform, ethyl acetate and aqueous, along with high-performance liquid chromatography with diode-array detection (HPLC-DAD). Further, the antioxidant and pulmonary protective potential was explored against carbon tetrachloride (CCl₄)-induced CCl4-induced pulmonary tissue damage in rats. The highest TPC, TFC and TTC were found in FXBM (133.29±4.19 mg/g), ethyl acetate fraction (279.55±10.35 mg/g), and chloroform fraction (0.79±0.06 mg/g), respectively. The most potent antioxidant capacity was depicted by FXBM (29.21±2.40 μg/mg) and ethyl acetate fraction (91.16±5.51 μg/mg). The HPLC-DAD analysis revealed the predominance of gallic, chlorogenic, vanillic and ferulic acid in FXBM. The administration of CCl₄ induced oxidative stress, suppressed antioxidant enzymes′ activities including catalase, peroxidase, superoxide dismutase, glutathione peroxidase, glutathione-S-transferase, and glutathione reductase. Further, it increased thiobarbituric acid reactive substances (TBARS) and H₂O₂ levels, induced DNA injuries and reduced the total protein and glutathione content in lung tissues. The treatment of rats with FXBM restored these biochemical parameters to the normal level. Moreover, the histopathological studies of lung tissues demonstrated that FXBM protected rats′ lung tissues from oxidative damage restoring normal lung functions. Thus, F. xanthoxyloides bark extract is recommended as adjuvant therapy as protective agent for patients with lung disorders.     

Blunted pulmonary pressor responses to hypoxia in blood perfused, ventilated lungs isolated from oxygen toxic rats: Possible role of prostaglandins

To determine the effects of high oxygen (O₂) tension on pulmonary vascular reactivity, we exposed rats either to 100% O₂ for 48 hrs or 40% O₂ for 3 to 5 weeks. Lungs from all rats were isolated, blood perfused and ventilated, and pressor responses to airway hypoxia and to infused angiotensin II were measured. We found that chronic subtoxic hyperoxia did not augment subsequent hypoxic vasoconstriction, and that 48 hrs of 100% O₂ markedly blunted hypoxic vasoconstriction. Meclofenamate restored hypoxic vasoconstriction to control levels in the lungs with blunted responses. Evidence for O₂ toxicity in the lungs exposed to 100% O₂ included interstitial swelling with alveolar exudates seen by light microscopy, and lung edema by water content calculations. We conclude that 1) chronic subtoxic hyperoxia does not influence subsequent hypoxic vasoconstriction, and 2) a dilator prostaglandin produced in the lung is a potent inhibitor of hypoxic vasoconstriction in O₂ toxic lungs.     

Synergistic protection against hyperoxia-induced lung injury by neutrophils blockade and EC-SOD overexpression

Background

Oxygen may damage the lung directly via generation of reactive oxygen species (ROS) or indirectly via the recruitment of inflammatory cells, especially neutrophils. Overexpression of extracellular superoxide dismutase (EC-SOD) has been shown to protect the lung against hyperoxia in the newborn mouse model. The CXC-chemokine receptor antagonist (Antileukinate) successfully inhibits neutrophil influx into the lung following a variety of pulmonary insults. In this study, we tested the hypothesis that the combined strategy of overexpression of EC-SOD and inhibiting neutrophil influx would reduce the inflammatory response and oxidative stress in the lung after acute hyperoxic exposure more efficiently than either single intervention.

Methods

Neonate transgenic (Tg) (with an extra copy of hEC-SOD) and wild type (WT) were exposed to acute hyperoxia (95% FiO₂ for 7 days) and compared to matched room air groups. Inflammatory markers (myeloperoxidase, albumin, number of inflammatory cells), oxidative markers (8-isoprostane, ratio of reduced/oxidized glutathione), and histopathology were examined in groups exposed to room air or hyperoxia. During the exposure, some mice received a daily intraperitoneal injection of Antileukinate.

Results

Antileukinate-treated Tg mice had significantly decreased pulmonary inflammation and oxidative stress compared to Antileukinate-treated WT mice (p < 0.05) or Antileukinate-non-treated Tg mice (p < 0.05).

Conclusion

Combined strategy of EC-SOD and neutrophil influx blockade may have a therapeutic benefit in protecting the lung against acute hyperoxic injury.     

Interactions of nitric oxide and oxygen in cytotoxicity: Proliferation and antioxidant enzyme activities of endothelial cells in culture

Nitric oxide (NO) shows cytotoxicity, and its reaction products with reactive oxygen species, such as peroxynitrite, are potentially more toxic. To examine the role of O₂ in the NO toxicity, we have examined the proliferation of cultured human umbilical vein endothelial cells in the presence or absence of NO donor, ((Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)-amino]diazen-1-ium-1,2-diolate) (DETA-NONOate) (100–500 μM), under normoxia (air), hypoxia (< 0.04% O₂) or hyperoxia (88–94% O₂). It was found that the dose dependency on NONOate was little affected by the ambient O₂ concentration, showing no apparent synergism between the two treatments. We have also examined the effects of exogenous NO under normoxia and hyperoxia on the cellular activities of antioxidant enzymes involved in the H₂O₂ elimination, since many of them are known to be inhibited by NO or peroxynitrite in vitro. Under normoxia DETA-NONOate (500 μM) caused 25% decrease in catalase activity and 30% increases in glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase activities in 24 h. Under hyperoxia NO caused about 25% decreases in activities of catalase, glutathione reductase and glucose-6-phosphate dehydrogenase. The H₂O₂ removal rate by NO-treated cells was computed on the mathematical model for the enzyme system. It was concluded that the cellular antioxidant function is little affected by NO under normoxia but that it is partially impaired when the cells are exposed to NO under hyperoxia.     

Contributions of nitric oxide synthase isoforms to pulmonary oxygen toxicity, local vs. mediated effects

Reactive species of oxygen and nitrogen have been collectively implicated in pulmonary oxygen toxicity, but the contributions of specific molecules are unknown. Therefore, we assessed the roles of several reactive species, particularly nitric oxide, in pulmonary injury by exposing wild-type mice and seven groups of genetically altered mice to >98% O2 at 1, 3, or 4 atmospheres absolute. Genetically altered animals included knockouts lacking either neuronal nitric oxide synthase (nNOS(-/-)), endothelial nitric oxide synthase (eNOS(-/-)), inducible nitric oxide synthase (iNOS(-/-)), extracellular superoxide dismutase (SOD3(-/-)), or glutathione peroxidase 1 (GPx1(-/-)), as well as two transgenic variants (S1179A and S1179D) having altered eNOS activities. We confirmed our earlier finding that normobaric hyperoxia (NBO2) and hyperbaric hyperoxia (HBO2) result in at least two distinct but overlapping patterns of pulmonary injury. Our new findings are that the role of nitric oxide in the pulmonary pathophysiology of hyperoxia depends both on the specific NOS isozyme that is its source and on the level of hyperoxia. Thus, iNOS predominates in the etiology of lung injury in NBO2, and SOD3 provides an important defense. But in HBO2, nNOS is a major contributor to pulmonary injury, whereas eNOS is protective. In addition, we demonstrated that nitric oxide derived from nNOS is involved in a neurogenic mechanism of HBO2-induced lung injury that is linked to central nervous system oxygen toxicity through adrenergic/cholinergic pathways.     

Adenovirus-mediated transfer of the SOCS-1 gene to mouse lung confers protection against hyperoxic acute lung injury

Suppressor of cytokine signaling-1 (SOCS-1) is a member of the suppressor of cytokine signaling family of proteins and an inhibitor of interleukin-6 (IL-6) signaling. SOCS-1 has been shown to protect cells from cellular damage and apoptosis induced by tumor necrosis factor (TNF), lipopolysaccharide (LPS), and interferon gamma (IL-γ). However, it is not known whether increased SOCS-1 is protective during pulmonary oxidative stress. Therefore, we hypothesized that increased SOCS-1 in the lungs of mice would be protective in the setting of hyperoxic lung injury. We administered SOCS-1 adenovirus (Ad-SOCS-1) intratracheally into the lungs and exposed the mice to 100% O₂. Mice infected with GFP adenovirus (Ad-GFP) were used as controls. Mice treated with Ad-SOCS-1 had enhanced survival in 100% oxygen compared to Ad-GFP-administered mice. After 3 days of hyperoxia, Ad-GFP mice were ill and tachypnic and died after 4 days. In contrast, all Ad-SOCS-1-treated mice survived for at least 6 days in hyperoxia and 80% survived beyond 7 days. Ad-SOCS-1 transfection protected mouse lungs from injury as indicated by lower lung wet/dry weight, alveolar–capillary protein leakage, reduced infiltration of inflammatory cells, and lower content of thiobarbituric acid-reactive substances in lung homogenate. Our results also indicated that Ad-SOCS-1 significantly inhibits hyperoxia-induced ASK-1 (apoptosis signal-regulating kinase 1) expression. Taken together, these findings show that increased expression of adenovirus-mediated SOCS-1 in the lungs of mice significantly protects against hyperoxic lung injury.     

Enzyme defense against reactive oxygen derivatives. II. Erythrocytes and tumor cells

Summary The enzymatic destruction of oxidizing products produced during metabolic reduction of oxygen in the cell (such as singlet oxygen, H₂O₂ and OH radical) involves the concerted action of superoxide dismutase-which removes O ₂ ^- and yields H₂O₂-and H₂O₂ removing enzymes such as catalase and glutathione peroxidase. A difference in distribution or ratio of these enzymes in various tissues may result in a different reactivity of oxygen radicals.It was found that in red blood cells superoxide dismutase and catalase are extracted in the same fraction as hemoglobin, while glutathione peroxidase appears to be loosely bound to the cellular structure. This suggests that in red blood cells catalase acts in series with superoxide dismutase against bursts of oxygen radicals formed from oxyhemoglobin, while glutathione & peroxidase may protect the cell membrane against low concentrations of H₂O₂. On the other hand, catalase activity is absent in various types of ascites tumor cells, while glutathione peroxidase and superoxide dismutase are found in the cytoplasm. However, the peroxidase/dismutase ratio is lower than in liver cells, and this may provide an explanation for the higher susceptibility of tumor cells to treatments likely to involve oxygen radicals.     

The physiological ecology of Mytilus californianus Conrad

Summary The rates of oxygen consumption, filtration and ammonia excretion by Mytilus californianus have been related to body size and to ration. The rate of oxygen consumption (VO₂) by individuals while immersed, measured on the shore, resembled rates recorded for mussels starved in the laboratory. VO₂ by M. californianus was relatively independent of change in temperature, with a Q ₁₀ (13–22° C) of 1.20. In contrast, the frequency of heart beat was more completely temperature dependent [Q ₁₀ (13–22° C)=2.10]. Filtration rate showed intermediate dependence on temperature change [Q ₁₀ (13–22° C)=1.49] up to 22° C, but declined at 26° C. Both VO₂ and filtration rate declined during starvation. The utilisation efficiency for oxygen was low (approx. 4%) between 13 and 22° C, but increased to 10% at 26° C. Three components of the routine rate of oxygen consumption are recognised and estimated; viz. a basal rate (0.136 ml O₂ h^-1 for a mussel of 1 g dry flesh weight), a physiological cost of feeding (which represented about 6% of the calories in the ingested ration), and a mechanical cost of feeding which was three times higher than the physiological cost. The ratio oxygen consumed to ammonia-nitrogen excreted was low, and it declined during starvation. These data are compared with previously published measurements on Mytilus edulis, and the two species of mussel are shown to be similar in some of their physiological characteristics, though possibly differing in their capacities to compensate for change in temperature. For M. californianus, the scope for growth was highest at 17–22° C and declined at 26° C; it is suggested that exposure to temperatures in excess of 22° C, as for example during low tides in the summer, might result in a cumulative stress on these populations of mussels by imposing a metabolic deficit which must be recovered at each subsequent high tide. The high mechanical cost of feeding imposes a more general constraint on the scope for activity of the species.     

Hyperoxia disrupts pulmonary epithelial barrier in newborn rats via the deterioration of occludin and ZO-1

Background

Prolonged exposure to hyperoxia in neonates can cause hyperoxic acute lung injury (HALI), which is characterized by increased pulmonary permeability and diffuse infiltration of various inflammatory cells. Disruption of the epithelial barrier may lead to altered pulmonary permeability and maintenance of barrier properties requires intact epithelial tight junctions (TJs). However, in neonatal animals, relatively little is known about how the TJ proteins are expressed in the pulmonary epithelium, including whether expression of TJ proteins is regulated in response to hyperoxia exposure. This study determines whether changes in tight junctions play an important role in disruption of the pulmonary epithelial barrier during hyperoxic acute lung injury.

Methods

Newborn rats, randomly divided into two groups, were exposed to hyperoxia (95% oxygen) or normoxia for 1–7 days, and the severity of lung injury was assessed; location and expression of key tight junction protein occludin and ZO-1 were examined by immunofluorescence staining and immunobloting; messenger RNA in lung tissue was studied by RT-PCR; transmission electron microscopy study was performed for the detection of tight junction morphology.

Results

We found that different durations of hyperoxia exposure caused different degrees of lung injury in newborn rats. Treatment with hyperoxia for prolonged duration contributed to more serious lung injury, which was characterized by increased wet-to-dry ratio, extravascular lung water content, and bronchoalveolar lavage fluid (BALF):serum FD4 ratio. Transmission electron microscopy study demonstrated that hyperoxia destroyed the structure of tight junctions and prolonged hyperoxia exposure, enhancing the structure destruction. The results were compatible with pathohistologic findings. We found that hyperoxia markedly disrupted the membrane localization and downregulated the cytoplasm expression of the key tight junction proteins occludin and ZO-1 in the alveolar epithelium by immunofluorescence. The changes of messenger RNA and protein expression of occludin and ZO-1 in lung tissue detected by RT-PCR and immunoblotting were consistent with the degree of lung injury.

Conclusions

These data suggest that the disruption of the pulmonary epithelial barrier induced by hyperoxia is, at least in part, due to massive deterioration in the expression and localization of key TJ proteins.     

Novel Peptide for Attenuation of Hyperoxia-induced Disruption of Lung Endothelial Barrier and Pulmonary Edema via Modulating Peroxynitrite Formation

Pulmonary damages of oxygen toxicity include vascular leakage and pulmonary edema. We have previously reported that hyperoxia increases the formation of NO and peroxynitrite in lung endothelial cells via increased interaction of endothelial nitric oxide (eNOS) with β-actin. A peptide (P326TAT) with amino acid sequence corresponding to the actin binding region of eNOS residues 326–333 has been shown to reduce the hyperoxia-induced formation of NO and peroxynitrite in lung endothelial cells. In the present study, we found that exposure of pulmonary artery endothelial cells to hyperoxia (95% oxygen and 5% CO₂) for 48 h resulted in disruption of monolayer barrier integrity in two phases, and apoptosis occurred in the second phase. NOS inhibitor N^G-nitro-l-arginine methyl ester attenuated the endothelial barrier disruption in both phases. Peroxynitrite scavenger uric acid did not affect the first phase but ameliorated the second phase of endothelial barrier disruption and apoptosis. P326TAT inhibited hyperoxia-induced disruption of monolayer barrier integrity in two phases and apoptosis in the second phase. More importantly, injection of P326TAT attenuated vascular leakage, pulmonary edema, and endothelial apoptosis in the lungs of mice exposed to hyperoxia. P326TAT also significantly reduced the increase in eNOS-β-actin association and protein tyrosine nitration. Together, these results indicate that peptide P326TAT ameliorates barrier dysfunction of hyperoxic lung endothelial monolayer and attenuates eNOS-β-actin association, peroxynitrite formation, endothelial apoptosis, and pulmonary edema in lungs of hyperoxic mice. P326TAT can be a novel therapeutic agent to treat or prevent acute lung injury in oxygen toxicity.     

Pulmonary oxygen toxicity: Lack of tolerance of mice of various ages

Prolonged continuous exposure of adult (3–4 months) and old (21 months) mice to hyperoxia did not lead to significant changes in the activities of superoxide dismutase and catalase in liver or blood. Lung superoxide dismutase activity increased by 25% during initial exposure to 100% O₂, but then fell progressively to below control level. Exposure of mice to 60% or 80% O₂ increased their susceptibility to further exposure to 100% O₂. The results clearly show that both adult and old mice are incapable of coping with the high oxygen environment and that antioxidant enzyme induction and the associated partial protection from pulmonary O₂ toxicity are not the general rule in mammalian lung exposed to subtoxic oxygen levels.     

Assessment of antioxidant defense system responses in the hepatopancreas of the freshwater crab Sinopotamon henanense exposed to lead

Lead (Pb) is a common pollutant in aquatic ecosystems, which produces a wide range of toxic biochemical effects in different organisms. The aim of the present study was to explore the mechanisms of acute toxicity and antioxidant defenses in freshwater crab Sinopotamon henanense induced by Pb. Hepatopancreas was collected from S. henanense exposed for 96 h to Pb (0, 9.188, 18.375, 36.75, 73.5, and 147 mg/l). Oxidative stress was examined using a suite of assays in crabs, including contents of hydrogen peroxide (H₂O₂) and malondialdehyde (MDA), activities of antioxidative enzymes such as superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx), and glutathione system-related parameters such as glutathione (GSH), glutathione S-transferase (GST), and glutathione reductase (GR). A dose-dependent increase of H₂O₂ and MDA was observed in the crabs after Pb exposure, while antioxidative enzymes activities were suppressed significantly (P < 0.05) at higher concentrations of Pb. The ratios of CAT/SOD, GPx/SOD, and GR/GPx were also suppressed. Our results suggested that acute exposure of Pb causes lipid peroxidation and harmful lessened antioxidant defenses of crabs. The above parameters were evaluated as potential biomarkers for Pb pollution monitoring and health assessment of crabs which is also important for the aquaculture of crabs.