DNA Methylation of <i>SLC6A4</i> and <i>NR3C1</i> in Term and Preterm Infants: A Pilot Study of NICU-Associated Stress

<b>Background:</b> Preterm infants requiring prolonged hospitalization in the neonatal intensive care unit (NICU) are often exposed to early-life stress, which may contribute to long-term health consequences. Epigenetic markers, particularly DNA methylation (DNAm) of stress-related genes such as <i>SLC6A4</i> and <i>NR3C1</i>, have been identified as potential biomarkers of early-life stress. Limited research has compared DNAm patterns between term and preterm infants or examined associations between NICU-related stress and DNAm in preterm populations. <b>Objectives:</b> This pilot study aimed to (1) compare DNAm patterns between preterm and term infants, (2) examine relationships between NICU-related stress and DNAm alterations in preterm infants, and (3) explore DNAm changes over time in preterm infants. <b>Methods:</b> Buccal swabs were collected from 10 term and 23 preterm infants within the first week of life. Eleven preterm infants returned for a repeat sampling three months post discharge. The Neonatal Infant Stressor Scale (NISS) was used to calculate early-life stress. DNAm at specific CpG sites were analyzed using Mann-Whitney <i>U</i> tests and Spearman's rho correlations. <b>Results:</b> Preterm infants exhibited significantly different DNAm patterns at select CpG sites compared to term infants. Among preterm infants, higher NISS scores during hospitalization correlated with altered DNAm patterns in both genes. At three months post-discharge, new and persistent DNAm alterations were identified; although, total average DNAm did not significantly differ over time. <b>Conclusion:</b> These findings suggest that NICU-related stress is associated with DNAm alterations in preterm infants, supporting the need for further investigation of DNAm as a biomarker of early-life stress.