Stability of Blood DNA Methylation Across Two Timepoints in Three Cohorts

Background: DNA methylation mediates the gene-environment interactions, with implications for health and disease. Studies with sampling at more than one timepoint revealed the considerable variability of the blood methylome, but comprehensive resources on genome-wide methylation stability are still lacking. We aimed to identify methylation sites that remain the most stable across two timepoints in human whole blood. Methods: Publicly available blood DNA methylation data from three cohorts were analysed, which included methylation profiles at two timepoints >1 year apart. The cohorts included pre-/post-pubertal children (Illumina 450k array), the elderly (Illumina 450k array), and middle-aged adults with obesity (Illumina EPIC array). Two metrics were used for the stability assessment: the mean absolute difference (MAD) of beta values between two measurements and the intraclass correlation coefficient (ICC). We searched for probes demonstrating high stability (low MAD and high ICC) across the three cohorts. Data from 51 children, 86 elderly adults, and 120 middle-aged participants were re-analysed. Results: The median interquartile range (IQR) of the maximum (from three datasets) MAD was 2.1% (1.5-2.9%), and the median of the minimum ICC agreement coefficient was 0.053 (-0.077-0.304). The Pearson's correlation coefficient for the ICC vs. maximum MAD was low (r = 0.34, p < 2.2 × 10^-16). We found only 239 probes that were highly stable based on both the maximum MAD (<5th percentile, <0.01) and ICC criterion (>95th percentile, >0.74). Conclusions: The whole-blood DNA methylation profile, as measured using microarrays, is dynamic over >1 year, but contains a fraction of stable probes, most of which are related to genomic variation. A resource describing probe stability is made publicly available, with the intention to support biomarker studies and the investigation of early epigenetic programming. The absolute error and correlation are two complementary facets of probe stability that may be considered in further research, especially to determine the stability of probes in health and disease across different tissues and populations.