Donor blood saves lives, yet the potential impact of recurrent large-volume phlebotomy on donor health and hematopoietic stem cells (HSCs) remains largely unexplored.

In our study, we conducted a comprehensive screening of 217 older male volunteer donors with a history of extensive blood donation (> 100 life-time donations) to investigate the phenomenon of clonal hematopoiesis (CH).

No significant difference in the overall incidence of CH was found in frequent donors (FD) compared to sporadic donors (< 10 life-time donations, 212 donors). However, upon deeper analysis of mutations in DNMT3A, the most commonly affected gene in CH, we observed distinct mutational patterns between the FD and control donor (CD) cohorts. Functional analysis of FD enriched DNMT3A variants examined in CRISPR-edited human HSCs demonstrated their competitive outgrowth potential upon erythropoietin (EPO) stimulation, whereas leukemogenic DNMT3A R882 mutations exhibited preferential responsiveness to inflammatory cues. Concurrent mutational and immunophenotypic single-cell profiling revealed no lineage bias in the contribution of FD DNMT3A variant mutant cells in primary homeostatic donor samples. Conversely, they promote erythroid biased differentiation in xenografts stressed by serial blood loss and EPO treatment. This contrasts with R882 mutations, which induce a pronounced myeloid bias during both homeostasis and erythroid stress.

Our data demonstrate a nuanced ongoing Darwinian evolution at the somatic stem cell level, with EPO identified as a novel environmental factor that favors HSCs carrying certain DNMT3A mutations. Crucially, our findings robustly demonstrate that frequent blood donation over many years does not promote the selection of malignant clones, thus strongly reaffirming its safety.

No conflict of interest to disclose