Presence of an epigenetic signature of prenatal cigarette smoke exposure in childhood

Highlights

• An epigenetic signature of prenatal exposure to cigarette smoke, originally discovered in other studies of cord blood at birth, is detectable in childhood blood samples at ages 3–5.

• Prenatal smoking-associated loci do not show similar patterns or magnitudes of DNA methylation differences for other prenatal exposures.

• DNA methylation patterns from 26 CpG sites at 3–5 years of age can accurately classify prenatal exposure to smoking.

Abstract

Prenatal exposure to tobacco smoke has lifelong health consequences. Epigenetic signatures such as differences in DNA methylation (DNAm) may be a biomarker of exposure and, further, might have functional significance for how in utero tobacco exposure may influence disease risk. Differences in infant DNAm associated with maternal smoking during pregnancy have been identified. Here we assessed whether these infant DNAm patterns are detectible in early childhood, whether they are specific to smoking, and whether childhood DNAm can classify prenatal smoke exposure status. Using the Infinium 450 K array, we measured methylation at 26 CpG loci that were previously associated with prenatal smoking in infant cord blood from 572 children, aged 3–5, with differing prenatal exposure to cigarette smoke in the Study to Explore Early Development (SEED). Striking concordance was found between the pattern of prenatal smoking associated DNAm among preschool aged children in SEED and those observed at birth in other studies. These DNAm changes appear to be tobacco-specific. Support vector machine classification models and 10-fold cross-validation were applied to show classification accuracy for childhood DNAm at these 26 sites as a biomarker of prenatal smoking exposure. Classification models showed prenatal exposure to smoking can be assigned with 81% accuracy using childhood DNAm patterns at these 26 loci. These findings support the potential for blood-derived DNAm measurements to serve as biomarkers for prenatal exposure.

Introduction

A considerable proportion (11%) of women in the United States actively smoke during pregnancy, a major risk factor for pregnancy complications(Castles et al., 1999; Shah and Bracken, 2000) and adverse health outcomes during infancy, childhood and later life(Salmasi et al., 2010; Shah and Bracken, 2000). Understanding the impact of early life exposure to tobacco smoke on future health has important public health implications.

DNA methylation (DNAm) is a type of epigenetic modification central to development and gene regulation. It is of interest as a mediating mechanism in exposure-disease associations, and may also have utility as a biological marker of exposure, even if not mechanistically implicated (Ladd-Acosta, 2015). Several groups have investigated associations between DNAm levels and in utero exposure to tobacco smoke. Using global (Guerrero-Preston et al., 2010), candidate gene-based (Murphy et al., 2012; Suter et al., 2010), and genome-scale (Joubert et al., 2014, 2012; Richmond et al., 2014; Suter et al., 2011) approaches, they identified associations between maternal smoking during pregnancy and DNAm levels in placental tissue and in DNA from cord blood (Lee and Pausova, 2013; Richmond et al., 2014). A recent study, using a low-density DNAm array showed detectible prenatal smoking associations in childhood, but could not assess the reported birth sample associations now confirmed by several groups, given the incompatible array content (Breton et al., 2014). A candidate gene-based study (Novakovic et al., 2014) of 11 individuals showed that comparable differences in DNAm at AHRR, a tobacco-related gene, were detectable at birth as well as at 18 months. Finally, a recent longitudinal investigation revealed some smoking-related DNAm alterations, initially detected in their sample at birth, persist within the same individuals over time (Richmond et al., 2014). While a few of the loci identified by that paper overlap with previous studies, the study did not specifically examine the set of 26 loci (Joubert et al., 2012) that have now been well replicated in other birth samples.

Here we attempted to replicate prenatal smoking-associated DNAm differences observed in infant cord blood, reported by Joubert et al. (2012)), in an independent set of 572 early childhood blood samples to determine if the DNAm pattern in childhood is consistent with DNAm “signatures” of prenatal smoking detected at birth. This study, focused on prenatal smoking, assesses the potential utility of a DNAm signature measured later in life as an epigenetic biomarker of prenatal exposure. This study also examines other issues relevant to DNAm's potential as a biomarker for prenatal smoke exposure. First, since it is possible that the DNAm changes previously reported in cord blood could be related to downstream responses to a range of prenatal exposures and, thus, are not tobacco smoke-specific, we explored associations of DNAm changes with other prenatal exposures, including maternal alcohol and medication use. Second, we evaluated associations between the previously reported DNAm changes and trimester-specific and sustained prenatal smoke exposure. Finally, we used machine learning and 10-fold cross-validation to assess whether childhood DNAm levels at these 26 sites can predict prenatal exposure to smoking.