Plasma testosterone heterogeneity reveals genetic and demographic determinants of donor health and red blood cell quality

Testosterone is a key determinant of red blood cell (RBC) biology and susceptibility to hemolysis; however, its contribution to donor-dependent variation in RBC quality and transfusion effectiveness remains unclear. Using data from 2,357 male participants in the NHLBI REDS-III RBC-Omics study, we quantified plasma total testosterone, free testosterone (bioavailable form), and androstenedione and evaluated their associations with donor characteristics, RBC hemolysis, and recipient post-transfusion hemoglobin increments, a clinical measure of RBC transfusion effectiveness. Subphysiological testosterone concentrations were common (~35%) and independently associated with increased storage and osmotic hemolysis. Frequent blood donations were independently associated with lower circulating free testosterone concentrations, even after adjustment for demographic and hematologic factors. Genome-wide association analyses identified loci associated with circulating testosterone, including regions near SHBG, TNFSF12, and the long noncoding RNA BALR6. Several testosterone variants were associated with RBC transfusion effectiveness. Specifically, variants associated with lower testosterone (e.g., TNFSF12) corresponded to higher post-transfusion hemoglobin increments (+9%), whereas BALR6 variants associated with higher testosterone were linked to reduced hemoglobin increments (-40%). In contrast, androstenedione-associated variants were not related to hemolysis or transfusion outcomes. Together, these findings demonstrate that genetic and physiologic variation in testosterone contributes to inter-individual variability in RBC quality and transfusion effectiveness.