Metabolic reprogramming under hypoxic storage preserves faster oxygen unloading from stored red blood cells

Abstract

Stored red blood cells (RBCs) incur biochemical and morphological changes, collectively termed the storage lesion. Functionally, the storage lesion manifests as slower oxygen unloading from RBCs, which may compromise the efficacy of transfusions where the clinical imperative is to rapidly boost oxygen delivery to tissues. Recent analysis of large real-world data linked longer storage with increased recipient mortality. Biochemical rejuvenation with a formulation of adenosine, inosine, and pyruvate can restore gas-handling properties, but its implementation is impractical for most clinical scenarios. We tested whether storage under hypoxia, previously shown to slow biochemical degradation, also preserves gas-handling properties of RBCs. A microfluidic chamber, designed to rapidly switch between oxygenated and anoxic superfusates, was used for single-cell oxygen saturation imaging on samples stored for up to 49 days. Aliquots were also analyzed flow-cytometrically for side-scatter (a proposed proxy of O2 unloading kinetics), metabolomics, lipidomics and redox proteomics. For benchmarking, units were biochemically rejuvenated at four weeks of standard storage. Hypoxic storage hastened O2 unloading in units stored to 35 days, an effect that correlated with side-scatter but was not linked to post-translational modifications of hemoglobin. Although hypoxic storage and rejuvenation produced distinct biochemical changes, a subset of metabolites including pyruvate, sedoheptulose 1-phosphate, and 2/3 phospho-D-glycerate, was a common signature that correlated with changes in O2 unloading. Correlations between gas-handling and lipidomic changes were modest. Thus, hypoxic storage of RBCs preserves key metabolic pathways and O2 exchange properties, thereby improving the functional quality of blood products and potentially influencing transfusion outcomes.

Biopreservation and Reversal of Oxidative Injury During Blood Storage by a Novel Curcumin-based Gel Formulation

“…The most common intervention strategies designed to control the biochemical consequences of RBC age-related changes include the targeting of (i) the source(s) of ROS by subjecting RBCs to periods of hypoxia in hypoxic chambers 33 , (ii) Hb oxidation with reductants such as ascorbic acid or caffeic acid 10,[34][35][36] and (iii) RBC membranes, speci cally band 3 complex proteins 37 . Individual or any combination of these methods may minimize biochemical changes and restore oxygen homeostasis in tissues.…”

Impaired O2 unloading from stored blood results in diffusion-limited O2 release at tissues: evidence from human kidneys

“…Rejuvenation before blood product use is labor intensive, and we estimate a burden of 3 to 4 hours that may limit its use in settings, such as major hemorrhage. Although there is capacity to improve the formulation using τ as a performance readout, and the possibility of rejuvenating blood before issue, future studies should evaluate alternative storage regimes (eg, hypoxic storage 26 ), the addition of phosphate and guanosine, or adjustments in pH. This may be particularly important in circumstances where the transfusion-load is a substantial fraction of the recipient’s blood volume or when blood is given to alleviate critical ischemia.…”

“…The kinetics of oxygen unloading from RBCs were studied using a modification of a previously published method. 10 , 26 …”

Erythrocyte metabolism

“…For instance, rejuvenation of erythrocytes during blood storage (using pyruvate, inosine, phosphate, and adenine) increased the levels of 2,3‐BGP and ATP similar to or even above the levels of fresh cells, probably by enhancing glycolytic flux 401 . Noteworthy, rejuvenated erythrocytes exhibited higher oxygen‐carrying capacity and faster oxygen‐unloading kinetics, reversing the adverse effects of blood storage 364,402 …”