Tracheal insufflation of tumor necrosis factor protects rats against oxygen toxicity

Tracheal insufflation of tumor necrosis factor (TNF; 5 micrograms or 1.2 x 10(5) U) markedly enhanced the survival of adult rats exposed to 100% O2: 12 of 17 rats (71%) survived for greater than 11 days, whereas 30 of 30 control (Hanks' balanced salt solution) insufflated rats (100%) died within 3 days of O2 exposure. Insufflation of gamma-interferon (5 micrograms) or intraperitoneal injection of up to 40 micrograms TNF did not afford any protection. At 55 h after O2 exposure, TNF-insufflated rats showed less pulmonary edema, as determined by the extravascular lung water content-to-bloodless lung dry weigh ratio and less alveolar capillary leak as determined by the protein content in the bronchoalveolar lavage fluid, than control insufflated rats similarly exposed. This protection against O2 toxicity by TNF insufflation was associated with increased lung superoxide dismutase, catalase, and glutathione peroxidase activities. The enhancement of lung antioxidant enzyme activities was noted at 55 h of O2 exposure, when control animals began to die of O2 toxicity. This temporal relationship suggests that TNF-induced increase in antioxidant enzyme activities contributes, at least in part, to the observed protection.     

Erythrocyte insufflation-induced protection against oxygen toxicity: role of cytokines.

We studied the pulmonary response of adult rats to erythrocyte (RBC) and RBC lysate insufflation to define the mechanism of RBC insufflation-induced protection against oxygen toxicity. Tracheal insufflation of 1 ml RBC (75%) or RBC lysate induced an intense pulmonary inflammatory response. Within 24 h of oxygen exposure, greater than 95% of insufflated RBCs was hemolyzed. The cell-free fraction of alveolar lavage fluids from RBC- or RBC lysate-insufflated rats had similar capacity in protecting endothelial cells against H2O2 cytotoxicity. However, RBC insufflation but not RBC lysate insufflation, protected rats against oxygen toxicity. There was marked erythrophagocytosis by alveolar macrophages of RBC-insufflated rats. Insufflation of RBCs, but not RBC lysate, resulted in production of tumor necrosis factor and interleukin 1, which could be recovered by bronchoalveolar lavages. When rats were insufflated with a combination of RBC lysate and cytokines at dosages within the range of cytokine levels achievable in alveolar lavage fluids by RBC insufflation, they became tolerant to oxygen. These results suggest that endogenously produced tumor necrosis factor and interleukin-1 as a result of RBC insufflation may play an important role in RBC insufflation-induced oxygen tolerance.     

Endotoxin-tolerant rats are still protected from oxygen toxicity by low-dose endotoxin treatment

To determine if we could reduce endotoxin's potential for toxicity, we produced "endotoxin-tolerant" rats by administering progressively increasing daily doses of endotoxin (10 ng, 100 ng, 1 microgram, 10 micrograms/kg). This dosage regimen produced a high degree of tolerance to the toxic actions of endotoxin: whereas only 3/17 (18%) of control rats survived a normally lethal dose of endotoxin (25 mg/kg), survival for the endotoxin-tolerant rats was 16/16. When endotoxin-tolerant rats received a standard protective dose of 500 micrograms/kg endotoxin just before transfer to 96-98% O2, 19/20 survived the 72-h exposure period vs. 20-30% survival for controls. Thus whereas the endotoxin-tolerant state blocked the tested lethal and toxic effects of endotoxin, it did not nullify the O2 protective action of endotoxin. In addition, endotoxin's stimulatory effects on the lung antioxidant enzymes in the 96-98% O2-exposed rats was also not blocked by the endotoxin-tolerant state. Thus the therapeutic ratio (TR) of endotoxin as an experimental pharmacological treatment against O2-induced lung damage has been markedly enhanced (TR = ratio of dose producing beneficial effects to dose producing toxic effects).     

Elevation of lung glutathione by oral supplementation of L-2-oxothiazolidine-4-carboxylate protects against oxygen toxicity in protein-energy malnourished rats.

The objectives of this study were to investigate whether oral supplementation of L-2-oxothiazolidine-4-carboxylate (OTC) is effective for increasing tissue glutathione (GSH) concentrations in rats fed a diet very low (0.5%) in protein-a model of wasting malnutrition-and to determine the efficacy of OTC for protection against pulmonary oxygen toxicity. Weanling rats, fed a 0.5 or 15% protein diet for 2 wk, were given an oral supplement of OTC, and tissue GSH concentrations were measured over a 24 h period. OTC supplementation to rats fed 0.5% protein significantly increased GSH concentrations in liver and lung, but not in kidney and blood, when compared with the 0.5% protein unsupplemented group. The liver GSH concentration in the 0.5% protein OTC-supplemented group was higher than the 15% control group. Daily supplementation of OTC protected rats from pulmonary oxygen toxicity during 4 days of 85% oxygen exposure as determined by lung-to-body weight ratios and in vivo proton magnetic resonance imaging. Although hyperoxia exposure increased lung GSH concentrations in all groups, OTC supplementation was effective for increasing lung GSH concentration in rats fed the 0.5% protein diet. This study demonstrated that oral administration of OTC to wasting malnourished rats is an effective procedure to increase GSH concentration rapidly in target organs such as lung, and that daily supplementation of a low dose of OTC has a sustained effect to protect against pulmonary oxygen toxicity during 4 days of hyperoxia exposure.     

Effects of bacterial endotoxin on protecting copper-deficient rats from hyperoxia

The administration of very low doses of bacterial endotoxin protects rats during exposure to hyperoxia and is associated with the induction of lung antioxidant enzyme activities. Copper-deficient rats have increased susceptibility to O2 toxicity, which may be related to their decreased lung superoxide dismutase activity (SOD) or decreased plasma ceruloplasmin concentrations. To determine whether endotoxin can protect against hyperoxia in this susceptible model, we exposed copper-deficient and control rats to a fractional inspiratory concentration of O2 greater than 0.95 for 96 h after pretreatment with 500 micrograms/kg of bacterial endotoxin or phosphate-buffered saline (PBS). Mortality in the copper-deficient and control rats given PBS and exposed to O2 for 96 h was 100%. Copper-deficient rats died significantly earlier during the exposure than controls. No mortality occurred in either group treated with endotoxin and hyperoxia despite the decreased activity of copper-dependent enzymes in the copper-deficient rats. Copper-deficient rats treated with endotoxin and exposed to hyperoxia did increase lung Cu-Zn-SOD activity, but activity remained below levels found in air-exposed controls. Mn-SOD activity was found to be induced above air-exposed controls in the copper-deficient rats treated with endotoxin and exposed to hyperoxia. Hyperoxic exposure resulted in a marked increase in plasma ceruloplasmin concentrations in the control rats, but no increases in ceruloplasmin occurred in the copper-deficient animals. Endotoxin protects copper-deficient rats from hyperoxia despite their decreased lung Cu-Zn-SOD activity, and decreased plasma ceruloplasmin.     

Protection against pulmonary oxygen toxicity by interleukin-1 and tumor necrosis factor: Role of antioxidant enzymes and effect of cyclooxygenase inhibitors

Rats injected with interleukin-1 (10 g) and tumor necrosis factor (10 g) and then exposed continuously to hyperoxia (> 99% O₂, 1 atm) survived longer, had increased lung reduced/oxidized glutathione ratios, smaller pleural effusions, less pulmonary hypertension and improv+++ed arterial blood gases. The percentage of animals surviving for 72 hours in hyperoxia increased from 8% to 94%. Although relatively small increases in glutathione redox cycle enzymes occurred four and sixteen hours following cytokine injection, dramatic increases in all major antioxidant enzymes including superoxide dismutase, glucose-6-phosphate dehydrogenase, glutathione reductase, glutathione peroxidase, and catalase had occurred following 72 hours of exposure to hyperoxia. The protective effect of IL-1 + TNF against lethal pulmonary O₂ toxicity could be partially inhibited by pre-injection of lysine acetylsalicylate or, less effectively, of ibuprofen.Recent studies have suggested that both IL-1 and TNF can induce manganese (mitochondrial) superoxide dismutase mRNA and protein synthesis in a variety of cell types. Preliminary studies suggest that IL-1 alone, in ample dosage, can provide protection against lethal pulmonary O₂ toxicity. Future studies should be directed toward identification of acute phase changes in lung antioxidant enzymes, surfactant proteins and/or lipid components, enzymes needed for synthesis of surfactant phospholipids, and/or other protective proteins. Additional work also needs to be done in identifying the lung cell types in which early enzyme induction occurs. These studies should provide a better understanding of mechanisms whereby protection against pulmonary O₂ toxicity can occur. An understanding of the molecular mechanisms inducing protective proteins should lead to more precise pharmacologic control of these processes.     

Apolipoprotein E protects against oxidative stress in mixed neuronal-glial cell cultures by reducing glutamate toxicity

Apolipoprotein E (ApoE) deficiency has been shown to adversely affect outcome after transient cerebral ischemia and head trauma. Since oxidative stress contributes to these injuries, the ability of ApoE to reduce irreversible oxidative damage was studied in primary mixed neuronal-glial cell cultures. Cells (13-16 days in vitro) were exposed to 50 microM hydrogen peroxide (H2O2) for 30 min, and toxicity was determined by the release of lactate dehydrogenase (LDH) 24 h after exposure. The presence of recombinant human ApoE2 (100, 300, or 1000 nM) in the culture media partially protected against oxidative injury. This protection was not reversed by pre-treatment with receptor associated protein. The NMDA receptor antagonist, MK-801, also provided partial protection against H2O2 toxicity. The degree of protection was similar to that conferred by ApoE treatment. The protective effects of ApoE and MK-801 were not additive; no ApoE protection was observed in cultures treated with MK-801 prior to H2O2 exposure. ApoE treatment had no effect on H2O2 stimulated glutamate release, but did increase the rate of glutamate uptake via the high affinity glutamate transporter in H2O2 treated cultures. Pre-treatment with ApoE also conferred partial protection against glutamate-induced LDH release. Taken together, these findings suggest that ApoE protects mixed neuronal-glial cell cultures against irreversible oxidative injury from H2O2 by reducing secondary glutamate excitotoxicity.     

Pulmonary and systemic inflammatory responses to intra-amniotic IL-1α in fetal sheep

Clinical and epidemiological studies implicate IL-1 as an important mediator of perinatal inflammation. We tested the hypothesis that intra-amniotic IL-1α would induce pulmonary and systemic fetal inflammatory responses. Sheep with singleton fetuses were given an intra-amniotic injection of recombinant sheep IL-1α (100 μg) and were delivered 1, 3, or 7 days later, at 124 ± 1 days gestation (n=5-8/group). A separate group of sheep were given two intra-amniotic IL-1α injections (100 μg dose each): 7 days and again 1 day prior to delivery. IL-1α induced a robust increase in monocytes, neutrophils, lymphocytes, and IL-8 protein in bronchoalveolar lavage fluid. H(2)O(2) secretion was increased in inflammatory cells isolated from lungs of IL-1α-exposed lambs upon LPS challenge in vitro compared with control monocytes. T lymphocytes were recruited to the lung. IL-1β, cyclooxygenase-1, and cyclooxygenase-2 mRNA expression increased in the lung 1 day after intra-amniotic IL-1α exposure. Lung volumes increased 7 days after intra-amniotic IL-1α exposure, with minimal anatomic changes in air space morphology. The weight of the posterior mediastinal lymph node draining the lung and the gastrointestinal tract doubled, inducible nitric oxide synthase (NOSII)-positive cells increased, and Foxp3-positive T-regulatory lymphocytes decreased in the lymph node after IL-1α exposure. In the blood, neutrophil counts and plasma haptoglobin increased after IL-1α exposure. Compared with a single exposure, exposure to intra-amniotic IL-1α 7 days and again 1 day before delivery had a variable effect (increases in some inflammatory markers, but not pulmonary cytokines). IL-1α is a potent mediator of the fetal inflammatory response syndrome.     

Dexamethasone protects against high-dose endotoxin without loss of tolerance to oxygen

Endotoxin (500 micrograms/kg)-treated rats are very tolerant to hyperoxia (greater than 95% O2, 1 ATA). We have now attempted to determine if dexamethasone given to rats 1 h before a usually lethal dose of endotoxin would diminish endotoxin's lethality without substantially abrogating its capacity to confer tolerance to hyperoxia. Endotoxin (20 mg/kg) given alone killed 70-80% of air- or O2-breathing rats within 24 h; dexamethasone (0.6 mg) given 1 h before endotoxin decreased mortality at 24 h to 10-15%. About 90% of the rats that were alive 24 h after receiving dexamethasone plus endotoxin (20 mg/kg) survived 72 h of hyperoxia. Dexamethasone plus endotoxin (10 mg/kg) provided as much protection against pulmonary edema resulting from 72 h of hyperoxia as did 500 micrograms/kg endotoxin alone. Tolerance to hyperoxia produced by dexamethasone plus high-dose endotoxin was accompanied by a rise in the activity in the lung of antioxidant enzymes. We conclude that dexamethasone protects rats against the lethal effects of high doses of endotoxin without interfering with endotoxin's capacity to engender tolerance to hyperoxia.     

Hyperoxic inhibition of newborn rat lung development: protection by deferoxamine.

Prolonged exposure to hyperoxia markedly inhibits normal lung development (alveolarization and respiratory surface area expansion) in immature animals. Since (a) hyperoxia results in excess hydroxyl radical (OH.) formation, (b) (OH.) is implicated in O2-induced lipid peroxidation and DNA alterations, and (c) both OH. formation and its interaction with DNA are Fe++ dependent; chelation of Fe++ should act to protect against pulmonary O2 toxicity and hyperoxic inhibition of lung development. We therefore treated litters of newborn rats with the iron chelator Deferoxamine mesylate (DES) (150 mg/kg/day) during a 10-day exposure to greater than 95% O2. Morphometric analysis demonstrated that compared to the mean airspace size in air control rat pups (Lm = 44.5 microns), hyperoxic exposure resulted in a 34% larger mean air space diameter in O2-saline rat lungs (59.5 microns) versus only an 11% enlargement in O2-DES lungs (51.1 microns*). Lung internal surface area (cm2) per 100-g body weight were air control = 4480, O2-saline = 3570 (decreases 20.3%), and O2-DES = 4125* (decreases 7.9%) (*p less than 0.05 versus O2-saline group). DES-treated animals also had significantly decreased lung conjugated diene levels during hyperoxic exposure and increased lung elastin content (reflective of preserved lung alveolar formation) compared to O2-saline rats. These results indicate that DES treatment substantially ameliorated the inhibitory effects of neonatal hyperoxic exposure on normal lung development.     

Development of O2 tolerance in rabbits with no increase in antioxidant enzymes

Instillation of exogenous surfactant into rabbits exposed to 100% O2 increases survival time and decreases alveolar epithelial injury. In this study we investigated whether rabbits with increased levels of endogenous pulmonary surfactant are more resistant to hyperoxia. Rabbits were exposed to 100% O2 for 64 h and then returned to room air for 8 days (preexposed). At this time, they had normal gas exchange and alveolar permeability to solute and increased levels of lavageable alveolar phospholipids compared with control rabbits breathing air (26 +/- 2 vs. 12 +/- 2 mumol/kg). Preexposed rabbits survived significantly longer than control rabbits when reexposed to 100% O2 (166 +/- 24 vs. 80 +/- 6 h; n = 7; P less than 0.05) and had significantly higher values of total lavageable phospholipids after 72 h in 100% O2 (15 +/- 2 vs. 5 +/- 2 mumol/kg). Controls developed arterial hypoxemia after 72 h in 100% O2. On the other hand, preexposed rabbits maintained arterial PO2 values greater than 100 Torr throughout the hyperoxic exposure and developed progressive respiratory acidosis. Specific activities of CuZn and Mn superoxide dismutase, catalase, and glutathione peroxidase in lung homogenates and isolated alveolar type II pneumocytes of preexposed rabbits were unchanged from those of controls before O2 reexposure and after 72 h in 100% O2. We concluded that 1) increases in pulmonary antioxidant enzyme specific activities are not necessary for the development of O2 tolerance in rabbits and 2) pulmonary surfactant may play a role in O2 adaptation.     

Effect of 0.25 ppm ozone exposure on pulmonary damage induced by bleomycin

To study the effect of ozone in a chronically damaged lung, we used a bleomycin (BLM) induced pulmonary fibrosis model. Both endotracheal instillation of BLM and O3 exposure both produce lung inflammation and fibrosis. Oxidative stress would be a common mechanism of damage for both BLM and O3. Our aim was to assess lung injury induced by 5 and 60 days of intermittent exposure to 0.25 ppm O3 in rats with bleomycin-induced pulmonary fibrosis. Thirty-day-old Sprague Dawley rats were endotracheally instilled with BLM (1 U/100 g body weight) and, 30 days later, exposed to 0.25 ppm 03 (0.25 ppm 4 h per day, 5 days a week). Histopatology controls were instilled with saline and breathing room air. Histopathological evaluation of lungs was done 5 and 60 days after O3 exposure. BLM-induced lung damage did not change after 60 days of intermittent O3 exposure. Five days of O3 exposure increased the mean score of BLM-induced pulmonary inflammation and fibrosis (p=0.06). Frequency of bronchopneumonia increased from 1/7 to 6/6 (p <0.001), suggesting that a short-term exposure to O3 in a previously damaged lung might be a risk factor for developing further lung injury.     

Inhaled carbon monoxide and hyperoxic lung injury in rats

Because carbon monoxide (CO) has been proposed to have anti-inflammatory properties, we sought protective effects of CO in pulmonary O(2) toxicity, which leads rapidly to lung inflammation and respiratory failure. Based on published studies, we hypothesized that CO protects the lung against O(2) by selectively increasing expression of antioxidant enzymes, thereby decreasing oxidative injury and inflammation. Rats exposed to O(2) with or without CO [50-500 parts/million (ppm)] for 60 h were compared for lung wet-to-dry weight ratio (W/D), pleural fluid volume, myeloperoxidase (MPO) activity, histology, expression of heme oxygenase-1 (HO-1), and manganese superoxide dismutase (Mn SOD) proteins. The brains were evaluated for histological evidence of damage from CO. In O(2)-exposed animals, lung W/D increased from 4.8 in normal rats to 6.3; however, only CO at 200 and 500 ppm decreased W/D significantly (to 5.9) during O(2) exposure. Large volumes of pleural fluid accumulated in all rats, with no significant CO treatment effect. Lung MPO values increased after O(2) and were not attenuated by CO treatment. CO did not enhance lung expression of oxidant-responsive proteins Mn SOD and HO-1. Animals receiving O(2) and CO at 200 or 500 ppm showed significant apoptotic cell death in the cortex and hippocampus by immunochemical staining. Thus significant protection by CO against O(2)-induced lung injury could not be confirmed in rats, even at CO concentrations associated with apoptosis in the brain.     

Interleukin-1beta swiftly down-regulates UCP-2 mRNA in beta-cells by mechanisms not directly coupled to toxicity

Regulation of uncoupling protein-2 (UCP-2) in beta-cells is presently unclear but may involve oxidative stress. We tested for regulation by beta-cell toxic cytokines. Exposure to interleukin-1beta (IL-1beta, 10 ng/ml) for 6 h down-regulated UCP-2 mRNA in clonal INS-1 cells, by 37 +/- 7%, and in rat pancreatic islets, by 55 +/- 8%. In contrast, a 6 h exposure to IL-1beta did not affect viability as assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, or mitochondrial membrane potential, or ATP cellular contents. Continued exposure to IL-1beta was accompanied by decreased viability and persisting down-regulation of UCP-2 mRNA. Exposure to a combination of IL-1beta and tumor necrosis factor (TNF)-alpha for 48 h additively decreased cell viability and UCP-2 mRNA. The constitutive nitric oxide (NO) synthase inhibitor N-omega-nitro-L-arginine methyl ester (L-NAME, 1 mM) partially protected against toxicity but failed to significantly affect UCP-2 mRNA expression. The inducible NO synthase inhibitor N(G)-monomethyl-L-arginine (L-NMMA, 1 mM) protected completely against cytokine-induced toxicity. L-NMMA per se down-regulated UCP-2 mRNA (by 64 +/- 7%). Transfection with a UCP-2-antisense nucleotide failed to affect IL-1beta induced toxicity. In conclusion, down-regulation of UCP-2 mRNA by IL-1beta is an early event of cytokine interaction with beta-cells which is not directly coupled to toxicity.     

Manganese superoxide dismutase is induced by IFN-gamma in multiple cell types. Synergistic induction by IFN-gamma and tumor necrosis factor or IL-1

We have previously characterized more than 20 proteins induced by the immunoregulatory lymphokine IFN-gamma in human fibroblasts by their m.w. and isoelectric points determined in two-dimensional gels. Some of these proteins are induced uniquely by IFN-gamma, whereas others are also induced by IFN-alpha, TNF, or IL-1. Recent technologic advances have allowed us to begin to rapidly identify proteins induced by IFN-gamma and other cytokines by sequencing the induced proteins from blots of preparative two-dimensional gels of total cell lysates. In this study, we show that the approximately 21 kDa, isoelectric point greater than 7 protein induced by IFN-gamma is manganese superoxide dismutase (Mn-SOD), a mitochondrial protective enzyme encoded by a nuclear gene. Mn-SOD is induced by IFN-gamma and also by TNF in all four human cell lines examined: HS153 fibroblasts, ACHN renal carcinoma, A549 lung carcinoma, and A375 melanoma. Induction of Mn-SOD mRNA is a primary, rapid, and dose-dependent response to IFN-gamma. In ACHN renal carcinoma cells, Mn-SOD mRNA and protein are induced synergistically by IFN-gamma in combination with either TNF or IL-1, and the induced protein is enzymatically active. IFN-gamma and TNF together induce Mn-SOD mRNA by more than 100-fold relative to its level in untreated ACHN cells. The induction of Mn-SOD by IFN-gamma and its synergistic induction by IFN-gamma in combination with TNF and IL-1 should protect healthy cells from the toxicity of O2- during an immune response, and may provide a mechanism for selective killing of infected cells.     

A role for platelet-derived growth factor beta-receptor in a newborn rat model of endothelin-mediated pulmonary vascular remodeling

Newborn rats exposed to 60% O2 for 14 days develop endothelin (ET)-1-dependent pulmonary hypertension with vascular remodeling, characterized by increased smooth muscle cell (SMC) proliferation and medial thickening of pulmonary resistance arteries. Using immunohistochemistry and Western blot analyses, we examined the effect of exposure to 60% O2 on expression in the lung of receptors for the platelet-derived growth factors (PDGF), which are implicated in the pathogenesis of arterial smooth muscle hyperplasia. We observed a marked O2-induced upregulation of PDGF-alpha and -beta receptors (PDGF-alphaR and -betaR) on arterial smooth muscle. This led us to examine pulmonary vascular PDGF receptor expression in 60% O2-exposed rats given SB-217242, a combined ET receptor antagonist, which we found prevented the O2-induced upregulation of PDGF-betaR, but not PDGF-alphaR, on arterial smooth muscle. PDGF-BB, a major PDGF-betaR ligand, was found to be a potent in vitro inducer of hyperplasia and DNA synthesis in cultured pulmonary artery SMC from infant rats. A critical role for PDGF-betaR ligands in arterial SMC proliferation was confirmed in vivo using a truncated soluble PDGF-betaR intervention, which attenuated SMC proliferation induced by exposure to 60% O2. Collectively, these data are consistent with a major role for PDGF-betaR-mediated SMC proliferation, acting downstream of increased ET-1 in a newborn rat model of 60% O2-induced pulmonary hypertension.     

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.     

Therapeutic hypercapnia prevents chronic hypoxia-induced pulmonary hypertension in the newborn rat

Induction of hypercapnia by breathing high concentrations of carbon dioxide (CO(2)) may have beneficial effects on the pulmonary circulation. We tested the hypothesis that exposure to CO(2) would protect against chronic pulmonary hypertension in newborn rats. Atmospheric CO(2) was maintained at <0.5% (normocapnia), 5.5%, or 10% during exposure from birth for 14 days to normoxia (21% O(2)) or moderate hypoxia (13% O(2)). Pulmonary vascular and hemodynamic abnormalities in animals exposed to chronic hypoxia included increased pulmonary arterial resistance, right ventricular hypertrophy and dysfunction, medial thickening of pulmonary resistance arteries, and distal arterial muscularization. Exposure to 10% CO(2) (but not to 5.5% CO(2)) significantly attenuated pulmonary vascular remodeling and increased pulmonary arterial resistance in hypoxia-exposed animals (P < 0.05), whereas both concentrations of CO(2) normalized right ventricular performance. Exposure to 10% CO(2) attenuated increased oxidant stress induced by hypoxia, as quantified by 8-isoprostane content in the lung, and prevented upregulation of endothelin-1, a critical mediator of pulmonary vascular remodeling. We conclude that hypercapnic acidosis has beneficial effects on pulmonary hypertension and vascular remodeling induced by chronic hypoxia, which we speculate derives from antioxidant properties of CO(2) on the lung and consequent modulating effects on the endothelin pathway.     

Induction of Mn-superoxide dismutase by tumor necrosis factor, interleukin-1 and interleukin-6 in human hepatoma cells.

Effects of Tumor Necrosis Factor (TNF), Interleukin-1 (IL-1), Interleukin-6 (IL-6) and Interferon-gamma (IFN-gamma) on the expression of Mn-superoxide dismutase (Mn-SOD) protein were investigated in human hepatoma cells, Hu-H1, which revealed resistance to the cytotoxicity of TNF and IL-1. Both TNF and IL-1 enhanced the Mn-SOD production to the level of 30- to 40-fold. IL-6 also increased the enzyme protein to 2- to 3-fold of the basal level without any cell proliferative effect. A specific antibody against IL-6 almost completely inhibited the induction of Mn-SOD. IL-6, as well as TNF and IL-1, appears to play some role in the Mn-SOD protein expression in human hepatoma cells.     

Characterization of LY231617 protection against hydrogen peroxide toxicity

The compound LY231617 [2,6-bis(1,1-dimethylethyl)-4-[[(1-ethyl)amino]methyl]phenol hydrochloride] has been reported to afford significant neuroprotection against hydrogen peroxide (H2O2)-induced toxicity in vitro and global ischemia in vivo. We now report on further mechanistic studies of H2O2 toxicity and protection by LY231617. Brief exposure to H2O2 (15 min) elicited an oxidative insult comparable with that generated by overnight treatment. H2O2-mediated cellular degeneration was characterized using lactate dehydrogenase (LDH) release, changes in total glutathione, and a new marker of oxidative stress, 8-epiprostaglandin F2alpha (8-isoprostane). LY231617 attenuated H2O2-mediated degeneration under a variety of exposure conditions, including a more clinically relevant posttreatment paradigm. Levels of 8-isoprostane paralleled LDH release under various treatment paradigms of 100 microM H2O2 +/- 5 microM drug. In contrast, despite affording significant protection, LY231617 had modest to no effects on cellular levels of glutathione. Taken together, these results are consistent with a membrane site of action for LY231617 and suggest that the compound affords cytoprotection via its antioxidant properties.