Ferroptosis regulates hemolysis in stored murine and human red blood cells

• QTL analyses identified >400 gene-metabolite associations in fresh and stored RBCs from 350 diversity outbred mice.

• Steap3 or ferroptosis-related genes FADS1/2, EPHX2, and LPCAT3 regulate lipid oxidation, PTR in mice, hemolysis, and Hgb increments in human RBCs.

Red blood cell (RBC) metabolism regulates hemolysis during aging in vivo and in the blood bank. However, the genetic underpinnings of RBC metabolic heterogeneity and extravascular hemolysis at population scale are incompletely understood. On the basis of the breeding of 8 founder strains with extreme genetic diversity, the Jackson Laboratory diversity outbred population can capture the impact of genetic heterogeneity in like manner to population-based studies. RBCs from 350 outbred mice, either fresh or stored for 7 days, were tested for posttransfusion recovery, as well as metabolomics and lipidomics analyses. Metabolite and lipid quantitative trait loci (QTL) mapped >400 gene-metabolite associations, which we collated into an online interactive portal. Relevant to RBC storage, we identified a QTL hotspot on chromosome 1, mapping on the region coding for the ferrireductase 6-transmembrane epithelial antigen of the prostate 3 (Steap3), a transcriptional target to p53. Steap3 regulated posttransfusion recovery, contributing to a ferroptosis-like process of lipid peroxidation, as validated via genetic manipulation in mice. Translational validation of murine findings in humans, STEAP3 polymorphisms were associated with RBC iron content, lipid peroxidation, and in vitro hemolysis in 13 091 blood donors from the Recipient Epidemiology and Donor Evaluation Study. QTL analyses in humans identified a network of gene products (fatty acid desaturases 1 and 2, epoxide hydrolase 2, lysophosphatidylcholine acetyl-transferase 3, solute carrier family 22 member 16, glucose 6-phosphate dehydrogenase, very long chain fatty acid elongase, and phospholipase A2 group VI) associated with altered levels of oxylipins. These polymorphisms were prevalent in donors of African descent and were linked to allele frequency of hemolysis-linked polymorphisms for Steap3 or p53. These genetic variants were also associated with lower hemoglobin increments in thousands of single-unit transfusion recipients from the vein-to-vein database.