Differential DNA Methylation of the Serotonin Receptor Signaling and Glutamatergic Synapse Pathways in Adult Twins Born Preterm.

Early-life environment may influence long-term neurodevelopment through epigenetic regulation. Serotonergic and glutamatergic pathways are central to brain development and have been implicated in DNA methylation changes following prenatal adversity. In this study, we examined whether preterm birth (PTB) in birthweight-discordant twins is associated with differential DNA methylation in the serotonin receptor signaling pathway and the glutamatergic synapse pathway in adult twins. Genome-wide DNA methylation data were obtained from whole blood samples of 288 individuals (144 monozygotic birthweight-discordant twin pairs), including a younger cohort (140 individuals; mean age 33 years) and an older cohort (148 individuals; mean age 63 years). DNA methylation was measured using the Illumina HumanMethylation450 BeadChip. Linear models were fitted for association testing, adjusting for leukocyte composition and twin pair correlation. Pathway-level differential methylation was assessed using Rotation Gene Set Testing. In the glutamatergic synapse pathway, no consistent directional enrichment of hypo- or hypermethylation was observed. However, gene-level analyses identified consistent hypomethylation of GRIA2 and GRIA4 across cohorts. In the serotonin receptor signaling pathway, the young cohort exhibited a mixed methylation pattern, whereas the old cohort showed significant enrichment of hypermethylation. At the gene level, HTR1A was hypomethylated in the young cohort but hypermethylated in the old cohort, indicating a cohort-dependent effect in the methylation patterns. These findings suggest that PTB is associated with long-term epigenetic variation in neurodevelopmentally relevant pathways, as reflected in blood cells. The results further indicate distinct methylation architectures across pathways, with more consistent pathway-level signals in the serotonergic system and more localized gene-level effects in the glutamatergic pathway.