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.     

Role of metallothionein in lung inflammation induced by ozone exposure in mice

Metallothionein (MT) is a free radical scavenger induced by inflammatory stimuli; however, its roles in inflammation have not been fully investigated. In the present study, we genetically determined the role of MT in ozone (O₃)-induced lung inflammation using MT-I/II null (–/–) mice. Subacute (65 h) exposure to O₃ (0.3 ppm) induced lung inflammation and enhanced vascular permeability, which was significantly greater in MT(–/–) than in corresponding wild-type mice. Electron microscopically, O₃ exposure induced vacuolar degeneration of pulmonary endothelial and epithelial cells, and interstitial edema with focal loss of the basement membrane, which was more prominent in MT(–/–) than in wild-type mice. O₃ -induced lung expression of interleukin-6 was significantly greater in MT(–/–) than in wild-type mice; however, lung expression of the chemokines examined was comparable in both genotypes of mice in the presence of O₃. Following O₃ exposure, the formation of oxidative stress-related molecules/adducts, such as heme oxidase-1, inducible nitric oxide synthase, 8-hydroxy-2′-deoxyguanosine, and nitrotyrosine, in the lung was significantly greater in MT(–/–) than in wild-type mice. Collectively, MT protects against O₃-induced lung inflammation, at least partly, via the regulation of pulmonary endothelial and epithelial integrity and its antioxidative property.     

Alteration of intracellular secretory acute phase response proteins expressed in human hepatocyte induced by exposure with interleukin-6

Interleukin-6 (IL-6) is a principal proinflammatory cytokine inducing the acute phase response in various tissues, including liver. Here, we adopt the FD-LC-MS/MS method, consisting of fluorogenic derivatization (FD), separation by liquid chromatography (LC), and identification of proteins by LC-tandem mass spectrometry (MS/MS), to reveal how exposure to IL-6 alters temporally the intracellular secretory acute phase response (sAPR) proteins expressed in human hepatocytes as compared to non-exposure. Nine altered sAPR proteins were identified in cultures in response to IL-6. Seven of them (serum amyloid A protein, haptoglobin, fibrinogen α chain, fibrinogen β chain, fibrinogen γ chain, α(1)-acid glycoprotein and α(1)-antitrypsin) were significantly increased and two (β(2)-glycoprotein 1 and transferrin) were significantly decreased in response to IL-6. In addition, the transmission speed of transferrin might be much faster than the other sAPR proteins. These results suggest a different molecular mechanism for protein synthesis and the secretory pathway among the sAPR proteins. In this study, we observed the simultaneously and temporally altered expression of sAPR proteins which had been induced by exposure to IL-6 in human hepatocytes, in contrast to previous reports, in all of which the proteins were tested from the time they were secreted into the medium from the cells.     

Pulmonary inflammation induced by subacute ozone is augmented in adiponectin-deficient mice: role of IL-17A

Pulmonary responses to ozone, a common air pollutant, are augmented in obese individuals. Adiponectin, an adipose-derived hormone that declines in obesity, has regulatory effects on the immune system. To determine the role of adiponectin in the pulmonary inflammation induced by extended (48-72 h) low-dose (0.3 parts per million) exposure to ozone, adiponectin-deficient (Adipo(-/-)) and wild-type mice were exposed to ozone or to room air. In wild-type mice, ozone exposure increased total bronchoalveolar lavage (BAL) adiponectin. Ozone-induced lung inflammation, including increases in BAL neutrophils, protein (an index of lung injury), IL-6, keratinocyte-derived chemokine, LPS-induced CXC chemokine, and G-CSF were augmented in Adipo(-/-) versus wild-type mice. Ozone also increased IL-17A mRNA expression to a greater extent in Adipo(-/-) versus wild-type mice. Moreover, compared with control Ab, anti-IL-17A Ab attenuated ozone-induced increases in BAL neutrophils and G-CSF in Adipo(-/-) but not in wild-type mice, suggesting that IL-17A, by promoting G-CSF release, contributed to augmented neutrophilia in Adipo(-/-) mice. Flow cytometric analysis of lung cells revealed that the number of CD45(+)/F4/80(+)/IL-17A(+) macrophages and γδ T cells expressing IL-17A increased after ozone exposure in wild-type mice and further increased in Adipo(-/-) mice. The IL-17(+) macrophages were CD11c(-) (interstitial macrophages), whereas CD11c(+) macrophages (alveolar macrophages) did not express IL-17A. Taken together, the data are consistent with the hypothesis that adiponectin protects against neutrophil recruitment induced by extended low-dose ozone exposure by inhibiting the induction and/or recruitment of IL-17A in interstitial macrophages and/or γδ T cells.     

Impact of interleukin-6 on hypoxia-induced pulmonary hypertension and lung inflammation in mice

Background

Inflammation may contribute to the pathogenesis of various forms of pulmonary hypertension (PH). Recent studies in patients with idiopathic PH or PH associated with underlying diseases suggest a role for interleukin-6 (IL-6).

Methods

To determine whether endogenous IL-6 contributes to mediate hypoxic PH and lung inflammation, we studied IL-6-deficient (IL-6^-/-) and wild-type (IL-6^+/+) mice exposed to hypoxia for 2 weeks.

Results

Right ventricular systolic pressure, right ventricle hypertrophy, and the number and media thickness of muscular pulmonary vessels were decreased in IL-6^-/- mice compared to wild-type controls after 2 weeks'' hypoxia, although the pressure response to acute hypoxia was similar in IL-6^+/+ and IL-6^-/- mice. Hypoxia exposure of IL-6^+/+ mice led to marked increases in IL-6 mRNA and protein levels within the first week, with positive IL-6 immunostaining in the pulmonary vessel walls. Lung IL-6 receptor and gp 130 (the IL-6 signal transducer) mRNA levels increased after 1 and 2 weeks'' hypoxia. In vitro studies of cultured human pulmonary-artery smooth-muscle-cells (PA-SMCs) and microvascular endothelial cells revealed prominent synthesis of IL-6 by PA-SMCs, with further stimulation by hypoxia. IL-6 also markedly stimulated PA-SMC migration without affecting proliferation. Hypoxic IL-6^-/- mice showed less inflammatory cell recruitment in the lungs, compared to hypoxic wild-type mice, as assessed by lung protein levels and immunostaining for the specific macrophage marker F4/80, with no difference in lung expression of adhesion molecules or cytokines.

Conclusion

These data suggest that IL-6 may be actively involved in hypoxia-induced lung inflammation and pulmonary vascular remodeling in mice.     

Effect of acute ozone induced airway inflammation on human sympathetic nerve traffic: a randomized, placebo controlled, crossover study

Background

Ozone concentrations in ambient air are related to cardiopulmonary perturbations in the aging population. Increased central sympathetic nerve activity induced by local airway inflammation may be one possible mechanism.

Methodology/Principal Findings

To elucidate this issue further, we performed a randomized, double-blind, cross-over study, including 14 healthy subjects (3 females, age 22–47 years), who underwent a 3 h exposure with intermittent exercise to either ozone (250 ppb) or clean air. Induced sputum was collected 3 h after exposure. Nineteen to 22 hours after exposure, we recorded ECG, finger blood pressure, brachial blood pressure, respiration, cardiac output, and muscle sympathetic nerve activity (MSNA) at rest, during deep breathing, maximum-inspiratory breath hold, and a Valsalva maneuver. While the ozone exposure induced the expected airway inflammation, as indicated by a significant increase in sputum neutrophils, we did not detect a significant estimated treatment effect adjusted for period on cardiovascular measurements. Resting heart rate (clean air: 59±2, ozone 60±2 bpm), blood pressure (clean air: 121±3/71±2 mmHg; ozone: 121±2/71±2 mmHg), cardiac output (clean air: 7.42±0.29 mmHg; ozone: 7.98±0.60 l/min), and plasma norepinephrine levels (clean air: 213±21 pg/ml; ozone: 202±16 pg/ml), were similar on both study days. No difference of resting MSNA was observed between ozone and air exposure (air: 23±2, ozone: 23±2 bursts/min). Maximum MSNA obtained at the end of apnea (air: 44±4, ozone: 48±4 bursts/min) and during the phase II of the Valsalva maneuver (air: 64±5, ozone: 57±6 bursts/min) was similar.

Conclusions/Significance

Our study suggests that acute ozone-induced airway inflammation does not increase resting sympathetic nerve traffic in healthy subjects, an observation that is relevant for environmental health. However, we can not exclude that chronic airway inflammation may contribute to sympathetic activation.     

Increased pulmonary responses to acute ozone exposure in obese db/db mice

Epidemiological studies indicate the incidence of asthma is increased in obese and overweight humans. Responses to ozone (O(3)), an asthma trigger, are increased in obese (ob/ob) mice lacking the satiety hormone leptin. The long form of leptin receptor (Ob-R(b)) is required for satiety; mice lacking this receptor (db/db mice) are also substantially obese. Here, wild-type (WT) and db/db mice were exposed to air or O(3) (2 ppm) for 3 h. Airway responsiveness, measured by the forced oscillation technique, was greater in db/db than WT mice after air exposure. O(3)-induced increases in pulmonary resistance and airway responsiveness were also greater in db/db mice. BALF eotaxin, IL-6, KC, and MIP-2 increased 4 h after O(3) exposure and subsided by 24 h, whereas protein and neutrophils continued to increase through 24 h. For each outcome, the effect of O(3) was significantly greater in db/db than WT mice. Previously published results obtained in ob/ob mice were similar except for O(3)-induced neutrophils and MIP-2, which were not different from WT mice. O(3) also induced pulmonary IL-1beta and TNF-alpha mRNA expression in db/db but not ob/ob mice. Leptin was increased in serum of db/db mice, and pulmonary mRNA expression of short form of leptin receptor (Ob-R(a)) was similar in db/db and WT mice. These data confirm obese mice have innate airway hyperresponsiveness and increased pulmonary responses to O(3). Differences between ob/ob mice, which lack leptin, and db/db mice, which lack Ob-R(b) but not Ob-R(a), suggest leptin, acting through Ob-R(a), can modify some pulmonary responses to O(3).     

The role of CCR7 in allergic airway inflammation induced by house dust mite exposure

House dust mite (HDM), the most common allergen, activate both the IgE-associated and innate immune responses. To clarify the process of sensitization, we investigated the role of the CCL21, CCL19, and CCR7 axis in a mouse model of HDM-induced allergic asthma. HDM inhalation without systemic immunization resulted in a HDM-specific IgE response. CCR7-knockout (CCR7KO) mice exhibited greater airway inflammation and IgE responses compared to wild-type mice. We examined FoxP3 expression in these mice to clarify the contribution of regulatory cells to the responses. FoxP3 expression was higher in the lungs but not in the lymph nodes of CCR7KO mice compared to wild-type mice. In CCR7KO mice, FoxP3-positive cells were found in lung, but we observed higher release of IL-13, IL-5, TGF-β, IL-17, and HMGB1 in bronchoalveolar lavage fluid. We demonstrate here that immuno-regulation through CCR7 expression in T cells plays a role in HDM-specific sensitization in the airway.     

Manipulation of inflammation modulates hyperlipidemia in apolipoprotein E-deficient mice: a possible role for interleukin-6

There are increasing evidences showing that inflammation participates in atherosclerosis. Therefore, the therapeutic use of anti-inflammatory agents should be considered. We have induced chronic, aseptic inflammation upon the injection of turpentine and tested the effect of dexamethasone on lipoprotein metabolism and, consequently, atherosclerosis in apolipoprotein E-deficient mice. Aseptic inflammation caused a significant decrease in hyperlipidemia. Treatment with dexamethasone elicited the opposite effect increasing hyperlipidemia through mechanisms related to the increase in the synthesis of triglyceride-rich lipoproteins. Changes in plasma lipids correlated with those observed in the size of atherosclerotic lesions. Our data suggest the presence of a common mechanism present in both observations and which is probably related to the cytokine secretion. Among the candidates, we chose to test the effect of interleukin-6 because it is involved in both processes, atherosclerosis and inflammation, and its expression is efficiently repressed by corticosteroids. The injection of recombinant interleukin-6 in our mice elicited the same effects observed in our model of inflammation. We conclude that manipulation of inflammation-related mechanisms modulates lipid homeostasis and development of atherosclerotic plaque in rodents.     

The wolf in sheep's clothing: the role of interleukin-6 in immunity, inflammation and cancer

Recent discoveries involving the cytokine interleukin (IL)-6 have originated from diverse disciplines, revealing roles in biological processes that are likewise varied. The most novel findings suggest a connection between inflammation and diseases, such as insulin resistance associated with diabetes mellitus and cancer, which had not or only weakly been appreciated previously. The IL-6 pathway is one of the mechanisms linking inflammation to these disease processes. In addition, new evidence points toward IL-6 as one of the mediators coordinating the interface between adaptive and innate immunity. Here, we review the evidence linking IL-6 to inflammatory diseases and cancer.     

Effects of pulmonary exposure to carbon nanotubes on lung and systemic inflammation with coagulatory disturbance induced by lipopolysaccharide in mice

Despite intensive research as to the pathogenesis of lipopolysaccharide (LPS)-related inflammation with coagulatory disturbance, their exacerbating factors have not been well explored. This study examined the effects of pulmonary exposure to two types of nano-sized materials (carbon nano-tubes: CNT [single-wall: SWCNT, and multi-wall: MWCNT]) on lung inflammation and consequent systemic inflammation with coagulatory disturbance induced by pulmonary exposure to LPS in mice and their cellular mechanisms in vitro. ICR male mice were divided into 6 experimental groups that intra-tracheally received the vehicle, two types of CNT (4 mg/kg), LPS (33 mu g/kg), or LPS plus either type of CNT. Twenty-four hours after treatment, both types of CNT alone induced lung inflammation with enhanced lung expression of proinflammatory cytokines, but did not synergistically exacerbate lung inflammation elicited by LPS. SWCNT significantly induced/ enhanced pulmonary permeability and hyperfibrinogenemia and reduced activated protein C in the absence or presence of LPS, whereas MWCNT did moderately. Both CNT moderately, but not significantly, elevated circulatory levels of proinflammatory cytokines and chemokines. In the presence of LPS, CNT tended to elevate the levels of the mediators with an overall trend, which was more prominent with SWCNT than with MWCNT. In vitro study showed that both CNT amplified LPS-induced cytokine production from peripheral blood monocytes. These results suggest that CNT can facilitate systemic inflammation with coagulatory disturbance, at least in part, via the activation of mononuclear cells, which is accompanied by moderate enhancement of acute lung inflammation related to LPS.     

The human haptoglobin gene promoter: interleukin-6-responsive elements interact with a DNA-binding protein induced by interleukin-6

[This corrects the article on p. 1145 in vol. 8, PMID: 2787245.].     

Effect of ozone exposure on prostacyclin synthesis in lung

The effect of ozone exposure on prostacyclin (PGI2) synthesis in the rat lung was studied. Male Wistar rats were exposed to 0.2, 0.4, 0.8, 1.2 and 1.8 ppm ozone for 24h. The higher concentration (1.8 ppm) significantly depressed-PGI2 synthesizing activity of lung homogenates. Time-courses (1, 3, 5, 7, 14 and 28 days) of the effect of ozone (0.4 and 0.8 ppm) exposure on the PGI2-synthesizing activity of lung homogenates were studied. The PGI2-synthesizing activity of the lung decreased, reaching a maximum at 5 days and then gradually returning to normal by day 14, and remaining normal at day 28, even though the ozone exposure continued. The formation of lipid peroxides due to ozone exposure may cause the depression of PGI2-synthesizing activity of lung. Induction of anti-oxidative enzymes may relate to the recovery of the PGI2-synthesizing activity.     

Role of interleukin-6 in murine airway responses to ozone

This study sought to examine the role of interleukin-6 (IL-6) in ozone (O(3))-induced airway injury, inflammation, and hyperresponsiveness (AHR). Subacute (72 h) exposure to 0.3 ppm O(3) significantly elevated bronchoalveolar lavage fluid (BALF) protein, neutrophils, and soluble TNF receptors (sTNFR1 and sTNFR2) in wild-type C57BL/6 (IL-6(+/+)) mice; however, all four outcome indicators were significantly reduced in IL-6-deficient (IL-6(-/-)) compared with IL-6(+/+) mice. Acute O(3) exposure (2 ppm for 3 h) increased BALF protein, KC, macrophage inflammatory protein(MIP)-2, eotaxin, sTNFR1, and sTNFR2 in IL-6(+/+) mice. However, MIP-2 and sTNFR2 were not significantly increased following O(3) exposure in IL-6(-/-) mice. Increases in BALF neutrophils induced by O(3) (2 ppm for 3 h) were also significantly reduced in IL-6(-/-) vs. IL-6(+/+) mice. Airway responsiveness to methacholine was measured by whole body plethysmography before and following acute (3 h) or subacute (72 h) exposure to 0.3 ppm O(3). Acute O(3) exposure caused AHR in both groups of mice, but there was no genotype-related difference in the magnitude of O(3)-induced AHR. AHR was absent in mice of either genotype exposed for 72 h. Our results indicate that IL-6 deficiency reduces airway neutrophilia, as well as the levels of BALF sTNFR1 and sTNFR2 following acute high dose and/or subacute low-dose O(3) exposure, but has no effect on O(3)-induced AHR.