Multicenter Ozone Study in oldEr Subjects (MOSES): Part 2. Effects of Personal and Ambient Concentrations of Ozone and Other Pollutants on Cardiovascular and Pulmonary Function.

Introduction: The Multicenter Ozone Study of oldEr Subjects (MOSES) was a multi-center study evaluating whether short-term controlled exposure of older, healthy individuals to low levels of ozone (O₃) induced acute changes in cardiovascular biomarkers. In MOSES Part 1 (MOSES 1), controlled O₃ exposure caused concentration-related reductions in lung function with evidence of airway inflammation and injury, but without convincing evidence of effects on cardiovascular function. However, subjects' prior exposures to indoor and outdoor air pollution in the few hours and days before each MOSES controlled O₃ exposure may have independently affected the study biomarkers and/or modified biomarker responses to the MOSES controlled O₃ exposures.

Methods: MOSES 1 was conducted at three clinical centers (University of California San Francisco, University of North Carolina, and University of Rochester Medical Center) and included healthy volunteers 55 to 70 years of age. Consented participants who successfully completed the screening and training sessions were enrolled in the study. All three clinical centers adhered to common standard operating procedures and used common tracking and data forms. Each subject was scheduled to participate in a total of 11 visits: screening visit, training visit, and three sets of exposure visits consisting of the pre-exposure day, the exposure day, and the post-exposure day. After completing the pre-exposure day, subjects spent the night in a nearby hotel. On exposure days, the subjects were exposed for 3 hours in random order to 0 ppb O₃ (clean air), 70 ppb O₃, and 120 ppm O₃. During the exposure period the subjects alternated between 15 minutes of moderate exercise and 15 minutes of rest. A suite of cardiovascular and pulmonary endpoints was measured on the day before, the day of, and up to 22 hours after each exposure.

In MOSES Part 2 (MOSES 2), we used a longitudinal panel study design, cardiopulmonary biomarker data from MOSES 1, passive cumulative personal exposure samples (PES) of O₃ and nitrogen dioxide (NO₂) in the 72 hours before the pre-exposure visit, and hourly ambient air pollution and weather measurements in the 96 hours before the pre-exposure visit. We used mixed-effects linear regression and evaluated whether PES O₃ and NO₂ and these ambient pollutant concentrations in the 96 hours before the pre-exposure visit confounded the MOSES 1 controlled O₃ exposure effects on the pre- to post-exposure biomarker changes (Aim 1), whether they modified these pre- to post-exposure biomarker responses to the controlled O₃ exposures (Aim 2), whether they were associated with changes in biomarkers measured at the pre-exposure visit or morning of the exposure session (Aim 3), and whether they were associated with differences in the pre- to post-exposure biomarker changes independently of the controlled O₃ exposures (Aim 4).

Results: Ambient pollutant concentrations at each site were low and were regularly below the National Ambient Air Quality Standard levels. In Aim 1, the controlled O₃ exposure effects on the pre- to post-exposure biomarker differences were little changed when PES or ambient pollutant concentrations in the previous 96 hours were included in the model, suggesting these were not confounders of the controlled O₃ exposure/biomarker difference associations. In Aim 2, effects of MOSES controlled O₃ exposures on forced expiratory volume in 1 second (FEV₁) and forced vital capacity (FVC) were modified by ambient NO₂ and carbon monoxide (CO), and PES NO₂, with reductions in FEV₁ and FVC observed only when these concentrations were "Medium" or "High" in the 72 hours before the pre-exposure visit. There was no such effect modification of the effect of controlled O₃ exposure on any other cardiopulmonary biomarker.

As hypothesized for Aim 3, increased ambient O₃ concentrations were associated with decreased pre-exposure heart rate variability (HRV). For example, high frequency (HF) HRV decreased in association with increased ambient O₃ concentrations in the 96 hours before the pre-exposure visit (-0.460 ln[ms²]; 95% CI, -0.743 to -0.177 for each 10.35-ppb increase in O₃; P = 0.002). However, in Aim 4 these increases in ambient O₃ were also associated with increases in HF and low frequency (LF) HRV from pre- to post-exposure, likely reflecting a "recovery" of HRV during the MOSES O₃ exposure sessions. Similar patterns across Aims 3 and 4 were observed for LF (the other primary HRV marker), and standard deviation of normal-to-normal sinus beat intervals (SDNN) and root mean square of successive differences in normal-to-normal sinus beat intervals (RMSSD) (secondary HRV markers).

Similar Aim 3 and Aim 4 patterns were observed for FEV₁ and FVC in association with increases in ambient PM with an aerodynamic diameter ≤ 2.5 μm (PM_2.5), CO, and NO₂ in the 96 hours before the pre-exposure visit. For Aim 3, small decreases in pre-exposure FEV₁ were significantly associated with interquartile range (IQR) increases in PM_2.5 concentrations in the 1 hour before the pre-exposure visit (-0.022 L; 95% CI, -0.037 to -0.006; P = 0.007), CO in the 3 hours before the pre-exposure visit (-0.046 L; 95% CI, -0.076 to -0.016; P = 0.003), and NO₂ in the 72 hours before the pre-exposure visit (-0.030 L; 95% CI, -0.052 to -0.008; P = 0.007). However, FEV₁ was not associated with ambient O₃ or sulfur dioxide (SO₂), or PES O₃ or NO₂ (Aim 3). For Aim 4, increased FEV₁ across the exposure session (post-exposure minus pre-exposure) was marginally significantly associated with each 4.1-ppb increase in PES O₃ concentration (0.010 L; 95% CI, 0.004 to 0.026; P = 0.010), as well as ambient PM_2.5 and CO at all lag times. FVC showed similar associations, with patterns of decreased pre-exposure FVC associated with increased PM_2.5, CO, and NO₂ at most lag times, and increased FVC across the exposure session also associated with increased concentrations of the same pollutants, reflecting a similar recovery. However, increased pollutant concentrations were not associated with adverse changes in pre-exposure levels or pre- to post-exposure changes in biomarkers of cardiac repolarization, ST segment, vascular function, nitrotyrosine as a measure of oxidative stress, prothrombotic state, systemic inflammation, lung injury, or sputum polymorphonuclear leukocyte (PMN) percentage as a measure of airway inflammation.

Conclusions: Our previous MOSES 1 findings of controlled O₃ exposure effects on pulmonary function, but not on any cardiovascular biomarker, were not confounded by ambient or personal O₃ or other pollutant exposures in the 96 and 72 hours before the pre-exposure visit. Further, these MOSES 1 O₃ effects were generally not modified, blunted, or lessened by these same ambient and personal pollutant exposures. However, the reductions in markers of pulmonary function by the MOSES 1 controlled O₃ exposure were modified by ambient NO₂ and CO, and PES NO₂, with reductions observed only when these pollutant concentrations were elevated in the few hours and days before the pre-exposure visit. Increased ambient O₃ concentrations were associated with reduced HRV, with "recovery" during exposure visits. Increased ambient PM_2.5, NO₂, and CO were associated with reduced pulmonary function, independent of the MOSES-controlled O₃ exposures. Increased pollutant concentrations were not associated with pre-exposure or pre- to post-exposure changes in other cardiopulmonary biomarkers. Future controlled exposure studies should consider the effect of ambient pollutants on pre-exposure biomarker levels and whether ambient pollutants modify any health response to a controlled pollutant exposure.