Grantee Research Project Results
1999 Progress Report: Mechanisms of Age-dependent Ozone Induced Airway Dysfunction
EPA Grant Number: R827447Title: Mechanisms of Age-dependent Ozone Induced Airway Dysfunction
Investigators: Shore, Stephanie , Murthy, G.G. Krishna , Laporte, Johanne , Want, Matt
Current Investigators: Shore, Stephanie , Laporte, Johanne , Murthy, G.G. Krishna , Want, Matt , Johnston, Richard
Institution: Harvard University
EPA Project Officer: Aja, Hayley
Project Period: July 1, 1999 through June 30, 2002 (Extended to July 31, 2003)
Project Period Covered by this Report: July 1, 1999 through June 30, 2000
Project Amount: $852,937
RFA: Children's Vulnerability to Toxic Substances in the Environment (1999) RFA Text | Recipients Lists
Research Category: Children's Health , Human Health
Objective:
Acute exposure to ozone (O3) causes damage to lung and airway epithelial cells, airway inflammation, decreases in lung function and airway hyperresponsiveness (AHR) in human subjects. The mechanistic basis for O3 induced AHR appears to be inflammation arising from oxidant injury to the lungs and airways. However, the precise aspect of the inflammatory cascade required for AHR is still not firmly established. O3 may be a particularly important respiratory hazard for children because they spend more time outdoors where O3 levels are higher, and because they are more physically active, and hence, have higher minute ventilations and, consequently, a higher delivered dose of O3. In addition, animal studies suggest that O3 may be more toxic to the lungs of immature than mature animals.The questions that this project seeks to address are: Is O3-induced AHR greater in juvenile than adult animals, and if so, why? What is the mechanism of O3-induced AHR? Is the mechanistic basis for O3-induced AHR the same or different in juvenile and adult animals? In particular, we propose to test the following hypotheses: (1) age-related differences in ventilation and in the effect of O3 on the ventilatory pattern contribute to age-related differences in O3-induced AHR; (2) TNFa formation induced by O3 is important for O3-induced AHR; and (3) O3-induced TNFa formation is different in juvenile and adult animals.
Progress Summary:
We already have made significant progress in establishing that there are age-related differences in ventilation and in the effect of O3 on the ventilatory pattern. It is known that during O3 exposure, adult rats decrease their minute ventilation (VE). To determine whether such changes also are observed in immature animals, we exposed Sprague Dawley rats, aged 2, 4, 6, 8, or 12 weeks, to O3 (2 ppm for 3 hours) in nose only exposure plethysmographs. Baseline VE normalized for body weight decreased with age from 2.1 + 0.1 ml/min/g in 2-week old rats to 0.72 + 0.03 ml/min/g in 12-week old rats, consistent with the higher metabolic rates of younger animals. In adult (8- and 12-week old) rats, O3 caused 40-50 percent decreases in VE that occurred primarily as the result of a decrease in tidal volume (VT). In 6-week old rats, O3 induced changes in VE were significantly less, and in 2- and 4-week old rats, no significant changes in VE were observed during O3 exposure. The increased baseline VE and the smaller decrements in VE induced by O3 in the immature rats imply that their delivered dose of O3 is about 5 times higher than in adult rats.Although changes in frequency were not consistently observed with O3 exposure in adult animals, there were changes in the timing of ventilation such that Ti decreased and Te increased: the decrease in Ti and the increase in Te were significant within 70 minutes of the initiation of O3 exposure (p<0.01 and p<0.02, respectively). Upon further examination, the increase in Te was found to result primarily from an increase in end expiratory pause (EEP), that occurred within 70 minutes of the onset of O3 exposure. Indeed, in some animals, 1-2 second apneas were occasionally observed during O3 exposure. For each of these ventilatory parameters, Ti, Te, and EEP, the effect of O3 was significantly greater in adult rats compared to each of the 2, 4, and 6 week old groups (p<0.001 in each case).
To determine whether these age-related differences in O3 dose influence the extent of injury, we measured BAL protein concentrations. The magnitude of the changes in BAL protein induced by O3 was significantly greater in 2-week old than 8-week old rats (267 + 47 percent versus 165 + 22 percent, respectively, p<0.05). O3 exposure also caused a significant increase in PGE2 in 2-week old but not adult rats. The results indicate that the ventilatory response to O3 is absent in 2-week old rats, and that lack of this response in conjunction with a greater specific ventilation leads to greater lung injury.
We also have made significant progress in establishing that TNFa acting through the p75 TNF receptor (type 2 TNF receptor, TNFR2) is important for O3 induced airway hyperresponsiveness. Wildtype mice (WT), TNF p55 or p75 receptor knockout mice (p55 TNFR -/- and p75 TNFR -/-) as well as double receptor knockout mice (p55/p75 TNFR -/-) were exposed to O3. Three hours after cessation of O3, airway responses to inhaled methacholine were determined by whole body plethysmography using changes in Penh as an index of airway narrowing. In Wildtype mice, O3 exposure (0.5 ppm, 3 hours) caused a significant increase in airway responsiveness as indicated by a 1.2 log leftward shift in the methacholine dose response curve. In contrast, in p55/p75 TNFR -/- mice, O3 caused only a 0.5 log shift in the dose response curve (p<0.05 compared to Wildtype). Similar results were obtained in p75 TNFR -/- mice. In contrast, O3 induced airway hyperresponsiveness was not different in WT and p55 TNFR -/- mice. These results indicate that TNFa acting through the p75 TNF receptor is necessary for O3-induced AHR. We do not yet know the precise role of TNF in these events.
During O3 exposure (1 ppm, 3 hours), VE decreased by 64 + 4 percent in Wildtype, but only 24 + 5 percent in p55/p75 TNFR -/- mice, indicating that despite their reduced O3-induced AHR, the TNFR deficient mice actually inhaled a greater dose of O3. Similar results were obtained in p75 -/- mice, whereas changes in VE induced by O3 were the same in Wildtype and p55 -/- mice. As in the experiments with rats described above, O3 exposure also caused a significant increase in EEP in these mice. The increase in EEP was significantly reduced in p55/p75 -/- mice indicating that not just the changes in ventilation, but also the changes in breathing pattern induced by O3 were mediated by TNF.
Polymorphonuclear leukocyte (PMN) numbers in bronchoalveolar lavage fluid recovered 21 hours after cessation of exposure to O3 (2 ppm, 3 hours), were significantly increased compared to after air exposure, but were not different in Wildtype and p55/p75 TNFR -/- mice. These results indicate that TNF contributes to the AHR but not the PMN emigration induced by acute O3 exposure.
Future Activities:
In the coming year, we will measure, in mice aged 2 weeks through 12 weeks, O3-induced changes in airway responsiveness to inhaled methacholine using both whole body plethysmography. In each mouse, we also will measure VE during the O3 exposures. We will then construct O3 dose (O3 conc. ? exposure time ? VE) response curves for O3-induced AHR. If our hypothesis that age-related differences in ventilation and in the effect of O3 on the ventilatory pattern contribute to age-related differences in AHR is correct, then we would expect to see no difference in O3 dose-AHR response curves between animals of different ages even though we see a leftward shift in the O3 concentration?AHR response curves in the younger animals. Similar experiments will be conducted using O3-induced changes in airway inflammation or injury, as assessed using bronchoalveolar lavage PMN and mediators, as outcome indicators.Journal Articles on this Report : 2 Displayed | Download in RIS Format
Other project views: | All 5 publications | 4 publications in selected types | All 4 journal articles |
---|
Type | Citation | ||
---|---|---|---|
|
Shore SA, Abraham JH, Schwartzman IN, Krishna Murthy GG, Laporte JD. Ventilatory responses to ozone are reduced in immature rats. Journal of Applied Physiology 2000;88(6):2023-2030. |
R827447 (1999) R827447 (2000) R827447 (Final) |
Exit Exit |
|
Shore SA, Schwartzman IN, Le Blanc B, Krishna Murthy GG, Doerschuk CM. Tumor necrosis factor receptor 2 contributes to ozone-induced airway hyperresponsiveness in mice. American Journal of Respiratory and Critical Care Medicine 2001;164(4):602-607. |
R827447 (1999) R827447 (Final) |
Exit Exit |
Supplemental Keywords:
ozone, exposure, health effects, age, biology, sensitive populations, mammalian., RFA, Health, Scientific Discipline, PHYSICAL ASPECTS, Air, air toxics, Health Risk Assessment, Risk Assessments, Susceptibility/Sensitive Population/Genetic Susceptibility, Biochemistry, Physical Processes, Children's Health, Molecular Biology/Genetics, tropospheric ozone, genetic susceptability, asthma, lung injury, ozone induced inflammation, sensitive populations, exposure and effects, stratospheric ozone, exposure, age-related differences, airway disease, dose response model, respiratory problems, ozone, air pollution, enzyme systems, ozone induced airway dysfunction, children, human exposure, assessment of exposure, children's vulnerablity, inhalation, age dependent response, biomedical research, acute exposure, environmental hazard exposuresRelevant Websites:
http://www.hsph.harvard.edu/facres/shr.htmlProgress and Final Reports:
Original AbstractThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.