Cantone et al. (version 9.1.3; SAS Institute Inc., Cary, NC, coefficient (r) = 0.7–1] for aluminum with and histone modifications in age-adjusted USA) and R (R Foundation for Statistical manganese, lead, and PM10; for manganese regression models. Computing, Vienna, Austria). with lead and PM10; for nickel with arsenic Association of levels of exposure to inhal- and iron; and between arsenic and iron, PM10 able metals with histone modifications. We Results and lead, and PM1 and PM10. Correlations evaluated whether levels of H3K4me2 and Subjects’ characteristics and exposure levels. The were moderate (r = 0.4–0.7) for PM1 with alu- H3K9ac were associated with the levels of per- mean age of the study subjects was 44.00 years minum, manganese, and lead, between nickel sonal exposure to metals in inhalable PM, as (range, 27–55 years). Twenty-five subjects and aluminum, and between iron and zinc. well as to PM mass, in both simple regression (40%) were current smokers, who reported a Except for PM1 and arsenic (r = –0.20), all models and multivariable models adjusted median number of 15 cigarettes smoked every other correlations were positive with r < 0.40. for age, BMI, education, pack-years, and per- day (range, 0–30 cigarettes/day). The BMI Associations of histone modifications with cent granulocytes. Results from unadjusted of the study participants showed a median of subjects’ characteristics and years of employ- and adjusted models showed similar results 26 kg/m2 (range, 20–33 kg/m2). Table 1 shows ment. The range of values of the histone mod- (Table 4). In adjusted models, H3K4me2 the average levels of inhalable air metal con- ifications was 0.26–1.20 OD for H3K4me2 increased in association with nickel, arsenic, centrations and PM mass estimated. For both (median, 0.99 OD) and –0.50 to 1.06 OD and iron. H3K4me2 was not associated with metal levels and PM mass, the study subjects for H3K9ac (median, 0.37 OD). aluminum, manganese, zinc, lead, PM10, or showed wide ranges of exposures. The subject H3K4me2 was moderately but signifi- PM1 exposure levels in unadjusted or adjusted with the maximum individual exposure level cantly correlated with H3K9ac (Pearson’s r regression models (Table 4). H3K9ac was was at least 17 times more exposed than the = 0.45, p < 0.001). H3K4me2 and H3K9ac positively but not significantly associated with subject with the minimum exposure level (e.g., were not associated with age, BMI, smok- nickel and iron levels. H3K9ac was not asso- for PM10: maximum, 1220.17 μg/m 3 ; mini- ing, number of cigarettes/days, smoking dura- ciated with the levels of the remaining metals, mum, 73.72 μg/m 3). For some of the expo- tion, pack-years, area of residence (city center, PM10 or PM1 (Table 4). sures (aluminum, manganese, zinc, lead), the suburbs, rural), self-reported traffic intensity Association of cumulative exposure to maximum individual exposure was > 200 times near home, or percent granulocytes (Table 3). inhalable metals with histone modifications. higher than the minimum individual exposure H3K9ac was positively associated with edu- Table 5 shows the associations of H3K4me2 (Table 1). cation (p = 0.04; Table 3). Both H3K4me2 and H3K9ac with cumulative exposures, esti- The correlation matrix between expo- and H3K9ac increased in association with mated as the product of level of exposure to sure levels showed correlations of different the years of employment of the study subjects PM metal components or total PM mass and strengths between the exposures (Table 2). We in the steel plant (Table 3). We confirmed years of employment. Again, unadjusted and found high correlations [Pearson’s correlation the association between years of employment adjusted models showed similar results. In adjusted models, H3K4me2 increased in asso- Table 1. Levels of personal exposure to metal components of PM and total PM mass. ciation with cumulative exposure to nickel Percentile and arsenic. H3K9ac showed significant asso- Exposurea (μg/m3) Mean ± SD Minimum 25th 50th 75th Maximum n ciations with cumulative exposure to nickel Aluminum 63 8.50 ± 18.07 0.40 1.48 2.05 7.41 84.07 and arsenic. Cumulative exposure to iron was Manganese 63 11.26 ± 30.41 0.11 1.20 4.63 10.77 174.79 positively but not significantly associated with Nickel 63 0.30 ± 0.18 0.02 0.23 0.25 0.46 0.72 both H3K4me2 and H3K9ac. Zinc 63 18.85 ± 26.37 0.26 1.47 8.45 32.28 129.06 Arsenic 63 0.10 ± 0.1 0.01 0.02 0.07 0.17 0.31 Discussion Lead 63 7.53 ± 17.46 0.13 0.63 2.87 9.52 99.90 The present study, based on a healthy worker Iron 63 32.02 ± 22.08 0.96 18.00 25.64 48.69 88.43 population from a steel plant near Brescia, Italy, PM10 63 233.42 ± 214.56 73.72 152.23 179.45 222.86 1220.17 showed that exposure to some metal compo- PM1 63 8.48 ± 6.18 1.71 3.51 9.01 11.35 30.49 nents of PM was associated with increased acti- aMetal components were measured on the PM10 fraction of PM mass. Coarse PM levels were calculated as the differ- ence between PM10 and PM1. vating histone modifications measured in blood leukocyte samples. In particular, we found that Table 2. Matrix of correlations (r)a among individual exposures (PM metal components and PM mass both H3K4me2 and H3K9ac were higher in measures). individuals with more years of employment Exposure Aluminum Manganese Nickel Zinc Arsenic Lead Iron PM10 PM1 in the plant and higher estimated cumulative Aluminum 1 exposures to arsenic and nickel. Manganese 0.75 1 To the best of our knowledge, this is the < 0.0001 first study showing associations between metal Nickel 0.46 0.39 1 exposures, such as nickel, arsenic, and iron, 0.0001 0.0017 and histone modifications in human subjects. Zinc 0.18 0.21 0.32 1 Our study was based on measures of histone 0.1625 0.1034 0.0105 Arsenic 0.15 0.31 0.84 0.25 1 modifications in blood leukocyte samples from 0.2378 0.0143 0.0000 0.0483 healthy subjects, thus suggesting that exposure- Lead 0.75 0.99 0.38 0.35 0.28 1 related alterations of histone modifications may < 0.0001 < 0.0001 0.0018 0.0047 0.0265 occur in normal tissues and possibly anticipate Iron 0.15 0.28 0.76 0.48 0.70 0.32 1 the onset of disease. Estimated effects of metal 0.2451 0.0261 < 0.0001 0.0001 < 0.0001 0.0113 exposures on H3K4me2 in blood leukocytes PM10 0.81 0.82 0.34 0.34 0.04 0.85 0.27 1 were consistent with previous in vitro toxi- < 0.0001 < 0.0001 0.0056 0.0070 0.7446 < 0.0001 0.0294 PM1 0.63 0.60 0.22 0.28 –0.20 0.64 0.26 0.90 1 cology studies that showed that carcinogenic < 0.0001 < 0.0001 0.0897 0.0277 0.1195 < 0.0001 0.0408 < 0.0001 metals increased H3K4me2 in A549 human lung carcinoma cells (Sun et al. 2009; Zhou X aPearson’s product-moment correlation coefficient and corresponding p-values. 966 119 | number 7 | July 2011 • Environmental Health Perspectives ovlume