Prenatal metal exposure moderates the effect of CRH-related DNA methylation in the placenta on infant communication outcomes at 2 months of age

BACKGROUND/ AIM: Prenatal stress and environmental metal mixtures impact neurobehavioral development, yet their interplay and potential mechanisms via placental epigenetics remain unclear. We investigated whether prenatal metal mixtures moderate the epigenetic pathway linking maternal stress at delivery (corticotropin-releasing hormone, CRH) to infant neurobehavioral development. METHODS: In a prospective cohort of 120 mother-infant pairs, we measured maternal plasma CRH at delivery, placental DNA methylation (Infinium EPIC v2.0), and placental concentrations of 18 metals (ICP-MS). Infant neurobehavioral development was evaluated at 2 months using the Ages & Stages Questionnaires, Third Edition. We performed causal mediation analysis to identify mediating differentially methylated regions (DMRs). We then tested for effect moderation by the metal mixtures, employing a weighted quantile sum (WQS) index to assess the simultaneous effect of the 18-metal mixture on neurobehavioral development, followed by sex-stratified analyses. RESULTS: Placental DMRs annotated to GNAS, ZMYND10, and CLIP4 significantly mediated the association between maternal CRH and infant neurobehavioral development across five epigenetic pathways (ACME p-value < 0.05). The pathway linking CRH to communication via CLIP4 was significantly moderated by metal mixtures (p for interaction = 0.048), driven by sodium (18.3%), potassium (15.2%), mercury (11.9%), and copper (11.5%). The adverse association between higher CLIP4 methylation and lower communication scores was strong and significant at low levels but weakened with increasing exposure. Sex-stratified analyses revealed that this moderating effect was significant in male infants (p for interaction = 0.030) but not in female infants (p for interaction = 0.734). CONCLUSION: Our study suggests that prenatal metal mixtures moderate the placental epigenetic pathway linking maternal stress to neurobehavioral development in a sex-dependent manner. This underscores the necessity of a multi-exposure framework to understand the developmental origins of health and disease.