Objectives We evaluated interactions between SERPINA1 PiMZ genotype, associated with intermediate α1-antitrysin deficiency, with outdoor particulate matter ≤10 µm (PM10), and occupational exposure to vapours, dusts, gases and fumes (VGDF), and their effects on annual change in lung function.
Methods Pre-bronchodilator spirometry was performed in 3739 adults of the Swiss Cohort Study on Air Pollution and Lung Disease in Adults (SAPALDIA) for whom SERPINA1 genotypes were available. At baseline in 1991, participants were aged 18–62 years; follow-up measurements were conducted from 2001 to 2003. In linear mixed regression models of annual change in lung function, multiplicative interactions were evaluated between PiMZ genotype (PiMM as reference) and change in PM10 (μg/m3), and VGDF exposure (high-level, low-level or no exposure as reference) during follow-up.
Results Annual declines in forced expiratory flow at 25–75% of forced vital capacity (FEF25–75%) (−82 mL/s, 95% CI −125 to −39) and forced expiratory volume in 1 s over forced vital capacity (FEV1/FVC) (−0.3%, 95% CI −0.6% to 0.0%) in association with VGDF exposure were observed only in PiMZ carriers (Pinteraction<0.0001 and Pinteraction=0.03, respectively). A three-way interaction between PiMZ genotype, smoking and VGDF exposure was identified such that VGDF-associated FEF25–75% decline was observed only in ever smoking PiMZ carriers (Pinteraction=0.01). No interactions were identified between PiMZ genotype and outdoor PM10.
Conclusions SERPINA1 PiMZ genotype, in combination with smoking, modified the association between occupational VGDF exposure and longitudinal change in lung function, suggesting that interactions between these factors are relevant for lung function decline. These novel findings warrant replication in larger studies.
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What this paper adds
There is evidence that α1-antitrypsin (AAT) deficiency, occupational exposure to vapours, gas, dusts and fumes (VGDF), ambient air pollution and smoking are all causative factors for adverse pulmonary health. The interactions between these factors are not well investigated.
In the present study, a three-way interaction was identified between SERPINA1 PiMZ genotype, which is associated with intermediate AAT deficiency, smoking and VGDF exposure. No interactions were present between PiMZ genotype and air pollution.
Genetic predisposition for intermediate AAT deficiency, in combination with smoking, may increase susceptibility to occupational exposure-related decline in lung function. These findings should be reassessed in larger studies.
Decline in lung function over time is a characteristic of aging, and accelerated decline is distinctive for obstructive lung diseases including asthma and chronic obstructive pulmonary disease (COPD).1 ,2 Epidemiological evidence from occupational and population-based studies indicates that occupational exposures to dusts, gases, and fumes are associated with additional decline in lung function and a causal risk factor of COPD.3 ,4 There is limited evidence to support that ambient air pollution is associated with lung function decline and COPD.4–6
The best known genetic risk factor for COPD is severe serum deficiency of α1-antitrypsin (AAT), an anti-inflammatory protease inhibitor (PI) of neutrophil elastase, a protease which breaks down elastin in the lung parenchyma.4 ,7 AAT is encoded by the SERPINA1 gene and allelic variants of SERPINA1 lead to inherited low AAT serum levels. Severe AAT deficiency is caused by rare SERPINA1 genotypes occurring at a frequency of <0.1% in the general population, like homozygosity for the PI deficient Z allele (PiZZ) or compound heterozygosity for PI deficient S and Z alleles (PiSZ).8 PiMZ and PiMS genotypes of SERPINA1 (M being the common non-deficient allele) lead to intermediate and mild AAT deficiency, respectively, and are more prevalent in the general populations representative of Middle European countries at 2% and 8% frequency, respectively.9 Meta-analyses offer mixed evidence to support an association between PiMZ genotype and risk of COPD, independent of smoking; there is no evidence of an association between PiMS genotype and risk of COPD.10 ,11
Earlier studies from the follow-up of the Swiss Cohort Study on Air Pollution and Lung Diseases in Adults (SAPALDIA 2) identified associations between improvements in the concentrations of outdoor particulate matter of 10 µm or less (PM10) and attenuation of lung function decline, with the greatest change in small airway function,5 and between high levels of occupational exposure to vapours, dusts, gases and fumes (VGDF) and incidence of COPD.12 A more recent study from the same survey showed associations between PiMZ genotype and age-related decline in forced expiratory flow between 25% and 75% of the forced vital capacity (ΔFEF25–75%) in participants with systemic low-grade inflammation, including persistent smokers.13 For this analysis, we evaluated whether the PiMZ genotype modified the associations between environmental and occupational exposures, including outdoor PM10, environmental tobacco smoke (ETS) exposure and occupational exposure to VGDF, and annual change in lung function in SAPALDIA, hypothesising that the associations between these exposures and lung function decline would be stronger in PiMZ carriers in comparison with PiMM carriers.
More detailed descriptions of the SAPALDIA study population and design are summarised in previous publications12–14 and in the online supplement.
SAPALDIA is a multicentre, population-based prospective cohort study consisting of a random sample of 9651 participants aged between 18 and 62 years that were recruited from eight regions in Switzerland, and were administered medical examinations, pre-bronchodilator spirometry testing and a detailed health questionnaire, at baseline in 1991.15 The follow-up survey (SAPALDIA 2) was conducted from 2001 to 2003, of which 8047 of the original study participants were present and of whom 6058 subjects provided blood samples and consented to DNA analysis. This analysis was restricted to 3739 participants who were either SERPINA1 PiMZ or PiMM carriers, completed spirometry testing at both surveys, reported information on important covariates, and for whom air pollution measurements (outdoor PM10) and occupational history during follow-up were available. Because we identified no presence of interaction between PiMS genotype and smoking in our previous analysis,13 and because there is no evidence of an association between PiMS genotype and risk of COPD, SAPALDIA participants who were PiMS carriers were not included in this analysis. See online supplementary figure E1 for additional details describing the selection of participants included in this analysis. The SAPALDIA cohort study complies with the Declaration of Helsinki. Written informed consent was obtained from participants in both surveys. The study was approved by the central ethics committee of the Swiss Academy of Medical Sciences and the respective Cantonal ethics committees of the eight study regions.
The spirometry protocol, which complied with American Thoracic Society (ATS) criteria,16 has been described in detail elsewhere.17–19 No bronchodilation was applied. The outcomes of interest for this study were annual changes in forced expiratory volume in 1 s (FEV1, mL), forced vital capacity (FVC, mL), the ratio of FEV1 over FVC (FEV1/FVC, %), and forced expiratory flow between 25% and 75% of FVC (FEF25–75, mL/s). Annual change in lung function was defined as the difference in each variable between the two examinations, divided by the follow-up time in years for the participant. See the online supplement for additional details describing methods for spirometry testing.
Occupational and environmental exposure assessment
ETS exposure at baseline and follow-up visits was assessed for different environments by the question ‘How many hours per day are you exposed to other people’s tobacco smoke: (i) at home; (ii) at the workplace?’ ETS exposure was categorised into three exposure groups: none, <3 h/day but not none, and ≥3 h/day.
Current job title reported by participants at the baseline survey (1991) and all job titles reported during the follow-up period were standardised according to the International Standard Classification of Occupations (ISCO-88) code's four-digit classification,20 and linked to the ALOHA GPJEM for COPD.21 ,22 Evaluation of occupational exposure in this analysis was restricted to exposure to high VGDF level during follow-up (low level or no exposure as reference) and continuous cumulative exposure (>0 years for exposed, 0 years for unexposed; unit, years) during follow-up. See the online supplement for additional descriptions of the ALOHA GPJEM and exposure variables.
Outdoor PM10 concentrations (μg/m3) outside of each subject's residence were estimated for the years 1990 and 2000 using a validated dispersion model (with different emissions inventories for both years) with a spatial resolution of 200×200 m. We estimated the change in PM10 exposure during follow-up (ΔPM10) for each participant, which was the difference between home outdoor mean PM10 level averaged over 1 year prior to the follow-up survey (in 2001–2003) and the corresponding mean PM10 level averaged over 1 year prior to the baseline survey (in 1990), divided by the time between baseline and follow-up examinations. As documented previously, overall exposure to home outdoor PM10 declined over the follow-up period (median, −5.3 μg/m3). Further details of the air pollution exposure assessment and modelling have been described previously.23 For this analysis, ΔPM10 was scaled per 10 μg increase over 10 years.
Genomic DNA was extracted from blood samples using the Puregene DNA Isolation Kit (Gentra Systems, USA). DNA samples of 6058 probands were shipped to the Center for Diagnosis of Inherited Alpha1-antitrypsin Deficiency in Pavia (Italy) and processed in a semi-automated medium throughput setup, assisted by liquid handling station (Freedom EVO75, Tecan Group, Switzerland). The PiS (rs17580) and PiZ (rs28929474) single nucleotide polymorphisms (SNPs) of SERPINA1 were genotyped using 5′-nuclease fluorescent real-time PCR (TaqMan Probes technology) on LightCycler480 (Roche). Genotype distributions for PiS and PiZ in this study sample were all in Hardy–Weinberg equilibrium.
We used linear mixed regression models of annual changes in FEV1, FVC, FEV1/FVC and FEF25–75%, with random intercept for study area, to evaluate multiplicative interactions between SERPINA1 PiMZ genotype (PiMM genotype as reference) and daily ETS, ΔPM10 and high-level VGDF exposures during follow-up. Interactions between PiMZ genotype and each exposure type were evaluated simultaneously in all models. We extracted the β coefficients to estimate the adjusted difference in annual change in lung function in association with each exposure in PiMZ and PiMM carriers from a single nested model with the multiplicative interaction terms. All models were adjusted for exposure and covariates ascertained at baseline including PM10 concentration, cumulative VGDF exposure, age, age squared, sex, height (cm), body mass index (BMI, kg/m2), early respiratory infection, parental asthma, and education; additional covariates included smoking status through follow-up (persistent smokers, former smokers, never smokers as reference), cumulative pack-years through follow-up, the interaction PiMZ genotype and smoking status, difference in BMI over follow-up period, and PiMZ genotype. Additionally, we tested for three-way interaction between smoking status through follow-up (ever smokers, never smokers), PiMZ genotype, and either ΔPM10 or high-level VGDF exposure. All analyses were done using SAS V.9.2. Two-sided p values <0.05 were interpreted as statistically significant for main and interaction effects.
A substantial number of participants with complete information at baseline were not included in the analysis due to non-participation, or incomplete genotyping, spirometry, or exposure and other covariate information at follow-up (49.5%). The non-participants were more likely to be older, female, heavier, smokers, have lower lung function, report more daily exposure to ETS, or have higher exposure to outdoor PM10 at the baseline survey (see online supplementary table E1). No significant difference was observed in the distribution of occupational VGDF exposure between the participants and non-participants. To account for the potential bias arising from differences between participants and non-participants, we weighted the linear mixed effect regression models by the inverse probability for inclusion in the analysis. We also evaluated whether baseline lung function attenuated the interactions between PiMZ genotype and each exposure, with additional adjustment for baseline lung function.
Characteristics of the study population stratified by SERPINA1 genotype are summarised in table 1. Similar distributions between genotypes were observed for BMI and height, but PiMZ carriers were slightly younger, and had higher lung function at baseline on average compared with PiMM carriers. The mean annual declines in FEV1 and FEV1/FVC over the follow-up period were slightly larger in PiMZ carriers compared with PiMM carriers, with a larger difference observed for FEF25–75% (−88 mL/s vs −71 mL/s, respectively). The prevalence of ever smokers and mean cumulative pack-years was lower in PiMZ carriers. Compared with PiMM carriers, fewer PiMZ carriers reported being exposed to ETS ≥3 h/day. Over the follow-up period, PiMZ carriers had a slightly larger mean reduction in outdoor PM10 on average compared with PiMM carriers. The prevalence of high-level VGDF exposure at baseline and during follow-up was also moderately lower in PiMZ carriers.
The adjusted differences in annual changes in lung function in association with ΔPM10 and high-level VGDF exposures during follow-up estimated in PiMZ and PiMM carriers are presented in table 2. A statistically significant interaction (p<0.0001) was observed between PiMZ genotype and high-level VGDF on annual change in FEF25–75%. High-level VGDF exposure was associated with accelerated annual decline in FEF25–75% (−82 mL/s, 95% CI −125 to −39) in PiMZ carriers, while a positive association was observed in PiMM carriers. A similar interaction of statistical significance (p=0.03) was observed between PiMZ genotype and high-level VGDF exposure on annual change in FEV1/FVC. Overall, larger annual declines in lung function in association with ΔPM10 were observed in PiMZ carriers than in PiMM carriers, but no statistically significant interactions were present. No interactions were present between PiMZ genotype and daily ETS exposure (data not shown).
After adjustment for baseline lung function (see online supplementary table E2), the estimated effects of ΔPM10 and high-level VGDF exposure on annual decline in lung function in PiMZ carriers were smaller compared to those presented in table 2, but the interaction between PiMZ genotype and high-level VGDF exposure on annual change in FEF25–75% remained statistically significant. The estimated associations presented in table 2 were similar to those after weighting the models according to the inverse probability of inclusion in the analysis (see online supplementary table E3); the interaction between PiMZ genotype and high-level VGDF exposure on annual change in FEV1/FVC and FEF25–75% remained statistically significant. When occupational VGDF exposure was characterised as continuous cumulative exposure (see online supplementary table E4), the interactions between PiMZ genotype and cumulative VGDF exposure during follow-up were similar to those presented for high-level VGDF exposure presented in table 2; statistically significant excess annual declines in FEV1/FVC and FEF25–75% per 10 years of cumulative VGDF exposure were observed only in PiMZ carriers.
Table 3 summarises the adjusted differences in annual change in lung function for ΔPM10 and high-level VGDF exposure by PiMZ genotype and smoking status, as estimated from the nested model with a three-way interaction between PiMZ genotype, smoking, and environmental (ΔPM10) or occupational (high-level VGDF) exposure, respectively. Overall, the largest annual declines in FEF25–75% (−109 mL/s; 95% CI −159 to −59) and FEV1/FVC (−0.4%; 95% CI −0.8% to −0.0%) associated with high-level VGDF exposure during follow-up were observed for ever smoking PiMZ carriers. A three-way interaction between ever smoking status, PiMZ genotype and high-level VGDF exposure on annual change in FEF25–75% was statistically significant (p=0.01).
In this prospective cohort study of Swiss working adults, we observed the SERPINA1 PiMZ genotype, in combination with smoking, to modify the association between occupational exposure to VGDF during follow-up and annual change in lung function, particularly for FEF25–75% and FEV1/FVC such that the excess annual decline in FEF25–75% and FEV1/FVC was observed in ever smoking PiMZ carriers. To a lesser extent, larger annual declines in lung function in association with ΔPM10 were observed in PiMZ carriers than in PiMM carriers, but no statistically significant interactions were present. We did not detect any interaction between PiMZ genotype and daily ETS exposure.
Earlier cross-sectional population-based studies of homozygous PiZ individuals have shown that lower FEV1 was observed in participants who reported occupational exposures compared to unexposed participants.24 ,25 Similarly, a prospective study of exposed iron-ore miners demonstrated that a larger decline in FEV1 was observed in workers with intermediate AAT deficiency compared with workers with normal AAT phenotype.26 These studies could not formally assess effect modification as they included only AAT deficient or only exposed individuals. A cross-sectional study of young farming apprentices and unexposed conscripts observed an interaction between farming occupation and PiMZ phenotype on bronchial hyperresponsiveness; no effect modification by PiMZ phenotype was observed between farming occupation and lung function.27 A recent study of New York City World Trade Center (WTC) firefighters observed that workers with moderate and mild AAT deficiency experienced larger declines in FEV1 in comparison with participants with no AAT deficiency during the first 4 years after September 11th.28 In the study of WTC firefighters, AAT deficiency also did not enhance the rate of FEV1 decline among the study participants pre-9/11, suggesting a novel interaction between AAT deficiency and exposures resulting from 9/11 on lung function decline. Our study is the first to identify interaction between SERPINA1 genotype for predisposition to intermediate AAT deficiency and chronic occupational exposure to high-levels of VGDF in a general population prospective cohort-based setting over a long duration of follow-up.
It is of interest that PiMZ carriers were less likely to be exposed to high-level VGDF exposures at baseline and during the follow-up period compared with PiMM carriers. The discrepancy in the VGDF exposure distribution by SERPINA1 genotype may be suggestive of a healthy selection effect associated with occupational inhalant exposures. It may be hypothesised that PiMZ carriers may select themselves away from occupational respirable exposures that may be irritable. They may not seek employment in occupations with high exposure to VGDF due to health conditions attributable to AAT deficiency, or they may leave occupations with high exposure to VGDF due to health aggravation or illness prior to the baseline survey, all of which may reduce power to observe an association between occupational exposure and lung function decline in PiMZ carriers. Because we did not have SERPINA1 genotype information at the time of the baseline survey, we cannot formally evaluate this question, but the influence of genetic susceptibility on healthy worker selection and survival effects merits further research.
There is mixed evidence to support that PiMZ is associated with accelerated lung function decline and increased risk of COPD,11 and our previous analysis in SAPALDIA did not identify any main associations between PiMZ genotype and lung function decline.13 However, the current findings suggest that PiMZ genotype, only in combination with smoking, may enhance susceptibility for occupational exposure associated lung function decline. The biological mechanism(s) explaining this susceptibility is not clear. Previous studies in other populations have also observed a combined effect between occupational exposure to VGDF and smoking on risk of COPD.29–31 Thus the combined interactive effects between PiMZ genotype, smoking and occupational VGDF exposure may be a reflection of smokers with intermediate AAT deficiency exceeding their ability to cope with additional occupational VGDF exposure, or due to poor mucus clearance among PiMZ smokers leading to prolonged inflammatory response after occupational VGDF exposure. Alternatively, it may be that PiMZ individuals may be more susceptible to lung function decline attributable to the combination of VGDF and cigarette smoke due to a diminished protection against tissue damage from elastase in AAT deficiency.32
The interaction between causative factors for adverse pulmonary health including AAT deficiency, smoking, and occupational VGDF exposure is also of special public health relevance, particularly because COPD and its related conditions are associated with high morbidity and mortality, and because the prevalence of genetic predisposition to intermediate AAT deficiency is estimated to be around 30 million worldwide.33 The role of genetic screening in the workplace is also raised as a topic of discussion34–36 in light of the numerous studies that have documented gene–occupation interactions. However, no genetic test related to an occupational disease has been validated or accepted for use, except the use of genetic biomarkers to measure the dose of a genotoxic exposure34; and there are many scientific, ethical, legal and social issues to consider for the use of genetic screening for susceptibility to workplace exposures. In this context, it serves as an important reminder to occupational health practitioners and researchers that occupational exposures are the primary cause of occupational disease and will remain the responsibility of the employer to control.35
This study has numerous strengths, including selection of SNPs with demonstrated functionality,9 prospective study design, detailed data on individual smoking and ETS and PM10 exposure, semi-quantitative estimates of occupational exposures to VGDF using a GPJEM, extensive control for confounding, and high quality of longitudinal lung function data. However, there are a number of limitations to be considered. We observed that effect modification by the PiMZ genotype on the association between VGDF exposure and lung function decline was strongest for FEF25–75%, consistent with previous studies of the SAPALDIA cohort that have also identified strong main and interacting associations between air pollution and FEF25–75%,5 ,37 and between PiMZ genotype and FEF25–75% in population subgroups indicative of low-grade systemic inflammation.13 FEF25–75% is also used as an indicator of small airways obstruction and bronchial reactivity and correlates highly with percentage predicted FEV1/FVC at levels of at least moderate airflow obstruction.38 The primary limitation of FEF25–75% is that its measurement precision within subjects is lower than of FEV1.39 In this study sample, the correlation between baseline and follow-up measurements of FEV1, FEF25–75% and FEV1/FVC were 0.92, 0.81 and 0.72, respectively. While it is unknown why an interaction was not observed between PiMZ and exposures on annual change in FEV1, a spirometry metric which has less error, we hypothesise, given the previous findings in the SAPALDIA study, that the interactive effects observed for FEF25–75% are of relevance in this study population.
The relatively high degree of non-participation in our analysis may have also biased our observed associations. However, the use of inverse probability weighting to account for this potential selection bias did not result in considerable differences in the estimated associations, which suggests that any bias from non-participation is likely to be minimal. Finally, VGDF-associated lung function decline was limited to ever smoking PiMZ carriers, which may be explained by insufficient power to identify an interaction between PiMZ genotype and occupational VGDF exposure in non-smokers; seven of the nine participants with high-level VGDF exposure and PiMZ genotype were ever smokers. An alternative explanation is that smoking may be a prerequisite for the interaction between PiMZ genotype and VGDF exposure to occur as it increases inflammation and therefore elastase activity in the lung.
The present findings suggest that interaction between causative factors plays an important role in the nature of lung function decline. Genetic predisposition for intermediate AAT deficiency, in combination with smoking, may increase susceptibility to occupational exposure-related decline in lung function. The interaction between PiMZ genotype and outdoor PM10 needs to be reassessed in larger studies.
We would like to thank the whole SAPALDIA team for their contribution to the study. Additionally, the study could not have been done without the help of the study participants, technical and administrative support and the medical teams and field workers at the local study sites. See the online supplement for a detailed description of the SAPALDIA team.
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AJM and GAT contributed equally.
Contributors AM wrote the manuscript; AM and GAT did the statistical analysis; NK, HP, TR, EWR, CS, ML and NPH were involved in the study design; GAT, MI and IF did the genotyping; DK, HK, HP and RV were involved in exposure data collection; MI, NK, HK, DM, CS, JS, RV, ML and NPH helped with discussion and interpretation of the results. All authors reviewed and approved the manuscript.
Funding This study was supported by Swiss Accident Insurance Fund (SUVA), Swiss National Science Foundation (grants no 33CS30_134276/1, 33CSCO-108796, 3247BO-104283, 3247BO-104288, 3247BO-104284, 3247-065896, 3100-059302, 3200-052720, 3200-042532, 4026-028099, 3233-054996, PDFMP3-123171), the Federal Office for Forest, Environment and Landscape, the Federal Office of Public Health, the Federal Office of Roads and Transport, the canton's government of Aargau, Basel-Stadt, Basel-Land, Geneva, Luzern, Ticino, Valais, Zurich, the Swiss Lung League, the canton's Lung League of Basel Stadt/Basel Landschaft, Geneva, Ticino, Valais and Zurich, Freiwillige Akademische Gesellschaft, UBS Wealth Foundation, Talecris Biotherapeutics GmbH and Abbott Diagnostics. The Center for Diagnosis of Inherited Alpha1-antitrypsin Deficiency in Pavia is supported by grants from Talecris Biotherapeutics GmbH, Kedrion S.p.A., IRCCS (Istituto di ricovero e cura a carattere scientifico) Foundation San Matteo Hospital, and Cariplo Foundation 2006 projects.
Competing interests DM is an employee of a Swiss workers compensation board (SUVA, Occupational Medicine Department), TR has participated in advisory boards sponsored by GlaxoSmithKline, Takeda, Grifols (formerly Talecris) and InterMune. ML received an unrestricted research grant from Talecris/Grifols, consultancy fees from Grifols and gave paid lectures for Kedrion. NPH has received an unrestricted research grant from Talecris. The grant money was applied to covering part of the salary costs for GAT.
Patient consent Obtained.
Ethics approval Central ethics committee of the Swiss Academy of Medical Sciences and the respective cantonal ethics committees of the eight study regions.
Provenance and peer review Not commissioned; externally peer reviewed.