Associations between Maternal Tobacco Smoke Exposure and the Cord Blood CD 4 + DNA Methylome

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.

The current study examined the impact of in utero exposure to maternal tobacco smoke on the cord blood $\text{CD}{4}^{+}$ DNA methylome.

The methylomes of 20 Hispanic white newborns ($n=10$ exposed to any maternal tobacco smoke in pregnancy; $n=10$ unexposed) from the Maternal and Child Health Study (MACHS) were profiled by whole-genome bisulfite sequencing (median coverage: $6.5\times$). 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).

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) $\left(p_{FDR}\right)<0.05$]. From these CpGs, RADMeth identified 557 differentially methylated regions (DMRs) that were overrepresented ($p<0.05$) in important regulatory regions, including enhancers. Of nine DMRs that could be queried in a reduced representation bisulfite sequencing (RRBS) study of adult $\text{CD}{4}^{+}$ cells ($n=9$ smokers; $n=10$ nonsmokers), four replicated ($p<0.05$). Additionally, a CpG in the promoter of SLC7A8 (percent methylation difference: $-9.4\%$ comparing exposed to unexposed) replicated ($p<0.05$) in an EPIC (Illumina) array study of cord blood $\text{CD}{4}^{+}$ cells ($n=14$ exposed to sustained maternal tobacco smoke; $n=16$ unexposed) and in a study of adult $\text{CD}{4}^{+}$ cells across two platforms (EPIC: $n=9$ smokers; $n=11$ nonsmokers; 450K: $n=59$ smokers; $n=72$ nonsmokers).

Maternal tobacco smoke exposure in pregnancy is associated with cord blood $\text{CD}{4}^{+}$ 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 $\text{CD}{4}^{+}$ cells. https://doi.org/10.1289/EHP3398.