DNA methylation in relation to gestational age and brain dysmaturation in preterm infants

Abstract

Preterm birth is associated with dysconnectivity of structural brain networks and is a leading cause of neurocognitive impairment in childhood. Variation in DNA methylation is associated with early exposure to extrauterine life but there has been little research exploring its relationship with brain development. Using genome-wide DNA methylation data from the saliva of 258 neonates, we investigated the impact of gestational age on the methylome and performed functional analysis to identify enriched gene sets from probes that contributed to differentially methylated probes or regions. We tested the hypothesis that variation in DNA methylation could underpin the association between low gestational age at birth and atypical brain development by linking differentially methylated probes with measures of white matter connectivity derived from diffusion MRI metrics: peak width skeletonized mean diffusivity, peak width skeletonized fractional anisotropy and peak width skeletonized neurite density index. Gestational age at birth was associated with widespread differential methylation at term equivalent age, with genome-wide significant associations observed for 8870 CpG probes (P < 3.6 × 10^-8) and 1767 differentially methylated regions. Functional analysis identified 14 enriched gene ontology terms pertaining to cell-cell contacts and cell-extracellular matrix contacts. Principal component analysis of probes with genome-wide significance revealed a first principal component that explained 23.5% of the variance in DNA methylation, and this was negatively associated with gestational age at birth. The first principal component was associated with peak width of skeletonized mean diffusivity (β = 0.349, P = 8.37 × 10^-10) and peak width skeletonized neurite density index (β = 0.364, P = 4.15 × 10^-5), but not with peak width skeletonized fraction anisotropy (β = -0.035, P = 0.510); these relationships mirrored the imaging metrics' associations with gestational age at birth. Low gestational age at birth has a profound and widely distributed effect on the neonatal saliva methylome that is apparent at term equivalent age. Enriched gene ontology terms related to cell-cell contacts reveal pathways that could mediate the effect of early life environmental exposures on development. Finally, associations between differential DNA methylation and image markers of white matter tract microstructure suggest that variation in DNA methylation may provide a link between preterm birth and the dysconnectivity of developing brain networks that characterizes atypical brain development in preterm infants.

Keywords: DNA methylation; MRI; brain; development; neonate.

Graphical Abstract

Figure 1

Manhattan plot for the significance [−log10 (P-value) used for visualization purposes] of the association between gestational age at birth (weeks) and DNA methylation, following adjustment for covariates and surrogate variables. The solid horizontal line shows the genome-wide significance level and red dots above this line represent probes that are significant at this threshold (P < 3.6 × 10⁻⁸).

Figure 2

Scatter plots with regression lines and 95% confidence intervals showing the relationships between gestational age at birth (weeks) and DNAm with PSMD and PSNDI, where peak width is the difference between the 95th and 5th centile of histogram values across the white matter skeleton. The associations between gestational age (weeks) and PSMD and PSNDI are shown in (A) and (B), respectively. The relationships between DNAm PC1 and PSMD and PSNDI are shown in (C) and (D), respectively. PS metrics are residualized for gestational age at scan; PSMD is additionally residualized for scanner variable.

Figure 3

Variation in DNAm probes selected by EWAS captured by principal component analysis, and the relationship with gestational age (weeks). A. A scree plot showing percentage of variance accounted for by the first 10 components, with a sharp elbow after the first PC, accounting for 23.5% of variance. B. A scatter plot, with regression line, showing the relationship between gestational age at birth (weeks) and PC1 (r = −0.622) with 95% confidence intervals.