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. 2019 Apr;127(4):047009.
doi: 10.1289/EHP3398.

Associations between Maternal Tobacco Smoke Exposure and the Cord Blood [Formula: see text] DNA Methylome

Caitlin G Howe  1 , Meng Zhou  2 , Xuting Wang  3 , Gary S Pittman  3 , Isabel J Thompson  3 , Michelle R Campbell  3 , Theresa M Bastain  1 , Brendan H Grubbs  4 , Muhammad T Salam  1   5 , Cathrine Hoyo  6 , Douglas A Bell  3 , Andrew D Smith  2 , Carrie V Breton  1
Affiliations

Associations between Maternal Tobacco Smoke Exposure and the Cord Blood [Formula: see text] DNA Methylome

Caitlin G Howe et al. Environ Health Perspect. 2019 Apr.

Abstract

Background: Maternal tobacco smoke exposure has been associated with altered DNA methylation. However, previous studies largely used methylation arrays, which cover a small fraction of CpGs, and focused on whole cord blood.

Objectives: The current study examined the impact of in utero exposure to maternal tobacco smoke on the cord blood [Formula: see text] DNA methylome.

Methods: The methylomes of 20 Hispanic white newborns ([Formula: see text] exposed to any maternal tobacco smoke in pregnancy; [Formula: see text] unexposed) from the Maternal and Child Health Study (MACHS) were profiled by whole-genome bisulfite sequencing (median coverage: [Formula: see text]). Statistical analyses were conducted using the Regression Analysis of Differential Methylation (RADMeth) program because it performs well on low-coverage data (minimizes false positives and negatives).

Results: We found that 10,381 CpGs were differentially methylated by tobacco smoke exposure [neighbor-adjusted p-values that are additionally corrected for multiple testing based on the Benjamini-Hochberg method for controlling the false discovery rate (FDR) [Formula: see text]]. From these CpGs, RADMeth identified 557 differentially methylated regions (DMRs) that were overrepresented ([Formula: see text]) in important regulatory regions, including enhancers. Of nine DMRs that could be queried in a reduced representation bisulfite sequencing (RRBS) study of adult [Formula: see text] cells ([Formula: see text] smokers; [Formula: see text] nonsmokers), four replicated ([Formula: see text]). Additionally, a CpG in the promoter of SLC7A8 (percent methylation difference: [Formula: see text] comparing exposed to unexposed) replicated ([Formula: see text]) in an EPIC (Illumina) array study of cord blood [Formula: see text] cells ([Formula: see text] exposed to sustained maternal tobacco smoke; [Formula: see text] unexposed) and in a study of adult [Formula: see text] cells across two platforms (EPIC: [Formula: see text] smokers; [Formula: see text] nonsmokers; 450K: [Formula: see text] smokers; [Formula: see text] nonsmokers).

Conclusions: Maternal tobacco smoke exposure in pregnancy is associated with cord blood [Formula: see text] DNA methylation in key regulatory regions, including enhancers. While we used a method that performs well on low-coverage data, we cannot exclude the possibility that some results may be false positives. However, we identified a differentially methylated CpG in amino acid transporter SLC7A8 that is highly reproducible, which may be sensitive to cigarette smoke in both cord blood and adult [Formula: see text] cells. https://doi.org/10.1289/EHP3398.

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Figures

Figures 1a, 1b, 1c, and 1d plot the methylation levels (ranging between 0% and 100%) (y-axis) for exposed and unexposed groups of participants and the genomic positions for DMRs chr11:47952753-47952964, -7.8 percent; chr6:2764191-2764383, 11.1 percent; chr9:72027018-7202781, 17.8 percent; and chr19:47273675-47273807, 6.7 percent, respectively (x-axis).
Figure 1.
The four differentially methylated regions (DMRs) identified in the Maternal and Child Health Study (MACHS), which replicated in the National Institute of Environmental Health Sciences Clinical Research Unit (NIEHS CRU) reduced representation bisulfite sequencing study of adult CD4+ cells, are depicted in (A–D). DMRs were considered replicated if a) the chromosome coordinates overlapped between the two studies; b) the p-value for the NIEHS CRU study DMR was <0.05; and c) the percent methylation difference was in the same direction. Coordinates for each DMR and the percent methylation difference (comparing maternal tobacco smoke exposed with unexposed newborns) are indicated at the top of each panel. The location of each DMR is indicated by a vertical red line in the ideogram of its respective chromosome. Plots show the percent methylation levels (y-axis) for each MACHS participant at each CpG contained in the DMR. Chromosome positions for CpGs are indicated in gray (x-axis) above the plot. Percent methylation levels for participants in the exposed group are shown as light blue dots, and mean percent methylation levels for each CpG are connected by light blue lines. Percent methylation levels for participants in the unexposed group are shown as dark blue dots, and mean percent methylation levels for each CpG are connected by dark blue lines. Vertical gray bars highlight CpGs that were identified as differentially methylated with both a raw p<0.05 and a p<0.05 after accounting for correlations with neighboring CpGs and multiple testing, using the Benjamini-Hochberg method to control the false discovery rate.
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