Abstract: Aging is associated with many complex diseases such as cancer and neurodegenerative diseases. Recently, many age-related DNA methylation biomarkers in peripheral whole blood have been identified. These biomarkers may reflect DNA methylation changes derived from changes in the number of a specific leukocyte cell type during aging. To clarify the source of these age-related DNA methylation changes, we analysed DNA methylation profile of peripheral whole blood from three independent cohorts of healthy subjects and identified age-related DNA methylation CpG sites (arCpGs) using the Spearman’s rank test with high reproducibility (Hypergeometric test, P=1.65×10^-11). Using a deconvolution algorithm, we found that the proportion of myeloid lineage cells was increased while that of lymphoid lineage cells was decreased in the peripheral whole blood with age (Spearman’s rank correlation test, P<0.05, r≤0.22). The CpG sites, whose methylation levels were significantly different in myeloid cells and lymphoid cells, were preferentially recognized as arCpGs in peripheral whole blood. Moreover, the arCpGs in CD4+ T cells significantly overlapped with that in peripheral whole blood (Hypergeometric test, P=6.14×10^-12) and 99.1% of the overlapping arCpGs had consistent positive or negative correlations with age. Though the arCpGs in CD14+ monocytes did not significantly overlap with that in peripheral whole blood (Hypergeometric test, P=0.232), 90.1% of 51 overlapping arCpGs were correlated with age in CD14+ monocytes, peripheral whole blood, and CD4+ T cells consistently. In summary, most of the methylation changes in arCpGs identified in peripheral whole blood come from common or specific DNA methylation changes in leukocyte subtypes, while part of them reflect alterations in the number of specific cell types of leukocytes.

Key words: peripheral whole blood, aging, DNA methylation, CD4+ T cells, CD14+ monocytes