Hypoxic storage of red blood cells improves metabolism and post-transfusion recovery

Abstract

Background: Blood transfusion is a lifesaving intervention for millions of recipients worldwide every year. Storing blood makes this possible but also promotes a series of alterations to the metabolism of the stored erythrocyte. It is unclear whether the metabolic storage lesion is correlated with clinically relevant outcomes and whether strategies aimed at improving the metabolic quality of stored units, such as hypoxic storage, ultimately improve performance in the transfused recipient.

Study design and methods: Twelve healthy donor volunteers were recruited in a two-arm cross-sectional study, in which each subject donated 2 units to be stored under standard (normoxic) or hypoxic conditions (Hemanext technology). End-of-storage measurements of hemolysis and autologous posttransfusion recovery (PTR) were correlated to metabolomics measurements at Days 0, 21, and 42.

Results: Hypoxic red blood cells (RBCs) showed superior PTR and comparable hemolysis to donor-paired standard units. Hypoxic storage improved energy and redox metabolism (glycolysis and 2,3-diphosphoglycerate), improved glutathione and methionine homeostasis, decreased purine oxidation and membrane lipid remodeling (free fatty acid levels, unsaturation and hydroxylation, acyl-carnitines). Intra- and extracellular metabolites in these pathways (including some dietary purines) showed significant correlations with PTR and hemolysis, though the degree of correlation was influenced by sulfur dioxide (SO2 ) levels.

Conclusion: Hypoxic storage improves energy and redox metabolism of stored RBCs, which results in improved posttransfusion recoveries in healthy autologous recipients-a Food and Drug Administration gold standard of stored blood quality. In addition, we identified candidate metabolic predictors of PTR for RBCs stored under standard and hypoxic conditions.

Conflict of interest statement

CONFLICT OF INTEREST

SN, SS, DS, FM, and NR have disclosed no conflicts of interest. ADA and TN are founders of Omix Technologies Inc and Altis Biosciences LLC. ADA and JAC are consultants for Hemanext Inc, with which ADu and TY are affiliated.

© 2020 AABB.

Figures

Fig. 1.

Hypoxic storage results in superior PTR and different metabolic reprogramming in stored RBCs. Overview of the experimental design. Twelve healthy donor volunteers donated 2 units of blood (two-arm, cross-sectional study) that were stored either under standard (normoxic) or hypoxic conditions for up to 42 days (A). At the end of storage, Food and Drug Administration gold standards for storage quality were tested for all 24 units, showing superior 24-hr posttransfusion recovery (PTR) (p

Fig. 2.

Overview of metabolites in central carbon and nitrogen metabolism affected by normoxic (red) or hypoxic (blue) storage at Storage Days 0, 21, and 42. Pantoth. = Pantothenic acid.

Fig. 3.

Circos plot of correlation networks among metabolites and PTR. Circos plots were generated for normoxic (top row) and hypoxic (bottom row) RBCs at Days 0, 21, and 42 (from left to right). Metabolites were connected by an edge if the module of the Spearman correlation between their relative levels or with end-of-storage PTR was higher than 65%. Edges in red highlight metabolites with significant correlation with end-of-storage PTR at each time point in each condition. For those instances in which >10 edges were identified, selected metabolites were graphed among the most significant ones for representative pathways.

Fig. 4.

Significant metabolic correlates to PTR differ in a storage-independent hypoxia-dependent fashion, as noted in the heat map on the left-hand side. Metabolites are color coded by metabolic pathway, according to the legend in the top right corner. Some metabolites showed significant (module of Spearman) correlation to PTR in normoxia, but not in hypoxia at any given storage day and vice versa (red in heat map). Some of the metabolites with high correlation with PTR in normoxia were only significant in hypoxic RBCs at the end of storage. All these metabolites were part of pathways significantly affected by SO2 levels of the unit. Indeed, the levels of these metabolites showed significant positive (bottom right panel in red) or negative (green) correlations to SO2 levels. Correlation curves are shown for glucose and lactate, two representative metabolites of glycolysis and the most differentially regulated pathway between normoxic and hypoxic RBCs.

Fig. 5.

Ranked Day 42 metabolic correlates to PTR measurements. PTR—in the tested units: all (A), normoxic (B) or hypoxic (C), and hemolysis (D). Highlights of representative correlation curves (E).