Fig. 1: Analyzing the neuronal methylome and hydroxymethylome in OUD.

a Experimental workflow. Fluorescence-activated nuclei sorting was used to isolate neuronal nuclei from postmortem OFC. Nuclei were processed for genomic DNA, undergoing reduced representation oxidative bisulfite sequencing to examine 5mC and 5hmC at CpG-dense loci. Figure created with BioRender.com. b An average of 10x coverage was obtained for ~3.5 million CpG sites. c 44% of CpG sites were in promoter regions and 30% in intergenic regions. d, e Neuronal 5mC occurs mainly in intergenic regions, while 5hmC occurs in introns, exons, and intergenic regions (n = 38 biologically independent samples; 26 OUD+; 12 OUD−). Data are presented as mean values ± SD, and p value were generated using ANOVA. f Contrasting mean 5mC levels at neuronal (NeuN+) vs. non-neuronal (NeuN−) marker genes.