The global clonal complexity of the hematopoietic system declines with age. (A) Schematic of experimental approach. The clonal complexity of the PB and BM of cohorts of Conf-E2a^Cre, Conf-VE^Cre, Conf-Flk1^Cre, and Conf-Vav1^Cre mice were examined at 2, 7, 12, 16, 20, 24, and 26 months of age (see supplemental Figure 1A-B for schematic of Confetti allele and Confetti color flow cytometry gating strategy). (Bi-ii) Schematic of the inverse relationship between numbers of initially labeled events and MtMV in the distribution of Confetti colors. (Ci) Analysis of the clonal complexity of the PB in cohorts of mice from 2 to 26 months of age. At 2 months old: Conf-E2a^Cre (n = 14), Conf-VE^Cre (n = 13), Conf-Flk1^Cre (n = 7), and Conf-Vav1^Cre (n = 11). At 26 months old, Conf-E2a^Cre (n = 10), Conf-VE^Cre (n = 5), Conf-Flk1^Cre (n = 6), and Conf-Vav1^Cre (n = 9). (Cii) Average PB clonal complexity of Conf-Flk1^Cre, Conf-VE^Cre, and Conf-Vav1^Cre mice over time relative to 2 months of age. Error bars indicated standard deviation. (Di) The clonal complexity of the major BM HSPCs from cohorts of mice were calculated at 2 and 26 months of age from previously cKit⁺-enriched BM. At 2 months, Conf-E2a^Cre (n = 3), Conf-VE^Cre (n = 6), Conf-Flk1^Cre (n = 7), and Conf-Vav1^Cre (n = 8). At 26 months, Conf-E2a^Cre (n = 7), Conf-VE^Cre (n = 4), Conf-Flk1^Cre (n = 5), and Conf-Vav1^Cre (n = 5). (Dii) Average BM HSPC clonal complexities of Conf-Flk1^Cre, Conf-VE^Cre, and Conf-Vav1^Cre mice over time relative to 2 months of age. Error bars indicate standard deviation (*P < .05; #P < .1). Source data are provided in supplemental Table 1. (Diii) Schematic of the consequences of aging on HSCs and PB clonal complexity. The absolute number of phenotypic HSCs increases with age (supplemental Figure 6A) due to the expansion of functionally impaired clones. In aged mice, “young-like” minimally expanded HSCs contribute disproportionately to PB, resulting in a less dramatic decrease in PB clonal complexity.