Imboden et al. Current and past smoking habits, exposure to ETS, and occupational exposure to dust and fumes were assessed with the same questions at both surveys. Participants who reported smok- ing < 20 packs of cigarettes and using < 360 g of tobacco in their lifetime at both time points were defined as never-smokers. Cumulative cigarette exposure of participants was assessed by pack-years smoked before the first examina- tion and pack-years smoked during follow-up. Participants were asked not to smoke in the hour before the examination, and smoking was validated by measuring the carbon mon- oxide concentration in exhaled breath using an EC 50 Micro-Smokerlizer (Bedfont Scientific, Rochester, UK). Atopic status was assessed at baseline using skin prick tests (Phazet; Pharmacia, Uppsala, Sweden) for eight com- mon inhalant allergens: dog epithelium; cat fur; pollen of timothy grass, Parietaria, and birch; the house dust mite Dermatophagoides ptero- nyssinus; and the molds Alternaria tenuis and Cladosporium herbarum (Martin et al. 1997; Wuthrich et al. 1996). Atopy was defined by an adjusted mean wheal diameter ≥ 3 mm to at least one allergen. Statistical analysis. We inferred CCND1 haplotypes from unphased genotype data using PHASE 2.1 algorithm software [see Supplemental Material, Table 1 (doi:10.1289/ ehp.0800430.S1)] (Stephens et al. 2001). Hardy-Weinberg equilibrium was tested using STATA gtab command for global κ-statistic testing (STATA version 10; StataCorp, College Station, TX, USA). All four SNPs were in Hardy-Weinberg equilibrium. We obtained Lewtonin’s linkage disequilibrium (LD) metric D´using STATA command pwld for pairwise LD. Descriptive analyses of the lung function parameters, PM10, smoking, socioeconomic variables, and other relevant covariates have been described previously and in detail in different SAPALDIA cohort study reports (Ackermann-Liebrich et al. 2005; Downs et al. 2007; Imboden et al. 2007). We compared baseline characteristics of the cohort participants included in this study (n = 4,326) with subjects participating only at baseline as well as with cohort participants excluded from this analysis because information on genotype or covariate data was missing. Results are presented as the estimated effect of a 10-µg/m3 decrease in the annual average PM10 exposure over the follow-up period (ΔPM10) on the average attenuation of the annual decline in FEF25–75. The association between ΔPM10 and the average annual rate of lung function decline had been previously assessed using mixed linear model analysis, and selection of relevant covariates was based on this previ- ous investigation (Downs et al. 2007): age at baseline (SAPALDIA1), age2, sex, height, parental smoking during childhood reported 1422 at baseline, sine and cosine function of day of examination to control for seasonal effects, level of education at baseline, change in level of education, Swiss nationality, self-reported occupational exposure to dust and occupa- tional exposure to fumes at SAPALDIA1 and SAPALDIA2 (yes/no), smoking status at follow-up (never, former, or current), pack-years up to SAPALDIA1, pack-years between SAPALDIA1 and -2, cigarettes per day at SAPALDIA1 and -2, atopy, body mass index (BMI) at SAPALDIA1, change in BMI, interaction between the two BMI vari- ables, and baseline PM10 exposure. Random effects were included to adjust for clustering of residuals within area and were assumed to be independent between the areas and to have an exchangeable correlation structure. To estimate main effects of gene variants of CCND1, p21, and p53 and rate of lung func- tion decline, we used the same mixed model with random area effects additionally adjusted for ΔPM10. To estimate modification of the ΔPM10 effect on average lung function decline by genotype, we introduced interaction terms between ΔPM10 and genotypes into the above-described covariate-adjusted mixed linear models independently for each SNP. Also, for each SNP we evaluated three differ- ent genetic models (codominant, dominant, and recessive) because previous cancer asso- ciation studies were not conclusive about the underlying genetic model of the SNP effects. The codominant model assumes that the gene effect depends on the number of alleles in a dose-dependent manner, the dominant model assumes that the gene effect depends on the presence of at least one of two high-risk alleles, and the recessive model assumes that the gene effect depends on the presence of both high- risk alleles. The three genetic models required different coding of genotypes. For each SNP we present the genetic model with the smallest interaction term p-value. We obtained effect estimates for the association between ΔPM10 and average annual lung function decline in genotype subgroups by creating genotype- specific PM10 exposures variables that we introduced into separate covariate-adjusted mixed linear models. Analyses were conducted using SAS release 9.1 (SAS Institute Inc., Cary, NC, USA) and STATA version 10. p-Values < 0.05 were interpreted as statistically signifi- cant for main and interaction effects. p-Values presented as main p-values in the tables are not corrected for multiple testing. Nevertheless, Bonferroni-corrected significance level (α = 0.05 divided by the number of tests) is indi- cated in tables and figures as appropriate. Results Population characterization. Table 1 describes the general characteristics of the study popula- tion (n = 4,326), including relevant predictors of lung function. This is a population con- sisting mostly of Caucasians. At follow-up participants were more likely to be women and less likely to be smokers than were non- participants. Participants tended to gain weight, give up smoking, and reduce expo- sure to dust or fumes at work or to ETS [for details, see Supplemental Material, Table 2 (doi:10.1289/ehp.0800430.S1)]. In 2002, 87% of the participants were liv- ing in the same area as in 1991, and 54% had the same address. In general, individual home outdoor concentrations of PM10 declined dur- ing the follow-up period (Table 1), as previ- ously described in detail (Downs et al. 2007; Liu et al. 2007). The median decline between examinations for subjects included in this analysis was 5.8 µg/m3 (interquartile range, 4.2–7.3 µg/m3). Mean decline was greatest for participants living in urban areas and low- est in alpine areas. Mean lung function for the cohort was within the range of predicted values for the general population and declined during the follow-up period (Table 1). The mean ± SD annual change in mid FEF25–75 during the 11-year follow-up period was –74.1 ± 70.3 mL/year in men and –68.9 ± 58.9 mL/ year in women included in this analysis. The main effects of the investigated genetic variants on change in lung function are described in more detail in Supplemental Material, Table 3 (doi:10.1289/ehp.0800430. S1). Briefly, we observed no statistically signi- ficant associations of the polymorphisms either with change in FEF25–75 or with FEV1 or FVC. Modification of the ΔPM10 effect on aver- age decline in lung function by genotype. As previously reported (Downs et al. 2007), a 10-µg/m3 decline in average annual home outdoor PM10 concentration over an 11-year period (ΔPM10) reduced the annual rate of decline in FEF25–75 on average by 11.2 mL/year or 16%. Here we report statistically significant modifications of this association of ΔPM10 with FEF25–75 decline by three of the four genetic variants investigated: CCND1 P242P [p-value for interaction (pint) = 0.017], CCND1 rs667515 (pint = 0.006), and p53 R72P (pint = 0.016). Figure 1A and Table 2 present results for the association of ΔPM10 with the aver- age annual change in FEF25–75 within geno- type and diplotype strata for the entire study population; for data for never-smokers, see Figure 1B and Supplemental Material, Table 4 (doi:10.1289/ehp.0800430.S1). Equivalent to the previously reported main effect of ΔPM10 (Downs et al. 2007), the genotype- specific results we observed in the entire study population were comparable to those in never- smokers. The A allele for CCND1 P242P reduced the attenuating effect estimate of ΔPM10 on FEF25–75 decline in a codominant ovlume 117 | number 9 | September 2009 • Environmental Health Perspectives